bromochloroacetic-acid and Cell-Transformation--Neoplastic

Keratins, the epithelial-predominant members of the intermediate filament superfamily, are expressed in a pairwise, tissuespecific and differentiation-dependent manner. There are 28 type I and 26 type II keratins, which share a common structure comprising a central coiled coil α-helical rod domain flanked by two nonhelical head and tail domains. These domains harbor sites for major posttranslational modifications like phosphorylation and glycosylation, which govern keratin function and dynamics. Apart from providing structural support, keratins regulate various signaling machinery involved in cell growth, motility, apoptosis etc. However, tissue-specific functions of keratins in relation to cell proliferation and differentiation are still emerging. Altered keratin expression pattern during and after malignant transformation is reported to modulate different signaling pathways involved in tumor progression in a context-dependent fashion. The current review focuses on the literature related to the role of keratins in the regulation of cell proliferation, differentiation and transformation in different types of epithelia.

Topics: Acetylation; Animals; Apoptosis; Carcinoma, Squamous Cell; Cell Differentiation; Cell Proliferation; Cell Transformation, Neoplastic; Epithelial Cells; Gene Expression Regulation, Neoplastic; Glycosylation; Humans; Keratins; Neoplasms; Phosphorylation; Protein Processing, Post-Translational; Protein Structure, Secondary; Signal Transduction

Two cases of adenomyoepithelioma of the breast with malignant transformation by monophasic population of cells are presented. The underlying benign adenomyoepithelioma with typical biphasic architectural pattern was identified and represented at least 30% of the tumor in each case. In both cases, malignant portion of tumor was composed of relatively uniform monophasic population of highly atypical cells. The malignant component in case 1 was positive for pan cytokeratin, myoepithelial markers, and basal-type cytokeratins and also focally positive for luminal-type of cytokeratins, but negative for hormone receptors (estrogen and progesterone) and HER-2/neu protein overexpression. The malignant component in case 2 was positive for spectrum of myoepithelial markers but negative for luminal cytokeratins, hormone receptors and HER-2/neu protein overexpression. The bilinear immunophenotype in the case 1 suggests that the malignant tumor may have developed from precursor multipotent cells that can differentiate into both luminal epithelial and myoepithelial cells, although malignant component in case 2 appears to be the of pure myoepithelial phenotype.

Topics: Adenomyoepithelioma; Adult; Breast Neoplasms; Cell Transformation, Neoplastic; Female; Humans; Keratins; Middle Aged

Keratins are the intermediate filament (IF)-forming proteins of epithelial cells. Since their initial characterization almost 30 years ago, the total number of mammalian keratins has increased to 54, including 28 type I and 26 type II keratins. Keratins are obligate heteropolymers and, similarly to other IFs, they contain a dimeric central α-helical rod domain that is flanked by non-helical head and tail domains. The 10-nm keratin filaments participate in the formation of a proteinaceous structural framework within the cellular cytoplasm and, as such, serve an important role in epithelial cell protection from mechanical and non-mechanical stressors, a property extensively substantiated by the discovery of human keratin mutations predisposing to tissue-specific injury and by studies in keratin knockout and transgenic mice. More recently, keratins have also been recognized as regulators of other cellular properties and functions, including apico-basal polarization, motility, cell size, protein synthesis and membrane traffic and signaling. In cancer, keratins are extensively used as diagnostic tumor markers, as epithelial malignancies largely maintain the specific keratin patterns associated with their respective cells of origin, and, in many occasions, full-length or cleaved keratin expression (or lack there of) in tumors and/or peripheral blood carries prognostic significance for cancer patients. Quite intriguingly, several studies have provided evidence for active keratin involvement in cancer cell invasion and metastasis, as well as in treatment responsiveness, and have set the foundation for further exploration of the role of keratins as multifunctional regulators of epithelial tumorigenesis.

Topics: Adenocarcinoma; Animals; Biomarkers, Tumor; Cell Transformation, Neoplastic; Cytoplasm; Epithelial Cells; Female; Health; Humans; Intermediate Filaments; Keratins; Male; Mice; Protein Biosynthesis; Protein Interaction Domains and Motifs; Protein Processing, Post-Translational

Topics: Biomarkers, Tumor; Breast Neoplasms; Cell Transformation, Neoplastic; Female; Humans; Immunophenotyping; Keratins

In the last 10 years, there has been a relative explosion of new rodent systems that recapitulate both genetic and cellular lesions that lead to the development of pancreatic cancer. These models now need to be considered when selecting an appropriate in vivo system to study disease etiology, cell signaling, and drug development. The majority of these evaluations have used transplantation of cancer cells and the use of carcinogens, which still maintain their value when investigating human cancer and epigenetic contributors. Xenograft models utilize cultured or primary pancreatic cancer cells that are placed under the skin or implanted within the pancreas of immunocompromised mice. Carcinogen-induced systems rely on administration of certain chemicals to generate cellular changes that rapidly lead to pancreatic cancer. Genetically modified mice are more advanced in their design in that relevant genetic mutations can be inserted into mouse genomic DNA in both a conditional and inducible manner. Generation of mice that develop spontaneous pancreatic cancer from a targeted genetic mutation is a valuable research tool, considering the broad spectrum of genes and cell targets that can be used, producing a variety of neoplastic lesions and cancer that can reflect many aspects of human pancreatic ductal adenocarcinoma.

Topics: Animals; Carcinogens; Cell Transformation, Neoplastic; Disease Models, Animal; Homeodomain Proteins; Humans; Keratins; Mice; Mice, Knockout; Mice, Transgenic; Pancreatic Elastase; Pancreatic Neoplasms; Trans-Activators; Transcription Factors; Xenograft Model Antitumor Assays

The cDNA microarrays allows the classification of breast cancers into 6 groups: luminal A, luminal B, luminal C, normal breast-like, human epidermal growth factor receptor 2-positive, and basal-like. This latter is characterized by the expression of basal cytokeratins (CKs), and frequent negativity for hormone receptors and human epidermal growth factor receptor 2. There is a marked parallelism between triple negative breast carcinomas and basal-like carcinoma, but these are not equivalent terms. Estimated concordance is around 80%. CK5 seems to be the best marker for the identification of these tumors. Other good markers to identify these tumors are CK14, CK17, and epidermal growth factor receptor. A subset of triple negative breast carcinomas has myoepithelial differentiation, with positivities for smooth muscle actin, p63, S-100, and CD10 among others. Recent studies suggest that basal like carcinomas are originated from mammary stem cells.

Topics: Angiogenesis Inhibitors; Animals; Biomarkers, Tumor; Breast Neoplasms; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Diagnosis, Differential; ErbB Receptors; Female; Humans; Incidence; Keratins; Mammary Glands, Human; Myeloid Progenitor Cells; Receptors, Cell Surface

Significant progress in the diagnosis and therapy of salivary gland diseases has been made in recent years. The new technique of diagnostic and interventional sialendoscopy has made an important contribution and is indicated in every case of obstructive sialadenitis. The number of open resections of salivary glands due to stones will clearly decrease in the future in favor of endoscopic removal. Due to recent publications on the appropriate extent of salivary gland resection in benign tumors, more and more specimens with reduced cuffs of healthy salivary gland tissue will be sent to the pathologists. Ultrasound will stay the procedure of first choice for imaging of salivary gland diseases in Germany. In combination with fine-needle aspiration cytology high sensitivity and specificity for the assessment of salivary gland tumors can be achieved. Diffusion-weighted magnetic resonance imaging (MRI) is a new imaging tool and the power of distinction of pleomorphic adenoma from malignant tumors is promising. The use of botulinum toxin for salivary glands diseases is increasing. Intraglandular injections have been shown to induce salivary gland atrophy in animal experiments. The availability of biologicals is currently yielding new aspects for the treatment of Sjögren's disease.

Topics: AIDS-Related Opportunistic Infections; Biomarkers, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Cysts; Diagnosis, Differential; Epithelium; Humans; Keratins; Lymph Nodes; Lymphoma, B-Cell, Marginal Zone; Parotid Diseases; Salivary Ducts; Salivary Gland Neoplasms; Salivary Glands; Sialadenitis; Sjogren's Syndrome

Miscellaneous primary tumors of the uterine cervix are rare. Markers which can be utilized to detect these tumors are very few and in most cases, have not been clinically validated. The information provided in this article will help in developing strategies to discover novel markers and initiate translational research in this ignored area. Based on the reported studies, cytokeratin markers are common in many tumors and few of these rare cancers demonstrate human papilloma-virus (HPV) and Epstein Bar virus (EBV) infection. Due to the very low prevalence of these tumors, epidemiological studies have not been conducted and the etiology of these tumors is largely unknown.

Topics: Biomarkers, Tumor; Carcinoma; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Lipoma; Melanoma; Neurilemmoma; Rare Diseases; Sarcoma; Uterine Cervical Neoplasms

The diversity of epithelial functions is reflected by the expression of distinct keratin pairs that are responsible to protect epithelial cells against mechanical stress and to act as signaling platforms. The keratin cytoskeleton integrates these functions by forming a supracellular scaffold that connects at desmosomal cell-cell adhesions. Multiple human diseases and murine knockouts in which the integrity of this system is destroyed testify to its importance as a mechanical stabilizer in certain epithelia. Yet, surprisingly little is known about the precise mechanisms responsible for assembly and disease pathology. In addition to these structural aspects of keratin function, experimental evidence accumulating in recent years has led to a much more complex view of the keratin cytoskeleton. Distinct keratins emerge as highly dynamic scaffolds in different settings and contribute to cell size determination, translation control, proliferation, cell type-specific organelle transport, malignant transformation and various stress responses. All of these properties are controlled by highly complex patterns of phosphorylation and molecular associations.

Topics: Animals; Cell Polarity; Cell Transformation, Neoplastic; Cytoskeleton; Desmosomes; Epithelial Cells; Humans; Intermediate Filaments; Keratins; Signal Transduction; Stress, Mechanical

Primary intracranial squamous cell carcinoma is extremely rare, with most of the cases arising from malignant transformation of an epidermoid or a dermoid cyst. We report here a case of a 45-year-old male patient who presented with 1-month history of intermittent headache and recent onset of altered sensorium. Imaging revealed a midline posterior fossa mass lesion compressing the fourth ventricle and causing hydrocephalus. A provisional diagnosis of dermoid cyst was considered. Histopathological examination revealed a squamous cell carcinoma possibly arising from an underlying epidermoid cyst. This entity is being reported for its rarity.

Topics: Brain Diseases; Calcinosis; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cholesterol; Cranial Fossa, Posterior; Diagnosis, Differential; Epidermal Cyst; Fourth Ventricle; Humans; Keratins; Male; Middle Aged; Skull Base Neoplasms; Tomography, X-Ray Computed

Intermediate filaments (IF) form the structural framework of the cytoskeleton. Although histopathological detection of IF proteins is utilized for examining cancer specimens as reliable markers, the molecular mechanisms by which IF are involved in the biology of cancer cells are still unclear. We found that site-specific phosphorylation of IF proteins induces the disassembly of filament structures. To further dissect the in vivo spatiotemporal dynamics of IF phosphorylation, we developed site- and phosphorylation state-specific antibodies. Using these antibodies, we detected kinase activities that specifically phosphorylate type III IF, including vimentin, glial fibrillary acidic protein and desmin, during mitosis. Cdk1 phosphorylates vimentin-Ser55 from prometaphase to metaphase, leading to the recruitment of Polo-like kinase 1 (Plk1) to vimentin. Upon binding to Phospho-Ser55 of vimentin, Plk1 is activated, and then phosphorylates vimentin-Ser82. During cytokinesis, Rho-kinase and Aurora-B specifically phosphorylate IF at the cleavage furrow. IF phosphorylation by Cdk1, Plk1, Rho-kinase and Aurora-B plays an important role in the local IF breakdown, and is essential for the efficient segregation of IF networks into daughter cells. As another part of our research on IF, we have set out to find the binding partners with simple epithelial keratin 8/18. We identified tumor necrosis factor receptor type 1-associated death domain protein (TRADD) as a keratin 18-binding protein. Together with data from other laboratories, it is proposed that simple epithelial keratins may play a role in modulating the response to some apoptotic signals. Elucidation of the precise molecular functions of IF is expected to improve our understanding of tumor development, invasion and metastasis.

Topics: Animals; Antibodies; Cell Transformation, Neoplastic; Humans; Intermediate Filaments; Keratins; Mitosis; Phosphorylation

We postulate that ductal carcinoma in situ (DCIS), and consequently breast carcinoma in general, is a lobar disease, as the simultaneously or asynchronously appearing, often multiple, in situ tumor foci are localized within a single lobe. Although the whole lobe is sick, carrying some form of genetic instability, the malignant transformation of the epithelial cells may appear localized to a part or different parts of the sick lobe at the same time or with varying time difference. It may be confined to terminal ductal lobular units (TDLUs), to ducts or both. The malignant transformation is often associated with aberrant branching and/or aberrant lobularization within the sick lobe. Involvement of a single individual TDLU or of a group of adjacent TDLUs generates a unifocal lesion. Multifocal lesions appear if distant TDLUs are involved. Diffuse growth pattern in DCIS indicates involvement of the larger ducts. The extent of the involved area in multifocal or diffuse cases varies considerably. Diffuse growth pattern with or without evidence of aberrant arborisation within the sick lobe seems to characterize a subgroup of DCIS with unfavourable prognosis. In this paper, we discuss the anatomical, embryological and pathological background of the theory of the sick lobe and present supporting evidence from modern radiological breast imaging, long-term follow-up studies and from our own series of 108 DCIS cases.

Topics: Biomarkers, Tumor; Breast Neoplasms; Carcinoma, Intraductal, Noninfiltrating; Cell Transformation, Neoplastic; Female; Humans; Keratins; Mammary Glands, Human; Precancerous Conditions

Until recently the myoepithelial cell has been studied relatively little in terms of its role in breast cancer. A number of malignancies showing myoepithelial differentiation have been reported in the literature, although they are still thought to be relatively rare and only limited studies are published. As a result of recent expression profiling experiments, one type of tumor with myoepithelial features, the so-called 'basal' breast cancer, has received a renewed interest, although it has been known to pathologists for more than two decades. These tumors, which express markers of both luminal and myoepithelial cells, are now being studied using antibodies against some new molecules that have emerged from studies of sorted normal luminal and myoepithelial cells. These immunohistochemical data, combined with genomic studies, may lead to better identification and management of patients with 'basal' tumors.

Topics: Animals; Biomarkers; Breast; Breast Neoplasms; Cell Differentiation; Cell Separation; Cell Transformation, Neoplastic; Epithelial Cells; Humans; Immunophenotyping; Keratins; Myoepithelioma; Neoplasms, Basal Cell; Stem Cells

Pathways to breast cancer have been difficult to define using morphology alone. Although epithelial hyperplasia of usual type (HUT) is associated with moderately elevated breast cancer risk, molecular evidence placing it in a cancer precursor pathway is not clear-cut. Recent evidence suggests that small numbers of cytokeratin 5/6 positive cells are precursors of separate lineages which acquire either cytokeratin 8/18/19 or smooth muscle actin (SMA) and cytokeratin 14 on separate pathways to fully differentiated epithelial and myoepithelial phenotypes (and may ultimately lose cytokeratin 5/6 expression). Immunohistochemistry shows that most HUT have a mixed precursor phenotype resembling normal breast with co-expression of cytokeratin 5/6, 8/18/19 and SMA, in contrast to atypical ductal hyperplasia (ADH) and ductal carcinoma in situ which typically have a 'mature' luminal phenotype positive for cytokeratin 8/18/19 but lacking cytokeratin 5/6 expression. While this supports the idea of a biological discontinuity between HUT and ADH/DCIS, caution is called for in the diagnostic use of these reagents until greater experience has been accumulated, and other published data do show features of HUT intermediate between normal breast lobules and ADH.

Topics: Breast; Breast Neoplasms; Carcinoma in Situ; Carcinoma, Ductal, Breast; Carcinoma, Intraductal, Noninfiltrating; Cell Transformation, Neoplastic; Epithelium; Female; Humans; Hyperplasia; Keratins; Loss of Heterozygosity; Neoplasm Invasiveness; Neoplasm Staging; Phenotype; Precancerous Conditions

The normal prostate shows a high degree of cellular organization. The basal layer is populated by prostate epithelial stem cells and a population of transiently proliferating/amplifying (TP/A) cells intermediate to the stem cells and fully differentiated cells. The luminal layer is composed of fully differentiated prostate epithelial cells. Neuroendocrine cells are scattered throughout the gland. This organization is also seen in prostate cancer, where the tumor cell origin (cancer stem cells) can be traced to a normal cell type by characteristic keratin expression patterns. Basal cells showed strong expression of K-[keratin]5, but they were only weakly positive for K18. Luminal cells strongly expressed K18. A subpopulation of basal cells coexpressed K5 and K14. These keratin expression patterns changed with the degree of cell differentiation as well as location. The least differentiated stem cells in the basal layer were positive for K5 and K14, with weak expression for K18. Intermediate stages of differentiation were identified by expression of K5 and K18. Neuroendocrine cells also expressed K5 as well as typical neuroendocrine cell markers (eg, chromogranin A). Evidence supporting the hypothesis that prostate cancer arises from malignant transformation of intermediate stem cells included the presence in prostate cancers of keratin patterns associated with the intermediate stages of differentiation, androgen independence of both prostate cancers and intermediate stem cells, and expression of c-met by both the TP/A intermediate stem cells and tumor cells.

Topics: Adenocarcinoma; Androgens; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Epithelial Cells; Humans; Immunophenotyping; Keratins; Male; Neoplasm Proteins; Neoplasms, Hormone-Dependent; Prostatic Neoplasms; Proto-Oncogene Proteins c-met; Stem Cells

Two cases of invasive carcinoma developing in extragonadal endometriosis are presented. Each case had a different clinical course. In addition to routine histopathologic studies immunohistochemical studies to assess the expression of cytoceratin and glycoprotein CD-44 were performed. In both cases CK-7 expression was higher in malignancy then in the endometrioid tissue. Very high expression of CD-44 protein (marker of metastatic potential) was found in patients with poor progress of the disease.

Topics: Abdomen; Adult; Biomarkers, Tumor; Carcinoma; Cell Transformation, Neoplastic; Endometriosis; Female; Gene Expression Regulation, Neoplastic; Humans; Hyaluronan Receptors; Keratin-7; Keratins; Perineum; Skin Neoplasms; Up-Regulation

Topics: Apoptosis; Biomarkers; Cell Differentiation; Cell Transformation, Neoplastic; Epithelial Cells; Homeostasis; Humans; Keratins; Male; Prostate; Prostatic Neoplasms; Receptors, Growth Factor; Stem Cells

This is Part 2 of a three-part review and deals with tumorigenic cell lines. Several immortalized and malignant adult human prostatic epithelial cell lines have been recently developed. The three most widely used carcinoma cell lines-DU-145, PC-3, and LNCaP-developed between 1977 and 1980, have greatly contributed to our current understanding of prostate cancer. Before a cell line can be accepted as having prostatic epithelial origin, some basic characteristics must be established. Expression of specific cytokeratins but absence of desmin and factor VIII should be first determined to establish epithelial origin. Responsiveness to androgens and expression of androgen receptor and prostate-specific antigen should be examined under stringent culture conditions to establish prostatic epithelial origin. Response to growth factors and expression of their receptors facilitates further characterization of cell behavior. Cell lines immortalized by human papillomaviruses (HPVs) are of special interest because HPVs are involved in a variety of anogenital cancers and may also play a role in prostate carcinogenesis. Malignant transformation of HPV-18 immortalized cells with the ras oncogene provides cell systems for investigating the multistep process of carcinogenesis. Each cell line has some unique characteristics, whether it arose directly from a carcinoma or resulted from immortalization with Simian virus 40 (SV40) or HPV, or was transformed in vitro by oncogenes. Comparisons of these characteristics should facilitate elucidation of the mechanisms involved in the initiation, promotion, and progression of prostate cancer. These cell lines will further serve as useful models for investigating tumor progression, invasion, metastasis, new therapeutic strategies, drug resistance, and its reversal and chemoprevention. The nontumorigenic cell lines were discussed in Part 1 [1]. This review summarizes the characteristics of several currently available tumorigenic, adult human prostatic epithelial cell lines.

Topics: Antigens, Neoplasm; Antigens, Surface; Carcinoma; Cell Line; Cell Transformation, Neoplastic; Epithelium; Glutamate Carboxypeptidase II; Growth Substances; Humans; Immunohistochemistry; Keratins; Male; Papillomaviridae; Prostate; Prostatic Neoplasms; Receptors, Androgen; Tumor Cells, Cultured

The use of oral exfoliative cytology in clinical practice declined due to the subjective nature of its interpretation and because there may be only a small number of abnormal cells identifiable in a smear. The more recent application of quantitative techniques, together with advances in immunocytochemistry, have refined the potential role of cytology, stimulating a reappraisal of its value in the diagnosis of oral cancer. This review considers the influence of the quantitative analysis of cytomorphology, DNA analysis and other tumour markers applied to oral exfoliative cytological samples. These studies indicate that oral cytology may provide an important adjunct in the assessment of the patient with a potentially cancerous oral lesion.

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Cytodiagnosis; DNA, Neoplasm; Flow Cytometry; Humans; Keratins; Mouth Mucosa; Mouth Neoplasms; Pathology, Oral; Process Assessment, Health Care

Malignant fibrous histiocytoma (MFH), arising in combination with a sacral chordoma in a 70-year-old men, is described. Intermediate spindle-shaped cells demonstrating keratin positivity, showed a gradual transition between the areas of conventional chordoma, and the spindle cell areas, lending credence to the theory of a multipotential neoplasm. We chose the descriptive term "chordoma with malignant spindle cell component" in the sense that high malignant sarcomatous components exists in conjunction with chordomas in the primary tumor and the local recurrence. A review of literature is undertaken chronicling the documented associations of chordoma and sarcoma, followed by a discussion of the various causes proposed to explain this phenomenon.

Topics: Aged; Biomarkers, Tumor; Cell Division; Cell Transformation, Neoplastic; Chordoma; Coccyx; Diagnosis, Differential; Histiocytoma, Benign Fibrous; Humans; Keratins; Male; Neoplasms, Multiple Primary; Sacrum; Spinal Neoplasms

Topics: Adult; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Keratins; Male; Mandibular Diseases; Mandibular Neoplasms; Neoplasm Invasiveness; Odontogenic Cysts

Clinical and experimental experience indicate that differentiation and malignancy are inversely correlated. However, more recent experimental studies using mouse and human keratinocyte systems have demonstrated that complete or even substantial loss in overall epithelial differentiation is not a prerequisite for malignant growth of cancer cells. Major defects in differentiation are also not a prerequisite for premalignant stages, in particular for cell immortalization, which is considered an early and essential step in the transformation process. Moreover, progressive dedifferentiation, often associated with advanced tumor stages, is also found in immortalized cell lines which are, however, nontumorigenic. On the other hand, malignant cell lines may have maintained a high degree of their normal differentiation program and sensitivity to differentiation modulators. However, to date no transformed keratinocyte cell lines with completely normal differentiation have been observed. Since epidermal keratinization is a very complex process involving many different parameters and is fully expressed only under in vivo conditions, an exact and quantitative comparison of such ill-defined phenomena (differentiation and malignancy) is still problematic. Obviously, both phenomena are under separate control and not causally linked. Nevertheless, a better understanding of factors and mechanisms regulating differentiation and of their disturbance in carcinogenesis would offer new possibilities to design novel tumor therapeutic strategies in the field of differentiation therapy.

Topics: Biomarkers; Carcinoma, Squamous Cell; Cell Differentiation; Cell Survival; Cell Transformation, Neoplastic; Disease Progression; Epidermal Cells; Epithelial Cells; Humans; Keratins; Neoplasm Proteins; Neoplasms; Protein Precursors; Skin Neoplasms; Tumor Cells, Cultured

CYFRA is a new marker which measures a fragment of cytokeratin 19 in the serum by an immune radiometric method. The test is based on the preservation of the cytokeratin expression on the epithelial cells during the course of malignant transformation. In immuno-histochemistry the antibodies which selectively recognise cytokeratin react with all histological types of bronchial cancer. The presence of cytokeratin in the serum of patients suffering from cancer would be linked to their liberation during the course of cellular death. The threshold of specificity for CYFRA 21-1 is 3.3.3.6 ng/ml in a population suffering from benign respiratory diseases. The study performed in bronchial cancer produced the following conclusions: the marker is, above all, useful for epidermoid cancer; it is more discriminating than other markers to separate bronchial cancers and non-malignant respiratory disease. An elevated level of CYFRA 21-1 is predictive of advanced disease but does not permit any prediction as to inoperability. In 65% of cases, variations of CYFRA 21-1 are concordant with the stage of the disease during chemotherapy. Finally, elevated levels of CYFRA 21-1 predict a poor prognosis independent of the state of the disease.

Topics: Biomarkers, Tumor; Bronchial Diseases; Bronchial Neoplasms; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; Humans; Immunoradiometric Assay; Keratins; Peptide Fragments; Prognosis; Sensitivity and Specificity

Malignant cutaneous cylindroma is a rare tumor. It has been described in 26 cases, both in the solitary form and in the autosomal dominant inherited multiple tumor form. The authors present two new cases that occurred in one family with a history of multiple cylindromas.. Clinical and histopathologic data of both tumors were compared with those of 26 other cases in the literature. Immunohistochemical examinations were performed.. The malignant tumors were distinguished from the benign lesions by rapid growth, long-standing ulceration, or bleeding. Histopathologic examination showed a well-differentiated carcinoma in one patient and a poorly differentiated tumor in the other. In the latter, lymph node metastasis developed, and the patient died 2.5 years later. Histopathologic criteria of malignancy included cell pleomorphism, frequent mitoses and loss of jigsaw pattern, peripheral palisading, hyaline sheaths, and dual cell population.. These observations are in accord with those in the literature. Malignant cutaneous cylindroma developed more often in the multiple tumor form than in the single tumor form. Malignant cylindroma is an aggressive carcinoma with a tendency to local destructive growth and metastases.

Topics: Aged; Antigens, Neoplasm; Carcinoma; Carcinoma, Adenoid Cystic; Cell Transformation, Neoplastic; Cytoplasm; Female; Humans; Hyalin; Keratins; Lymphatic Metastasis; Male; Membrane Glycoproteins; Mucin-1; S100 Proteins; Skin Neoplasms

Squamous differentiation in poral adnexal neoplasms is a rare event. Two cases are presented of malignant eccrine poroma in which areas of squamous differentiation showing the features of squamous cell carcinoma were present. The areas of squamous differentiation were found within the invasive components of the lesions and appeared to result from direct transformation of the poral epithelial cells. A review of the literature on this unusual phenomenon is presented.

Topics: Acrospiroma; Aged; Aged, 80 and over; Carcinoma, Squamous Cell; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Cytoplasmic Granules; Foot Diseases; Humans; Hyalin; Keratinocytes; Keratins; Male; Neoplasm Invasiveness; Sweat Gland Neoplasms; Thigh

Topics: Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Cellular Senescence; Epithelial Cells; Epithelium; Humans; Intermediate Filament Proteins; Intermediate Filaments; Keratins

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Biomarkers, Tumor; Cell Transformation, Neoplastic; Chromosome Aberrations; Chromosome Disorders; Karyotyping; Keratins; Mice; Papilloma; Precancerous Conditions; Skin Neoplasms; Tetradecanoylphorbol Acetate; Time Factors

Topics: Carcinogens; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Humans; Keratins

Topics: Animals; Calcium; Cell Differentiation; Cell Transformation, Neoplastic; Keratins; Mice; Neoplastic Stem Cells; Skin Neoplasms

The field of cellular senescence (cytogerontology) is reviewed. The historical precedence for investigation in this field is summarized, and placed in the context of more recent studies of the regulation of cellular proliferation and differentiation. The now-classical embryonic lung fibroblast model is compared to models utilizing other cell types as well as cells from donors of different ages and phenotypes. Modulation of cellular senescence by growth factors, hormones, and genetic manipulation is contrasted, but newer studies in oncogene involvement are omitted. A current consensus would include the view that the life span of normal diploid cells in culture is limited, is under genetic control, and is capable of being modified. Finally, embryonic cells aging in vitro share certain characteristics with early passage cells derived from donors of increasing age.

Topics: Aging; Blood; Cell Division; Cell Fusion; Cell Line; Cell Survival; Cell Transformation, Neoplastic; Epidermal Cells; Fibroblasts; Growth Substances; Humans; Keratins

Topics: Animals; Antigens, Polyomavirus Transforming; Antigens, Viral, Tumor; Breast; Carcinoma, Squamous Cell; Cell Survival; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cells, Cultured; Epithelium; Female; Humans; Keratins; Male; Methods; Oncogene Proteins, Viral; Oncogenes; Organ Specificity; Phenotype; Pregnancy; Simian virus 40; Skin

The cytoskeleton (CSK) of eukaryotic cells is composed of a complex interconnected network of filaments which is important in a wide variety of cellular functions including changes in cell shape, cell motility, mitosis, anchorage-dependent growth, and the localization of cellular organelles such as mitochondria, polyribosomes, and secretory granules. The various proteins comprising the cytoskeleton include actin in microfilaments, tubulin in microtubules, and the heterogeneous group of intermediate filament proteins that are associated with different cell types (keratin in epithelial cells, vimentin in fibroblasts, desmin in muscle cells, glial filament protein in glial cells, and the neurofilament protein subunits in neural tissue). Many other proteins in glial cells, and the neurofilament protein subunits in neural tissue). Many other proteins are closely associated with the cytoskeleton and influence its organization. In neoplastic cells, the expression of these different CSK proteins, especially the intermediate filament proteins, reflects their morphologic and functional differentiation. The carcinomas contain keratin; identification of individual keratin components may allow further sub-classification of carcinomas which is consistent with their tissue of origin. The sarcomas of muscle origin contain desmin. Vimentin is found primarily with cells of mesenchymal origin, but may coexist with other intermediate filament proteins in other tumors. One example is the coexistence of keratin and vimentin in tumors, such as mesotheliomas, which are derived from epithelial cells of embryonic origin. Glial fibrillary acidic protein is the most specific marker for glial tumors. Tumors of neural origin are characterized by the presence of neurofilament subunits. Therefore, analysis of CSK composition would be useful in diagnosis of clinical specimens and aid in studies of lineage relationships of neoplasms. Although no consistent differences in cytoskeletal structure between neoplastic and normal cells have been identified so far, the presence of more subtle biochemical alterations in the cytoskeletal structure of neoplastic cells that contributes to malignant behavior has not been ruled out. Since the cytoskeletal network plays an important role in cell shape and cell locomotion, which in turn are thought to be involved in growth control, invasion, and metastasis, further work is directed at identifying the various alterations in cytoskeletal architecture that

Topics: Actin Cytoskeleton; Actins; Animals; Astrocytes; Carcinoma; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Cytoskeleton; Desmin; Epithelium; Glial Fibrillary Acidic Protein; Glioma; Granulocytes; Humans; Intermediate Filament Proteins; Keratins; Leukemia; Lymphoma; Macrophages; Microtubules; Molecular Weight; Muscles; Neoplasm Metastasis; Neoplasms; Neoplasms, Nerve Tissue; Neurons; Sarcoma; Tissue Distribution; Vimentin

Epidermal tumourigenesis can be achieved in rodents by the application of a single subthreshold dose of a carcinogen (initiation) followed by repeated applications of a tumour promoter such as 12-0-tetradecanoyl phorbol, 13-acetate (TPA). TPA induces terminal differentiation in the majority of epidermal keratinocytes in vitro. However, transformed keratinocytes respond weakly to this terminal differentiation signal, and it is suggested that this property allows initiated cells and their progeny to obtain a selective advantage over their normal counterparts during promotion of papilloma formation by TPA. New data are reviewed which suggest that a putative wound hormone TGF-beta has similar differential effects on normal and transformed epithelial cells to those of TPA. It is proposed that the release of TGF-beta from platelets following deep skin wounding may be an explanation as to why wounding is a promoting stimulus but milder forms of epidermal injury are not. Weakly promoting hyperplasiogenic agents are also discussed within the context of a selection theory of tumour promotion.

Topics: Animals; Carcinogens; Cell Differentiation; Cell Transformation, Neoplastic; Epidermis; In Vitro Techniques; Keratins; Mice; Mitosis; Oncogenes; Papilloma; Peptides; Phenotype; Phorbols; Protein Kinase C; Skin; Skin Neoplasms; Tetradecanoylphorbol Acetate; Transforming Growth Factors

Topics: Actins; Cell Adhesion; Cell Differentiation; Cell Division; Cell Membrane; Cell Nucleus; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Gene Expression Regulation; Keratins; Mitosis; Tubulin; Vimentin; Virus Replication

Mallory's body filament assembly includes polypeptides of the cytokeratin class of intermediate filaments and also higher molecular weight polypeptides normally found only in the cytokeratins of mature keratinocytes of the epidermis. These additional polypeptides may alter both the morphologic characteristics and increase the resistance to dissolution of the filaments by Ca++-activated protease activity. Thus, it is likely that the kinetics of Mallory's body filament assembly and dissolution favor growth of the filaments. In rodents fed certain carcinogens, Mallory's body formation has been accompanied by the induction of the oncofetoenzyme gamma-glutamyl transpeptidase (GGT), suggesting that Mallory's body formation, like GGT induction, is a phenotypic change related to the process of neoplastic transformation in rodents.

Topics: Animals; Carcinogens; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Cytoskeleton; gamma-Glutamyltransferase; Griseofulvin; Humans; Intermediate Filament Proteins; Keratins; Liver; Liver Diseases; Liver Neoplasms; Liver Neoplasms, Experimental; Lung; Lung Diseases; Phosphorylation; Protein Precursors

Topics: 9,10-Dimethyl-1,2-benzanthracene; Amino Acids; Antigens; Cell Transformation, Neoplastic; Cells, Cultured; Culture Media; Culture Techniques; DNA; Epidermal Cells; Epidermis; Humans; Immunoglobulin G; Keratins; Melanocytes; Microscopy, Electron

Topics: Animals; Carcinogens; Cell Differentiation; Cell Division; Cell Survival; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Growth Inhibitors; Hematopoietic Stem Cells; Homeostasis; Hyperplasia; Keratins; Metaplasia; Mice; Mitogens; Models, Biological; Phorbol Esters; Skin Neoplasms; Wound Healing

The leucine rich repeats and immunoglobulin-like protein 1 (LRIG1) is a newly discovered negative regulator of epidermal growth factor receptor (EGFR) and a proposed tumor suppressor. It is not universally downregulated in human cancers, and its role in neoplastic transformation and tumorigenesis is not well-documented. In this study, we show the expression of LRIG1 as a novel potential marker for neoplastic transformation in ocular-surface squamous neoplasia (OSSN). The following two groups were included in this study: 1) benign group (3 cases; 1 with papilloma and 2 with dysplasia) and 2) malignant group (3 cases with squamous cell carcinoma (SCC)). In both groups, immunofluorescence analysis was firstly performed for keratins 4, 12, 13, and 15 to characterize the state of differentiation, and for Ki67 to evaluate the proliferation activity. Subsequently, LRIG1 and EGFR expression was analyzed. Either keratin 4 and/or 13, both non-keratinized epithelial cell markers, were generally expressed in both groups, except for 1 severe SCC case. Keratin 15, an undifferentiated basal cell marker, was more strongly expressed in the malignant cases than in the benign cases. The Ki67 index was significantly higher (P<0.002) in the malignant group (33.2%) than in the benign group (10.9%). LRIG1 expression was limited to basal epithelial cells in normal corneal epithelial tissue. Interestingly, LRIG1 was expressed throughout the epithelium in all the benign cases. In contrast, its expression was limited or totally disappeared in the malignant cases. Inversely, EGFR staining was faintly expressed in the benign cases, yet strongly expressed in the malignant cases. Malignant tissue with proliferative potential presented EGFR overexpression and inverse downregulation of LRIG1, consistent with LRIG1 being a suppressor of neoplastic transformation by counteracting the tumor growth property of EGFR. Our findings indicate that downregulation of LRIG1 is possibly a novel potential marker of transformation and tumorigenesis in OSSN cases.

Topics: Adult; Aged; Aged, 80 and over; Animals; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Down-Regulation; ErbB Receptors; Eye Neoplasms; Eye Proteins; Female; Gene Expression Regulation, Neoplastic; Humans; Keratins; Male; Membrane Glycoproteins; Middle Aged; Neoplasm Proteins

Consistent with cancer stem cell driven pattern of growth, human basal cell carcinomas (BCCs) demonstrate differentiation along hair follicle (HF) lineages.. To define the pattern of differentiation and therapeutic targets that promote BCC differentiation and therefore BCC cancer stem cell exhaustion.. An alkaline phosphatase substrate kit was used to determine dermal papilla cells within the BCC stroma. Autonomous HF cycle-dependent gene expression was identified by analysis of the human homologues of a murine gene set (total 2289 genes) that is differentially expressed in hair cycle phases. The findings were validated by quantitative real-time PCR and immunofluorescence, as well as in vitro transforming growth factor (TGF)-β2 stimulation of BCC cancer stem cell colonies.. As in the HF, keratin expression in the inner root sheath and matrix in BCC correlated with proliferative index and was tightly regulated, despite the absence of dermal papilla cells. Cross-species microarray analysis comparing human BCC and murine synchronous HF growth cycle datasets revealed 74% concordance with telogen differentiation compared with anagen (23%, P < 0.01) and catagen (49%; P < 0.01). Incomplete anagen differentiation within BCC was characterized by reduced expression of the anagen master regulator DLX3 (-5.5-fold), and increased expression of telogen-associated genes: AEBP1 (2.2-fold), DEFB8 (35.3-fold), MMP3 (106.0-fold) and MMP12 (12.9-fold). Restoration of dermal papilla signals by in vitro addition of TGF-β2 enhanced anagen differentiation.. Our findings show that BCC cells differentiate along HF lineages and may be susceptible to exogenous HF cycle modulators.

Topics: Animals; Carcinoma, Basal Cell; Cell Differentiation; Cell Transformation, Neoplastic; Fluorescent Antibody Technique; Gene Expression; Hair Follicle; Humans; Keratins; Mice; Oligonucleotide Array Sequence Analysis; Real-Time Polymerase Chain Reaction; Skin Neoplasms

Fibroblast growth factor receptor 1 (FGFR1) is frequently amplified in human small-cell lung cancer (SCLC), but its contribution to SCLC and other lung tumors has remained elusive. Here, we assess the tumorigenic capacity of constitutive-active FGFR1 (FGFR1

Topics: Adenocarcinoma of Lung; Animals; Bronchi; Cell Transformation, Neoplastic; Cisplatin; Drug Resistance, Neoplasm; Gene Expression Regulation, Neoplastic; Humans; Integrases; Keratins; Lung Neoplasms; Mice; Mutation; Nasal Cavity; Neurosecretory Systems; Oncogenes; Pulmonary Alveoli; Receptor, Fibroblast Growth Factor, Type 1; Retinoblastoma Protein; Small Cell Lung Carcinoma; Tumor Suppressor Protein p53

We used allogeneic bone marrow transplantation (BMT) and a mouse multistage cutaneous carcinogenesis model to probe recruitment of bone marrow-derived epithelial cells (BMDECs) in skin tumors initiated with the carcinogen, dimethylbenz[a]anthracene (DMBA), and promoted with 12-O-tetradecanolyphorbol-13-acetate (TPA). BMDECs clustered in the lesional epithelium, expressed cytokeratins, proliferated, and stratified. We detected cytokeratin induction in plastic-adherent bone marrow cells (BMCs) cultured in the presence of filter-separated keratinocytes (KCs) and bone morphogenetic protein 5 (BMP5). Lineage-depleted BMCs migrated towards High Mobility Group Box 1 (HMGB1) protein and epidermal KCs in ex vivo invasion assays. Naive female mice receiving BMTs from DMBA-treated donors developed benign and malignant lesions after TPA promotion alone. We conclude that BMDECs contribute to the development of papillomas and dysplasia, demonstrating a systemic contribution to these lesions. Furthermore, carcinogen-exposed BMCs can initiate benign and malignant lesions upon tumor promotion. Ultimately, these findings may suggest targets for treatment of non-melanoma skin cancers.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Bone Marrow Cells; Bone Marrow Transplantation; Bone Morphogenetic Protein 5; Cell Movement; Cell Plasticity; Cell Transformation, Neoplastic; Coculture Techniques; Epithelial Cells; Female; Hair Follicle; HMGB1 Protein; Keratinocytes; Keratins; Male; Mice; Mice, Inbred C57BL; Neoplasm Invasiveness; Papilloma; Skin Neoplasms; Stem Cells; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured

Malignant transformation of fibrous dysplasia (FD) is exceedingly rare, occurring in less than 1% of all FD cases, and has been described in both monostotic and polyostotic forms of this entity. We report a case of a large proximal femur mass arising in a 45-year-old man. The biopsy revealed a high-grade pleomorphic malignancy that focally expressed multiple keratins. Based on the presence of keratin immunoreactivity, the morphologic differential diagnosis included metastatic sarcomatoid carcinoma. However, review of the clinical information revealed a history of polyostotic FD, and imaging findings were compatible with malignant transformation of FD. The resected neoplasm was biphasic and composed of areas of conventional FD admixed with a high-grade pleomorphic malignancy. Activating GNAS mutations were identified in both components. To the best of our knowledge, this is the first description of keratin expression in malignant transformation of FD.

Topics: Biomarkers, Tumor; Biopsy; Cell Transformation, Neoplastic; Chromogranins; DNA Mutational Analysis; Femoral Neoplasms; Fibrous Dysplasia, Polyostotic; GTP-Binding Protein alpha Subunits, Gs; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Mutation; Osteotomy

The aim of this study was to investigate the relationship the expression of cytokeratins (CK10 and CK13) and the cell proliferation index determined by Ki-67 of lip squamous cell carcinoma and actinic cheilitis with different degrees of dysplasia.. Forty-five paraffin-embedded actinic cheilitis with and without dysplasia and 20 lip squamous cell carcinoma were analyzed by immunohistochemistry using anti-human anti-CK10, anti-CK13, and anti-Ki-67 antibodies.. The majority of actinic cheilitis showed immunopositivity for CK10 and CK13 with decrease or loss of expression in dysplastic areas. In lip squamous cell carcinoma of the lip, heterogeneous expression of CK13 and immunonegativity for CK10 were observed. There was a statistically significant difference between CK10 expression in lip squamous cell carcinoma and in actinic cheilitis with or without dysplasia (p < 0.001). The cell proliferation index was higher in actinic cheilitis with dysplasia and lip squamous cell carcinoma than in actinic cheilitis without epithelial dysplasia. A significant correlation was found between the intensity of the epithelial dysplasia and the cell proliferation index (p < 0.001).. These results provide evidence that there is a downregulation of CK10 expression in dysplastic areas of patients with actinic cheilitis and in those with lip squamous cell carcinoma (LSCC) and that the index of cell proliferation, determined by Ki-67, is directly correlated with the intensity of the epithelial dysplasia.. Altogether, these results suggest that CK10 expression and the epithelial cell proliferation index can help to identify malignant transformation in the lip region.

Topics: Carcinoma, Squamous Cell; Cell Proliferation; Cell Transformation, Neoplastic; Cheilitis; Humans; Keratins; Lip Neoplasms

To predict carcinogenic potential of AgNPs on the respiratory system, BEAS-2B cells (human bronchial epithelial cells) were chronically exposed to low- and non-cytotoxic dose (0.13 and 1.33μg/ml) of AgNPs for 4months (#40 passages). To assess malignant cell transformation of chronic exposure to AgNPs, several bioassays including anchorage independent agar colony formation, cell migration/invasion assay, and epithelial-mesenchymal transition (EMT) were performed in BEAS-2B cells. Chronic exposure to AgNPs showed a significant increase of anchorage independent agar colony formation and cell migration/invasion. EMT, which is the loss of epithelial markers (E-Cadherin and Keratin) and the gain of mesenchymal marker (N-cadherin and Vimentin), was induced by chronic exposure to AgNPs. These responses indicated that chronic exposure to AgNPs could acquire characteristics of tumorigenic cells from normal BEAS-2B cells. In addition, caspase-3, p-p53, p-p38, and p-JNK were significantly decreased, while p-ERK1/2 was significantly increased. MMP-9 related to cell migration/invasion was upregulated, while a MMP-9 inhibitor, TIMP-1 was down-regulated. These results indicated that BEAS-2B cells exposed to AgNPs could induce anti-apoptotic response/anoikis resistance, and cell migration/invasion by complex regulation of MAPK kinase (p38, JNK, and ERK) and p53 signaling pathways. Therefore, we suggested that long-term exposure to low-dose of AgNPs could enhance malignant cell transformation in non-tumorigenic BEAS-2B cells. Our findings provide useful information needed to assess the carcinogenic potential of AgNPs.

Topics: Antigens, CD; bcl-2-Associated X Protein; Cadherins; Caspase 3; Cell Line; Cell Movement; Cell Survival; Cell Transformation, Neoplastic; Epithelial-Mesenchymal Transition; Humans; Keratins; Matrix Metalloproteinase 9; Metal Nanoparticles; Proto-Oncogene Proteins c-bcl-2; RNA, Messenger; Silver; Tissue Inhibitor of Metalloproteinase-1; Tumor Suppressor Protein p53; Vimentin

Chronic venous leg ulcers (VLU), especially long-lasting non-healing ulcers, are among the risk factors for squamous cell carcinoma (SCC). Malignant transformation of a VLU is a rare finding and the relative risk of carcinomatous transformation is quite low (about 5.8). SCC arising in the context of a VLU has a particularly aggressive behavior. A 76-year-old male patient with no relevant medical familial history, with chronic venous insufficiency CEAP C6 for 10 years [recurrent leg ulcers with favorable outcome (healing) after specific local and systemic treatment], showing for about three years one ulcerated lesion located on the anterior upper third of the right calf non-responsive to specific treatment, which subsequently increased their size and merged. Biopsy sample was taken. Histopathology showed epidermal acanthosis, papillomatosis, intense parakeratosis, pseudoepitheliomatous hyperplasia, dysplasia and moderately differentiated squamous cell carcinoma with areas of acantholysis. Immunohistochemistry (Ki67, EMA, cytokeratin 34βE12 and p63) was performed and all types of immunostaining were moderately to intense positive. Above-knee leg amputation and specific oncologic treatment were proposed as possible curative solutions but the patient refused. Ten months after diagnosis and discharge form the Department of Dermatology, the patient died. Patients with chronic venous leg ulcers and clinically suspicious lesions should be evaluated for malignant transformation of the venous lesion. When diagnosed, malignancy complicating a chronic venous leg ulcer requires a resolute treatment as it may be fatal.

Topics: Aged; Amputation, Surgical; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Fatal Outcome; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Leg; Leg Ulcer; Male; Mucin-1; Risk Factors; Skin Neoplasms; Transcription Factors; Tumor Suppressor Proteins; Varicose Ulcer

We aimed to delineate the morphogenesis of aberrant nuclear features of urothelial carcinoma (UC) cells in association with cytokeratin (CK) expression patterns and cell proliferation activity. Correlation analysis of the nuclear area by morphometry and the expression patterns of CK5, CK20 and Ki-67 by triple immunofluorescence analysis was applied to 1699 cells from five low-grade and seven high-grade cases of UC. The majority of UC cells showed aberrant cellular differentiation represented by abnormal CK expression patterns of CK5+ / CK20+ (40.5%) or CK5- / CK20+ (56.0%). CK5+ / CK20- cells, a phenotype of cancer stem/progenitor cells, represented a very small population (1.9%) and showed a low proliferation activity. Ki-67+ cells showed a significantly different CK expression pattern compared with that of Ki-67(-) cells. The nuclear areas of CK5- / CK20+ cells (71.3 ± 25.9 μm2) were significantly larger than those of CK5+ / CK20+ cells (66.6 ± 25.5 μm2). Negativity for CK5 was related to the grade of UC and an increased number of CK5- / CK20+ / Ki-67+ cells was related to a higher malignant potential. We conclude the nuclear morphology is related to cell differentiation represented by CK expression and cell proliferative activity.

Topics: Aged; Biomarkers, Tumor; Cell Nucleus; Cell Transformation, Neoplastic; Female; Fluorescent Antibody Technique; Humans; Japan; Keratins; Ki-67 Antigen; Male; Middle Aged; Neoplastic Stem Cells; Urologic Neoplasms; Urothelium

Bone marrow-derived stem cells (BMDCs) have the ability to differentiate into lung epithelial cells in response to damage; however, their role in squamous cell carcinoma (SCC) formation is unknown. This study aimed to determine whether BMDC-derived lung epithelial cells could contribute to SCC formation. A model of lung SCC induced with N-nitrosodiethylamine (NDEA) in recipient female mice transplanted with green fluorescent protein (GFP)-positive BMDCs from male donors was established. Incorporation of BMDCs in lung tissue was determined using immunohistochemistry and immunofluorescence to detect GFP expression and fluorescence in situ hybridization to Y chromosomes. BMDC appeared at three stages of lung SCC progression: metaplasia, dysplasia, and carcinoma. There was a significantly higher proportion of GFP-positive (GFP(+)) cells within SCC than was found in metaplasia and dysplasia 16 weeks post-transplantation (both P < 0.017); GFP(+) BMDCs were also observed in clusters within several SCC nests. Furthermore, most GFP(+) cells in SCC were pancytokeratin-positive (PCK(+)) epithelial cells, and some exhibited proliferative activity as determined by Ki67 staining (9.7 ± 3.92 %). The presence of GFP(+)Ki67(+)PCK(+) cells within SCC nests suggested that some donor BMDCs differentiated into proliferating epithelial cells. Finally, analysis of p63 expression, a marker of SCC cells, indicated that the presence of GFP(+)p63(+) cells (green) in inner parts of the SCC. These findings strongly suggest that BMDC-derived lung epithelial cells could participate in lung SCC formation and partially contribute to tumor growth, which might have significant potential implications for both clinical cancer therapy using BMDCs.

Topics: Animals; Bone Marrow Cells; Bone Marrow Transplantation; Carcinoma, Squamous Cell; Cell Differentiation; Cell Movement; Cell Proliferation; Cell Transformation, Neoplastic; Diethylnitrosamine; Epithelial Cells; Female; Green Fluorescent Proteins; Keratinocytes; Keratins; Ki-67 Antigen; Lung Neoplasms; Male; Mice; Mice, Inbred C57BL; Phosphoproteins; Stem Cells; Trans-Activators

Topics: Aged, 80 and over; Carcinoma; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Male; Skin Neoplasms; Tumor Suppressor Protein p53

Our aim was to examine cell transition events by detecting the frequency of intrapithelial α-smooth muscle actin (SMA)(+)/cytokeratin (CK)(+) cells during colorectal adenoma-carcinoma sequence, in relation to E-cadherin expression. Our further aim was to determine the proliferative activity of intraepithelial α-SMA(+) cells. Histologically healthy, adenoma, and colorectal cancer (CRC) biopsy samples were taken during routine colonoscopy and were included into tissue microarrays (TMAs). Slides immunostained for Ki-67, α-SMA, E-cadherin and pan-cytokeratin were digitalized and analyzed by using a digital microscope software. The proportion of α-SMA(+)/CK(+) cells was significantly higher in CRC samples (3.34 ± 1.01%) compared to healthy (1.94 ± 0.69%) or adenoma (1.62 ± 0.78%) samples (p < 0.01). E-cadherin expression negatively correlated with the number of α-SMA(+) cells. The majority of intraepithelial α-SMA(+) cells were in the proliferative phase. During tumor progression, the appearance of dot-like α-SMA staining in CK positive cells may indicate the initial phase of the epithelial-to-mesenchymal transition (EMT). The high proportion of intraepithelial α-SMA(+) proliferating cells may refer to their increased plasticity compared to differentiated cells. The negative correlation between E-cadherin and intraepithelial α-SMA expression suggests that EMT is facilitated by a loss of epithelial cell contact.

Topics: Actins; Adenoma; Biomarkers, Tumor; Cadherins; Cell Differentiation; Cell Transformation, Neoplastic; Colon; Colorectal Neoplasms; Disease Progression; Epithelial-Mesenchymal Transition; Follow-Up Studies; Humans; Immunoenzyme Techniques; Keratins; Longitudinal Studies; Neoplasm Staging; Prognosis; Rectum; Tissue Array Analysis

Central mucoepidermoid carcinoma (MEC) is a rare neoplasm arising intraosseously in the jaws. To clarify the clinicopathologic profile and pathogenesis of central MEC, clinicopathologic findings and follow-up data of 39 cases were collected and analyzed. There were 16 male and 23 female patients (median age, 43 y). Sixteen cases affected the maxilla, and 23 occurred in the mandible. Radiographically, most cases (32 of 39) showed a unilocular or multilocular radiolucency with bone destruction, and 7 were found with scattered calcification. The margins of the lesions were ill defined or diffused in 14 cases and relatively well defined in 25 cases. Most cases (26 of 39) were classified as low-grade MECs, whereas 13 were moderate-to-high grade. Follow-up data were available for 35 patients with a median period of 36 months. All cases were found to be primary; local recurrence occurred in 8 cases, most (75.0%) of which were low-grade tumors. Four cases showed regional lymph node metastasis, and 1 developed distant metastasis. Of 11 cases with a clinical history of the jaw cyst, 8 initially showed a typical odontogenic cyst with local MEC-like proliferation. In summary, the most likely pathogenesis of central MEC is neoplastic transformation of the epithelial lining of an odontogenic cyst, diagnosis of which should be based on clinical, radiographic, and histopathologic findings. The immunohistochemical profile of keratins is helpful in differential diagnosis. Radical surgery is the treatment of choice, whereas the role of radiotherapy or chemotherapy is still controversial, and careful long-term follow-up is necessary.

Topics: Adolescent; Adult; Aged; Asian People; Carcinoma, Mucoepidermoid; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Jaw Neoplasms; Keratins; Male; Middle Aged; Neoplasm Grading; Odontogenic Cysts; Young Adult

Epithelial-myoepithelial carcinoma (EMC) is a rare low-grade salivary gland malignancy of presumed intercalated duct origin comprising 1% of all salivary gland tumours. High grade transformation (HGT) in EMC is a recently recognised entity with only a few cases reported in the literature. The authors report an additional case of EMC with HGT involving the submandibular gland. The patient was a 60-year-old woman who requested examination of the rapid growth of a mass in the left submandibular area, which she had first noticed 20 years previously. Histologically, the tumour had two distinct carcinomatous components. One component had features of a low grade EMC. The second component consisted of polygonal cells, arranged in a solid and nested pattern, with marked nuclear pleomorphism, brisk mitotic activity, and frequent necrosis. The Ki-67 labelling index of the EMC component was 9%, and that of the high grade component was 40%. The patient developed multiple pulmonary metastases 15 months after surgery. The aggressive behaviour of EMC with HGT suggests that it is important to recognise this variant of EMC to avoid misdiagnosis and inappropriate treatment.

Topics: Calcium-Binding Proteins; Calmodulin-Binding Proteins; Calponins; Carcinoma; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Diagnostic Errors; Female; Follow-Up Studies; Humans; Keratins; Ki-67 Antigen; Lung Neoplasms; Membrane Proteins; Microfilament Proteins; Middle Aged; Mitotic Index; Necrosis; Neoplasm Invasiveness; Submandibular Gland Neoplasms

Malignant transformation is rarely seen in the disease course of mature cystic teratoma (MCT) of the ovary. Adenocarcinoma arising from MCT is especially rare. We herein present the case of a premenopausal woman with a mucinous borderline-like tumor arising from a MCT. Based on the histological transition between the borderline-like tumor and gastrointestinal elements of the MCT, we consider that the tumor derived from teratomatous gastrointestinal epithelium. Immunohistochemistry showed that the proliferating mucinous cells were diffusely positive for cytokeratin 20 and partially positive for cytokeratin 7. MUC5AC was partially positive, whereas MUC2 and MUC6 were positive in a small number of tumor cells. The immunophenotype of cytokeratins and mucins in the present case was compatible with malignant transformation of the teratomatous gastrointestinal epithelium.

Topics: Adenocarcinoma, Mucinous; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Middle Aged; Mucins; Ovarian Neoplasms; Phenotype; Teratoma

Recent studies have suggested that some ovarian and pelvic serous carcinomas could originate from the fimbriated end of the distal fallopian tube. To test this hypothesis, we immortalized a normal human fallopian tube epithelial (FTE) cell line by using retrovirus-mediated overexpression of the early region of the SV40 T/t antigens and the human telomerase reverse transcriptase subunit (hTERT). These immortalized FTEs were then transformed by ectopic expression of oncogenic human HRAS (V12) . Tumorigenicity of the immortalized and/or transformed cells was subsequently tested by anchorage-independence growth assay and inoculation into nude mice via subcutaneous and intraperitoneal injection. As expected, the HRAS (V12) -transformed FTEs produced tumors through both subcutaneous and intraperitoneal injections, whereas no tumor growth was observed in immortalized FTEs. Unexpectedly, histopathological examination of tumors resulting from subcutaneous as well as intraperitoneal injections revealed largely poorly differentiated mucinous adenocarcinoma mixed with undifferentiated carcinoma. The tumor implants invaded extensively to the liver, colon, spleen, omentum, adrenal gland and renal capsule. Immunohistochemical staining of tumor cells showed positive staining for the epithelial cell markers cytokeratin AE1/AE3 and Müllerian lineage marker PAX8. Our study demonstrates that FTEs can generate poorly differentiated mucinous adenocarcinoma mixed with undifferentiated carcinoma through genetic modifications. Thus, we provide the first experimental evidence that fimbrial epithelial cells of the fallopian tube could be a potential source of ovarian mucinous adenocarcinoma.

Topics: Adenocarcinoma, Mucinous; Animals; Antigens, Polyomavirus Transforming; Carcinoma; Carcinoma, Ovarian Epithelial; Cell Line, Transformed; Cell Transformation, Neoplastic; Epithelial Cells; Fallopian Tubes; Female; Humans; Immunohistochemistry; Keratins; Mice; Mice, Nude; Neoplasms, Glandular and Epithelial; Ovarian Neoplasms; Paired Box Transcription Factors; PAX8 Transcription Factor; Proto-Oncogene Proteins p21(ras); Telomerase

Myelosarcomas are, due to their rarity, a difficult differential diagnosis. Not infrequently, extensive immunohistochemical staining for characterization of the tumor is performed, if one does not directly think of myelosarcoma. In the present case, there was a positivity of the myeloid blasts for cytokeratin. This may complicate the discrimination of myelosarcoma from carcinoma, in particular small cell carcinoma, not only in the mediastinum, but also in the skin, e.g., Merkel cell carcinoma.

Topics: Adenocarcinoma; Biomarkers, Tumor; Cell Transformation, Neoplastic; Diagnosis, Differential; Female; Humans; Keratins; Leukemia, Myeloid, Acute; Lung Neoplasms; Lymph Node Excision; Lymph Nodes; Lymphatic Metastasis; Mediastinum; Middle Aged; Myeloid Cells; Neoplasms, Multiple Primary

Standard cancer cell lines do not model the intratumoural heterogeneity situation sufficiently. Clonal selection leads to a homogeneous population of cells by genetic drift. Heterogeneity of tumour cells, however, is particularly critical for therapeutically relevant studies, since it is a prerequisite for acquiring drug resistance and reoccurrence of tumours. Here, we report the isolation of a highly tumourigenic primary pancreatic cancer cell line, called JoPaca-1 and its detailed characterization at multiple levels. Implantation of as few as 100 JoPaca-1 cells into immunodeficient mice gave rise to tumours that were histologically very similar to the primary tumour. The high heterogeneity of JoPaca-1 was reflected by diverse cell morphology and a substantial number of chromosomal aberrations. Comparative whole-genome sequencing of JoPaca-1 and BxPC-3 revealed mutations in genes frequently altered in pancreatic cancer. Exceptionally high expression of cancer stem cell markers and a high clonogenic potential in vitro and in vivo was observed. All of these attributes make this cell line an extremely valuable model to study the biology of and pharmaceutical effects on pancreatic cancer.

Topics: AC133 Antigen; Aldehyde Dehydrogenase 1 Family; Alleles; Animals; Antigens, CD; Antimetabolites, Antineoplastic; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Deoxycytidine; Disease Models, Animal; Drug Resistance, Neoplasm; Gemcitabine; Genomic Instability; Glycoproteins; GPI-Linked Proteins; Humans; Isoenzymes; Keratins; Male; Mesothelin; Mice; Middle Aged; Mutation; Neoplasm Metastasis; Neoplastic Stem Cells; Pancreatic Neoplasms; Peptides; Polyploidy; Retinal Dehydrogenase; Transplantation, Heterologous; Tumor Microenvironment

Jaagsiekte sheep retrovirus (JSRV) is the causative agent of a contagious lung cancer in sheep. The envelope protein (Env) is the oncogene, as it can transform cell lines in culture and induce tumors in animals, although the mechanisms for transformation are not yet clear because a system to perform transformation assays in differentiated type II pneumocytes does not exist. In this study we report culture of primary rat type II pneumocytes in conditions that favor prolonged expression of markers for type II pneumocytes. Env-expressing cultures formed more colonies that were larger in size and were viable for longer periods of time compared to vector control samples. The cells that remained in culture longer were confirmed to be derived from type II pneumocytes because they expressed surfactant protein C, cytokeratin, displayed alkaline phosphatase activity and were positive for Nile red. This system will be useful to study JSRV Env in the targets of transformation.

Topics: Alkaline Phosphatase; Alveolar Epithelial Cells; Animals; Cell Proliferation; Cell Transformation, Neoplastic; Cells, Cultured; Colony-Forming Units Assay; Jaagsiekte sheep retrovirus; Keratins; Male; Oncogene Proteins, Viral; Peptides; Rats; Rats, Sprague-Dawley; Viral Envelope Proteins

Assessing epithelial dysplasia to predict malignant transformation remains problematic in many tissues because grading systems are poorly structured and individual features poorly defined. Dysplasia grading is criticised for lack of reproducibility and poor predictive value. Grading systems for upper aerodigestive tract dysplasia have evolved over several decades and are not supported by good outcome experimental data..  This study analysed the individual features of dysplasia in 86 oral dysplastic lesions and determined the reproducibility of scoring for each, and correlated them with other features and clinical factors using complex clustering analyses.. A uniform pattern of dysplasia was found in 37 lesions, focal dysplasia in 36 and in 13 lesions dysplasia formed complex discontinuous patterns. There was wide variation in reproducibility of scoring of individual features and many, including thickness, some types of rete morphology, basaloid cell anisonucleosis, basal dyscohesion, and dyskeratosis as deep single cells correlated with sub-sites. Rete morphology, type of keratinisation, hyperchromatism of the basaloid compartment, prickle cell anisonucleosis and extension down salivary ducts correlated with smoking. Conventional grading and oral intraepithelial neoplasia (OIN) grading by 'thirds affected' showed strong correlation overall but scores obtained with the OIN system tended to a higher grade at all sites except soft palate/fauces. There was poor correlation between the systems for moderate dysplasia and also severe dysplasia at some sites. Individual features could not be shown to cluster to form distinct patterns of dysplasia.. These variations may account in part for the lack of reproducibility and poor predictive value of the grading systems in current use and could inform the design of future grading systems.

Topics: Carcinoma in Situ; Cell Adhesion; Cell Nucleus; Cell Transformation, Neoplastic; Chromatin; Epithelial Cells; Epithelium; Female; Humans; Keratins; Leukoplakia, Oral; Lip Neoplasms; Male; Mitosis; Mouth Floor; Mouth Mucosa; Mouth Neoplasms; Neoplasm Grading; Palatal Neoplasms; Palate, Soft; Precancerous Conditions; Reproducibility of Results; Salivary Ducts; Tongue Neoplasms

This laboratory has generated a series of seven cadmium (Cd(+2))- and six arsenite (As(+3))-transformed urothelial cancer cell lines by exposure of parental UROtsa cells to each agent under similar conditions of exposure. In this study, the seven Cd(+2)-transformed cell lines were characterized for the expression of keratin 6, 16, and 17 while the six As(+3) cell lines were assessed for the expression of keratin 7 and 19. The results showed that the series of Cd(+2)-transformed cell lines and their respective transplants all had expression of keratin 6, 16, and 17 mRNA and protein. The expression of keratin 6, 16, and 17 was also correlated with areas of the urothelial tumor cells that had undergone squamous differentiation. The results also showed that four of the six As(+3)-transformed cell lines had expression of keratin 7 and 19 mRNA and protein and produced subcutaneous tumors with intense focal staining for keratin 7 and 19. The other two As(+3)-transformed cell lines had very low expression of keratin 7 mRNA and protein and produced subcutaneous tumors having no immunoreactivity for keratin 7; although keratin 19 expression was still present. The peritoneal tumors produced by one of these two cell lines regained expression of keratin 7 protein. The present results, coupled with previous studies, indicate that malignant transformation of UROtsa cells by Cd(+2) or As(+3) produce similar patterns of keratin 6, 7, 16, 17, and 19 in the resulting series of cell lines and their respective tumors.

Topics: Animals; Arsenites; Biomarkers, Tumor; Blotting, Western; Cadmium; Cell Line; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; Humans; Keratins; Mice; Mice, Nude; Neoplasm Transplantation; Real-Time Polymerase Chain Reaction; RNA, Messenger; Transplantation, Heterologous; Urinary Bladder Neoplasms; Urothelium

Stable cell lines obtained by spontaneous immortalization might represent early stages of malignant transformation and be useful experimental models for studies of mechanisms of cancer development. The FHC (fetal human cells) cell line has been established from normal fetal colonic mucosa. Detailed characterization of this cell line and mechanism of spontaneously acquired immortality have not been described yet. Therefore, we characterized the FHC cell line in terms of its tumorigenicity, cytogenetics, and TP53 gene mutation analysis. FHC cells displayed capability for anchorage-independent growth in semisolid media in vitro and formed solid tumors after transplantation into SCID (severe combined immunodeficiency) mice. This tumorigenic phenotype was associated with hypotriploidy and chromosome number ranging from 66 to 69. Results of comparative genetic hybridization arrays showed that most chromosomes included regions of copy number gains or losses. Region 8q23 approximately 8q24.3 (containing, e.g., MYC proto-oncogene) was present in more than 20 copies per nucleus. Moreover, we identified mutation of TP53 gene in codon 273; triplet CGT coding Arg was changed to CAG coding His. Expression of Pro codon 72 polymorphic variant of p53 was also detected. Mutation of TP53 gene was associated with abolished induction of p21(Waf1/Cip1) and MDM-2 proteins and resistance to apoptosis after genotoxic treatment. Because of their origin from normal fetal colon and their relative resistance to the induction of apoptosis, FHC cells can be considered a valuable experimental model for various studies.

Topics: Animals; Apoptosis; Carcinoembryonic Antigen; Cell Adhesion; Cell Growth Processes; Cell Line, Transformed; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Comparative Genomic Hybridization; Cytogenetic Analysis; DNA Damage; DNA Mutational Analysis; Female; Fetus; Genes, p53; HCT116 Cells; Humans; In Situ Hybridization, Fluorescence; Karyotyping; Keratins; Mice; Mice, SCID; Neoplasm Transplantation; Phenotype; Proto-Oncogene Mas; Signal Transduction

Global hypomethylation is a common epigenetic event in cancer. Keratocystic odontogenic tumor (KCOT) and ameloblastoma are different tumors but posses the same tissue in origin. Here, we investigated long interspersed nuclear element-1 (LINE-1 or L1) methylation status between ameloblastoma and KCOT.. We studied the methylation levels of the long interspersed nucleotide element-1 (LINE-1) in ameloblastoma and KCOT. After collecting ameloblastoma cells and epithelium lining cells of KCOT by laser capture microdissection from paraffin embedded tissue, combined bisulfite restriction analysis of LINE-1 (COBRALINE-1) was performed to measure LINE-1 methylation levels.. The LINE-1 methylation level in KCOT (53.16 +/- 12.03%) was higher than that in ameloblastoma (36.90 +/- 16.52%), with a statistical significance of P = 0.001. The ranges of LINE-1 methylation of both lesions were not associated with either age or sex.. We found LINE-1 hypomethylation levels between ameloblastoma and KCOT are different. Therefore, global methylations between these tumors are processed differently.

Topics: Adult; Aged; Aged, 80 and over; Ameloblastoma; Cell Transformation, Neoplastic; Child; DNA Methylation; DNA, Neoplasm; Female; Humans; Jaw Neoplasms; Keratins; Long Interspersed Nucleotide Elements; Male; Middle Aged; Odontogenic Tumors; Promoter Regions, Genetic; Restriction Mapping; Young Adult

Luminal cells are believed to be the cells of origin for human prostate cancer, because the disease is characterized by luminal cell expansion and the absence of basal cells. Yet functional studies addressing the origin of human prostate cancer have not previously been reported because of a lack of relevant in vivo human models. Here we show that basal cells from primary benign human prostate tissue can initiate prostate cancer in immunodeficient mice. The cooperative effects of AKT, ERG, and androgen receptor in basal cells recapitulated the histological and molecular features of human prostate cancer, with loss of basal cells and expansion of luminal cells expressing prostate-specific antigen and alpha-methylacyl-CoA racemase. Our results demonstrate that histological characterization of cancers does not necessarily correlate with the cellular origins of the disease.

Topics: Animals; Biomarkers, Tumor; Cell Separation; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Flow Cytometry; Humans; Keratins; Male; Mice; Mice, Inbred NOD; Mice, SCID; Prostate; Prostatic Intraepithelial Neoplasia; Prostatic Neoplasms; Proto-Oncogene Proteins c-akt; Receptors, Androgen; Trans-Activators; Transcriptional Regulator ERG; Transduction, Genetic

Adamantinomas of the long bones are low-grade malignant tumours. They seem to be related to osteofibrous dysplasia with a mesenchymal-to-epithelial transformation. We report a case of an adamantinoma with a revertant sarcomatoid transformation that showed a complete loss of epithelial differentiation. It corresponded to a 41-year-old male presented with an 8-cm multilobated lesion in the centre of the distal tibia. On the en bloc resection specimen, areas of classic adamantinoma were found but most of the tumor corresponded to a high-grade neoplasm with 2 histologic patterns: one made up by epithelial nests with a basaloid arrangement and positive for pankeratins and so-called glandular keratins, and a second sarcomatoid component, negative for these epithelial markers. Five months after surgery the patient had a massive relapse that consisted solely of the second sarcomatous component also negative for epithelial antibodies.Three cases of adamantinomas with sarcomatoid transformation of the epithelial component have been described but the tumours still preserved an epithelial immunophenotype. However, our case represents the extreme end of the sarcomatoid dedifferentiation of a classic adamantinoma with complete sarcomatoid transformation and total loss of epithelial markers. To our knowledge this has not been described previously.

Topics: Adamantinoma; Adult; Biomarkers; Biomarkers, Tumor; Bone Neoplasms; Cell Transformation, Neoplastic; Humans; Keratins; Male; Mesoderm; Neoplasms, Second Primary; Phenotype; Sarcoma; Tibia; Treatment Outcome

The c-MET proto-oncogene, encoding the p190 hepatocyte growth factor tyrosine kinase receptor, can acquire oncogenic potential by multiple mechanisms, such as gene rearrangement, amplification and overexpression, point mutation, and ectopic expression, all resulting in its constitutive activation. Hepatocyte growth factor receptor truncated forms are generated by post-translational cleavage: p140 and p130 lack the kinase domain and are inactive. Their C-terminal remnant fragments are generally undetectable in normal cells, but a membrane-associated truncated form is recognized by anti-C-terminus antibodies in some human tumors, suggesting that a hepatocyte growth factor receptor lacking the ectodomain, but retaining the transmembrane and intracellular domains (Met-EC-), could acquire oncogenic properties. Herein we show that NIH-3T3 cells transduced with MET-EC- expressed a membrane-associated constitutively tyrosine-phosphorylated 60-kDa protein and, similarly to NIH-3T3 cells expressing the cytosolic oncoprotein Tpr-Met, showed activated extracellular regulated kinase 1/2 mitogen-activated protein kinase and Akt downstream transducers. Compared to control NIH-3T3 cells, NIH-3T3-Met-EC- cells grew faster and showed anchorage-independent growth and invasive properties in all aspects similar to cells expressing the transforming TPR-MET. Nude female mice injected subcutaneously with NIH-3T3-Met-EC- cells developed visible tumors, displaying the typical morphology of carcinomas with polygonal cells, in contrast to sarcomas with spindle-shaped cells induced by the injection of NIH-3T3-Tpr-Met cells. It is suggested that the different subcellular localization of the oncoproteins, more than differences in signal transduction, could be responsible for the tumor phenotype. All together, these data show that deletion of the ectodomain activates the hepatocyte growth factor receptor and its downstream signaling pathways, unleashing its transforming, invasive, and tumorigenic potential.

Topics: Animals; Cell Line, Tumor; Cell Transformation, Neoplastic; Female; Green Fluorescent Proteins; Immunohistochemistry; Keratins; Lentivirus; MAP Kinase Signaling System; Mice; Mice, Nude; NIH 3T3 Cells; Phosphorylation; Plasmids; Protein Processing, Post-Translational; Protein Structure, Tertiary; Proto-Oncogene Mas; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-met; Sequence Deletion; Transduction, Genetic; Tyrosine; Vimentin; Xenograft Model Antitumor Assays

Ameloblastomatous epithelium containing clusters of ghost cells is the typical histopathology of calcifying cystic odontogenic tumor (CCOT). This paper aimed to assess keratins AE1-AE3, K7, K10/13, K14, K18, K19, vimentin, laminin, and collagen IV in 08 CCOTs to discuss their histopathogenesis. Similarity to the immunoprofile of the stratified squamous epithelium was seen in the with the basal layer expressing K14 and the upper cells expressing K10/13. When compared to the immunoprofile of the normal odontogenic epithelium, of odontogenic tumor epithelia and of the ghost cells described in the literature, it was possible to suggest that the CCOT epithelium differentiates towards squamous type.

Topics: Cell Differentiation; Cell Transformation, Neoplastic; Collagen Type IV; Epithelium; Humans; Immunohistochemistry; Jaw Neoplasms; Keratins; Laminin; Odontogenic Cyst, Calcifying; Vimentin

Inorganic arsenic is a known human skin carcinogen. Chronic arsenic exposure results in various human skin lesions, including hyperkeratosis and squamous cell carcinoma (SCC), both characterized by distorted cytokeratin (CK) production. Prior work shows the human skin keratinocyte HaCaT cell line, when exposed chronically for >25 weeks to a low level of inorganic arsenite (100nM) results in cells able to produce aggressive SCC upon inoculation into nude mice. In the present study, CK expression analysis was performed in arsenic-exposed HaCaT cells during the progressive acquisition of this malignant phenotype (0-20 weeks) to further validate this model as relevant to epidermal carcinogenesis induced by arsenic in humans. Indeed, we observed clear evidence of acquired cancer phenotype by 20 weeks of arsenite exposure including the formation of giant cells, a >4-fold increase in colony formation in soft agar and a approximately 2.5-fold increase in matrix metalloproteinase-9 secretion, an enzyme often secreted by cancer cells to help invade through the local extra-cellular matrix. During this acquired malignant phenotype, various CK genes showed markedly altered expression at the transcript and protein levels in a time-dependent manner. For example, CK1, a marker of hyperkeratosis, increased up to 34-fold during arsenic-induced transformation, while CK13, a marker for dermal cancer progression, increased up to 45-fold. The stem cell marker, CK15, increased up to 7-fold, particularly during the later stages of arsenic exposure, indicating a potential emergence of cancer stem-like cells with arsenic-induced acquired malignant phenotype. The expression of involucrin and loricrin, markers for keratinocyte differentiation, increased up to 9-fold. Thus, during arsenic-induced acquired cancer phenotype in human keratinocytes, dramatic and dynamic alterations in CK expression occur which are consistent with the process of epidermal carcinogenesis helping validate this as an appropriate model for the study of arsenic-induced skin cancer.

Topics: Arsenites; Biomarkers, Tumor; Carcinogens; Cell Line; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; Humans; Keratinocytes; Keratins; Membrane Proteins; Phenotype; Skin Neoplasms; Tumor Stem Cell Assay

Breast cancer is characterized by malignant transformation of epithelial cells, but stromal cells also play an important role in tumorigenesis. While tumor-derived fibroblasts display unique phenotypic properties, it is unclear whether they also represent are a specific subpopulation.. Stromal fibroblasts deriving from malignant tissue of 10 women with invasive breast cancer, and from normal breast tissue of 10 women with benign breast disorders, were subjected to differential complementary DNA Microarray Analysis by using a 2,400 gene cDNA array. Individual gene expression pattern were confirmed by RT-PCR.. In a cDNA array that allows to analyze the differential gene expression of more than 2,400 genes, the mRNA expression of 135 genes were increased more than 2 fold in fibroblasts from malignant breast tumors. The majority of these genes encode tumor-promoting cytokines, transcription factors and cell-matrix associated proteins. The mRNA expression of 110 genes decreased to less than 0.5 fold. The remaining 2,155 genes were not significantly altered. RT-PCR performed on individual biopsies from breast cancer and normal breast tissues confirmed the validity of the pooled gene expression signature.. Breast cancer-derived stromal fibroblasts show a distinctive gene expression pattern that differentiates them from normal breast stroma. Our observation of increased expression of tumor promotion-associated genes even in the absence of adjacent malignant epithelium suggests that tumor stroma is comprised of a fibroblastic subpopulation that provides for a microenvironment which supports tumor growth and invasion.

Topics: Apoptosis; Breast Neoplasms; Cell Transformation, Neoplastic; Epithelial Cells; Fibroblasts; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Keratins; Leukocyte Common Antigens; Nucleic Acid Hybridization; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Vimentin

Carcinomas are widely thought to derive from epithelial cells with malignant progression often associated with an epithelial-mesenchymal transition (EMT). We have characterized tumors generated by spontaneously transformed human mesenchymal cells (TMC) previously obtained in our laboratory. Immunohistopathological analyses identified these tumors as poorly differentiated carcinomas, suggesting that a mesenchymal-epithelial transition (MET) was involved in the generation of TMC. This was corroborated by microarray and protein expression analysis that showed that almost all mesenchymal-related genes were severely repressed in these TMC. Interestingly, TMC also expressed embryonic antigens and were able to integrate into developing blastocysts with no signs of tumor formation, suggesting a dedifferentiation process was associated with the mesenchymal stem cell (MSC) transformation. These findings support the hypothesis that some carcinomas are derived from mesenchymal rather than from epithelial precursors.

Topics: Animals; Carcinoma; Cell Dedifferentiation; Cell Transformation, Neoplastic; Cells, Cultured; Epithelial Cells; Humans; Keratins; Male; Mesenchymal Stem Cells; Mice; RNA, Messenger; Vimentin

Oral submucous fibrosis (OSF) is characterized by abnormal collagen metabolism in the submucosal connective tissue. Its influence on the overlying epithelium is not known but about 14% of OSF cases undergo malignant transformation to squamous cell carcinoma indicating association with abnormality of the epithelium. Here, we have defined the keratin expression profile, by immunohistochemistry and quantitative image analysis, using a panel of 22 anti-keratin monoclonal antibodies on 28 OSF samples. We observed an increase of K1 and K10 in the suprabasal layers, induction of K6 in the basal layer and complete loss of K19 in the epithelium. Furthermore, there was increased K17 expression in the suprabasal layers, which correlated with disease severity. In a subset of the most severe OSF cases (14%), K17 expression was completely lost in the basal layer which might define them to be at most risk to undergo malignant transformation. There was no detectable expression of K8, K18, K7 and K9 and the expression of K4, K13, K14, K15 and K16 did not change in OSF. We propose that the altered keratin profiles could be useful as histological diagnostic markers and provide important insights into the pathogenesis of the disease and its predisposition to malignancy.

Topics: Biomarkers; Carcinoma, Squamous Cell; Case-Control Studies; Cell Transformation, Neoplastic; Gene Expression; Humans; Immunohistochemistry; Keratin-17; Keratinocytes; Keratins; Mouth Neoplasms; Oral Submucous Fibrosis; Phenotype; Photography, Dental; Precancerous Conditions; Severity of Illness Index

Calcifying odontogenic cyst was described first by Gorlin et al. in 1962; since then several hundreds of cases had been reported. In 1981, Praetorius et al. proposed a widely used classification. Afterwards, several authors proposed different classifications and discussed its neoplastic potential. The 2005 WHO Classification of Odontogenic Tumours re-named this entity as calcifying cystic odontogenic tumour (CCOT) and defined the clinico-pathological features of the ghost cell odontogenic tumours, the CCOT, the dentinogenic ghost cell tumour (DGCT) and the ghost cell odontogenic carcinoma (GCOC).. The aim of this paper was to review the clinical-pathological features of 122 CCOT, DGCT and GCOC cases retrieved from the files of the oral pathology laboratories from 14 institutions in Mexico, South Africa, Denmark, the USA, Brazil, Guatemala and Peru. It attempts to clarify and to group the clinico-pathological features of the analysed cases and to propose an objective, comprehensive and useful classification under the 2005 WHO classification guidelines.. CCOT cases were divided into four sub-types: (i) simple cystic; (ii) odontoma associated; (iii) ameloblastomatous proliferating; and (iv) CCOT associated with benign odontogenic tumours other than odontomas. DGCT was separated into a central aggressive DGCT and a peripheral non-aggressive counterpart. For GCOC, three variants were identified. The first reported cases of a recurrent peripheral CCOT and a multiple synchronous, CCOT are included.. Our results suggest that ghost cell odontogenic tumours comprise a heterogeneous group of neoplasms which need further studies to define more precisely their biological behaviour.

Topics: Adolescent; Adult; Age Distribution; Aged; Cell Transformation, Neoplastic; Child; Female; Humans; International Cooperation; Jaw Neoplasms; Keratins; Male; Middle Aged; Neoplasm Recurrence, Local; Odontogenic Cyst, Calcifying; Odontogenic Tumors; Retrospective Studies; Sex Distribution; Tooth, Unerupted

Microglandular adenosis (MGA) of the breast is widely known as a benign lesion that can mimic invasive carcinoma. In situ and invasive carcinomas have been described as arising in MGA, but which cases of MGA will progress to carcinoma is unclear. Criteria for distinguishing uncomplicated MGA, MGA with atypia (AMGA), and carcinoma arising in MGA (MGACA) are not standardized. The primary objective of this study was to illustrate the clinical, histopathologic, and immunophenotypical characteristics of MGA, AMGA, and MGACA in an effort to provide criteria for distinguishing the 3 types. We retrospectively identified 108 cases seen at M.D. Anderson Cancer Center between 1983 and 2007 that had a diagnosis of MGA. Of the 108 cases, 65 cases had available material for review. Inclusion criteria were glands of MGA expressing S-100 protein and lacking myoepithelial layer (smooth muscle actin negative). Eleven out of 65 cases qualified to have an MGA component; myoepithelial layer was detected in the remaining 54 cases and were classified as adenosis. Out of the 11 MGA patients, there were 3 patients with uncomplicated MGA, 2 had AMGA, and 6 had MGACA. Staining indices for the cell cycle markers p53 and Ki-67 were used to compare the 3 tumor categories. Additional staining for other tumor markers [estrogen and progesterone receptors, HER2, epidermal growth factor receptor (EGFR), c-kit, CK5/6, and CK18] were performed. Patient demographics, tumor radiologic features, and clinical follow-up data were collected for all cases. Multiple invasive histologic components were identified in each of the MGACA cases. All invasive MGACAs had a duct-forming component. In addition, basal-like component was present in 2 cases, aciniclike in 2, matrix producing in 4, sarcomatoid in 1, and adenoid cystic in 1. All tumors had strong and diffuse CK8/18 and EGFR expression but no estrogen receptor, progesterone receptor, HER2 (ie, triple negative), or CK5/6 expression. C-kit was focally expressed in 2 of the MGACAs. Ki-67 and p53 labeling indices was < 3% in all MGAs, 5% to 10% in the AMGAs, and > 30% in MGACAs. In a follow-up ranging from 14 days to 8 years, none of the MGA cases recurred. One of the AMGA cases recurred as invasive carcinoma in a background of AMGA after 8 years following incomplete excision of the lesion. Three out of 6 MGACA cases (50%) required multiple consecutive resections ending up with mastectomy due to involved margins by invasive or in situ carcinoma. Two ou

Topics: Actins; Adult; Aged; Biomarkers, Tumor; Breast Neoplasms; Carcinoma; Carcinoma in Situ; Cell Transformation, Neoplastic; Diagnosis, Differential; Diagnostic Errors; Disease Progression; ErbB Receptors; Female; Fibrocystic Breast Disease; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Mastectomy; Middle Aged; Neoplasm Invasiveness; Precancerous Conditions; Proto-Oncogene Proteins c-kit; Receptor, ErbB-2; Receptors, Estrogen; Receptors, Progesterone; Retrospective Studies; S100 Proteins; Texas; Time Factors; Treatment Outcome; Tumor Suppressor Protein p53

Origin of basal cell carcinoma (BCC) is still unclear. We studied the cytokeratin (CK) profile in BCC using monoclonal antibodies against 12 CKs to further investigate the suggested origin of the tumor from follicular matrix cells or from follicular outer root sheath cells and to determine if BCC subtypes can be identified on the basis of their CK profiles. Cases of pilomatricoma and samples of fetal skin served as controls to establish the CK profile in matrical cells and developing follicles during intrauterine life, that of the epidermis and cutaneous adnexa in adult life having been determined in a previous study. The most significant findings were as follows: (a) CK 5 and CK 17 positivity in all the BCCs studied; (b) CK 7, CK 8, CK 18, and CK 19 positivity in 30/52, 33/52, 42/52, and 14/52 BCCs, respectively; (c) CK 14 negativity in almost all the BCCs studied; and (d) lack of CK 1 expression only in 2/2 morpheiform BCCs and 4/10 nodular BCCs. The study suggests a tumorous differentiation toward follicular outer root sheath cells and, in most cases, also toward the glandular components of the pilosebaceous-apocrine unit. No significant difference in the CK profile among the BCC subtypes studied was found.

Topics: Biomarkers, Tumor; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Fetus; Gestational Age; Hair Diseases; Hair Follicle; Humans; Keratinocytes; Keratins; Pilomatrixoma; Skin; Skin Neoplasms

We report a case of primary sarcoma of the skin with a biphasic histological pattern, being composed of areas of mixed mesenchymal-epithelial cell proliferation and areas of purely sarcomatous growth. The tumor occurred in the posterior cervical region of a 93-year-old man, and its history was marked by sudden, rapid enlargement after many years of stable duration. The excised lesion was about 4 cm in diameter, had a firm consistency and was covered by intact skin. Histological examination showed a multifocal proliferation of follicular germinative cells arranged in corymbiform and petaloid shapes with an overall retiform growth pattern. Epithelial cords and strands were composed of cytologically uniform cells with bland nuclear features and were surrounded by a prominent, fibroblast-rich stroma reminiscent of a perifollicular sheath. In many areas of the tumor the stroma showed abrupt transition into a pleomorphic proliferation of large sarcomatous cells with frequent and often atypical mitoses. Multinucleated neoplastic cells infiltrated the epithelial structures to cause their partial or total obliteration in many fields of the lesion. Immunohistochemically, the epithelial cells displayed expression of various keratins, with a particularly intense staining for 34betaE12, and were partly positive for the CD10 antigen. A strong immunostaining for this antigen was also observed in malignant-appearing stromal areas, where no expression of cytokeratins was detected. Moreover, nuclear positivity for p53 protein was seen in sarcomatous cells, whereas it resulted in total lack of epithelial elements. Our case emphasizes that high-grade sarcoma may occur in the spectrum of trichoblastic tumors and that it may share some features of other noncutaneous biphasic neoplasms, such as mammary cystosarcoma phyllodes.

Topics: Aged, 80 and over; Cell Proliferation; Cell Transformation, Neoplastic; Hair Diseases; Hair Follicle; Humans; Keratins; Male; Neprilysin; Sarcoma; Skin Neoplasms; Stromal Cells; Tumor Suppressor Protein p53

Mature cystic teratoma of the ovary (MCTO) is the most common type of ovarian teratoma and also the most frequent tumor originating from germ cells. It is usually diagnosed in early adulthood and, by definition, is composed of well-differentiated tissues, which originate from all three germ cell layers. Unusual types of tissues can be found in MCTO, such as kidney, adrenal, and prostatic tissues. Malignant transformation is reported in less than 2% of teratomas. Squamous cell carcinoma is the most common malignancy arising in these otherwise benign tumors. We present the first case of MCTO containing a chordoma. The chordoma differentiation was supported by immunohistochemical staining and interphase fluorescence in situ hybridization (IP-FISH) technique showing 19% of the nuclei of the MCTO displaying polysomy for the chromosome X, while 28% of the chordoma nuclei showed chromosome 7 mosaicism. These results are concordant with previous studies, showing chromosomal anomalies in chromosomes X and 7 in MCTO and chordomas, respectively.

Topics: Adult; Cell Differentiation; Cell Transformation, Neoplastic; Chordoma; Chromosomes, Human, Pair 7; Chromosomes, Human, X; Diagnosis, Differential; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Keratins; Ki-67 Antigen; Mosaicism; Mucin-1; Ovarian Neoplasms; S100 Proteins; Teratoma; Tumor Suppressor Protein p53; Vimentin

We have studied cytokeratin (CK) expression in two cases of well-differentiated and poorly differentiated squamous cell carcinoma (SCC) arising from hidradenitis suppurativa (HS) (acne inversa). In both cases, type A (infundibular-like keratinized) epithelia were observed. In type A epithelia, CK 1 and 10 expressions were decreased, and CK 14 and 17 were detectable in the whole layers. CK 7, 8, 15, 16 and 18 were not detected in type A epithelia. In tumor nests of well-differentiated SCC, CK 1 and 10 expressions were downregulated, and CK 14 expression was upregulated. In tumor nests of poorly differentiated SCC, CK 1 and 10 were not expressed, but simple epithelial keratins (CK 8, 18 and 19) were expressed. These changes of CK expression are related to malignant transformation from the sinus tract (type A epithelium) in HS to SCC.

Topics: Aged; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Fatal Outcome; Hidradenitis Suppurativa; Humans; Immunoenzyme Techniques; Keratins; Male; Middle Aged; Skin Neoplasms

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Keratins; Kidney Neoplasms; Male; Middle Aged; Sarcoma

Research was initiated to develop an in vitro system to identify disinfection by-products with a potential to transform normal human colonocytes into malignant cells. Tribromomethane and bromochloroacetic acid, rodent colon carcinogens, dibromonitromethane and tribromonitromethane, recently identified in drinking water, and azoxymethane, a classic colon carcinogen, were tested for the ability to transform NCM460 cells. The chronic toxicity was determined for the series of trihalomethanes, haloacetic acids and halonitromethanes as well as NCM460 cell enzymatic capabilities. The order of cytotoxicity was halonitromethanes > haloacetic acids > trihalomethanes. Cytotoxicity within a series increased with the degree of bromination and decreased with the molecular weight. The genotoxicity profile was similar to that for cytotoxicity. Enzymatic analysis demonstrated that NCM460 cells possess glutathione-S transerase-1-1 and CYP450 activity similar to that measured in the large intestine. NCM460 cells were exposed to 10(-6) M of the test chemicals for three days. While NCM460 cells from all treatments had the ability to grow in soft agar to some extent, only cells exposed to azoxymethane or tribromomethane were able to grow in media lacking serum and growth factors. When sub cultured, NCM460 cells exposed to 10(-9) M azoxymethane for three weeks formed colonies with morphology distinct from untreated cells.

Topics: Acetates; Azoxymethane; Carcinogens; Cell Survival; Cell Transformation, Neoplastic; Cells, Cultured; Colon; Cytochrome P-450 Enzyme System; Glutathione Transferase; Humans; Hydrocarbons, Brominated; Inhibitory Concentration 50; Time Factors; Trihalomethanes

Poroid hidradenoma (PH), a less common subtype of poroid neoplasm (PN) than eccrine poroma (EP), has not been immunohistochemically studied before. Six cases of PH (four solitary PH and two PH coexisted with other types of PN) were included in the study. Fifteen cases of EP were also included for comparison. Hematoxylin and eosin, Masson-Zimmerman silver stain, and a variety of immunohistochemical stains were used. Microscopically, PH is not connected to the epidermis. All six PH contained small poroid cells and larger, paler cuticular cells. Some PH showed separate or clusters of sebocytes (2/6), horn cysts (1/6), juxtaposed lymphoid follicles in the stroma (1/6) and foci of keratohyaline granules (2/6), none of which was seen in the 15 EP. Immunohistochemically, the keratin distribution of PH was very similar to that of EP. PH has a very small number of Langerhans cells (significantly lower than the overlying epidermis, P=0.045), and a sparse deposition of melanin. We conclude that except the location, the histopathological and immunochemical differences between PH and EP were small. Sebaceous differentiation in two PH lesions suggested the possibility of an apocrine origin. The deeper parts of eccrine apparatus other than basaloid cells may have been more actively involved in the histogenesis of PH.

Topics: Adenoma, Sweat Gland; Aged; Cell Transformation, Neoplastic; Eccrine Glands; Epidermis; Humans; Immunohistochemistry; Keratins; Male; Melanins; Middle Aged; Skin Neoplasms; Sweat Gland Neoplasms

Notch pathway is crucial for stem/progenitor cell maintenance, growth and differentiation in a variety of tissues. Using a transgenic cell ablation approach, we found in our previous study that cells expressing Notch1 are crucial for prostate early development and re-growth. Here, we further define the role of Notch signaling in regulating prostatic epithelial cell growth and differentiation using biochemical and genetic approaches in ex vivo or in vivo systems. Treatment of developing prostate grown in culture with inhibitors of gamma-secretase/presenilin, which is required for Notch cleavage and activation, caused a robust increase in proliferation of epithelial cells co-expressing cytokeratin 8 and 14, lack of luminal/basal layer segregation and dramatically reduced branching morphogenesis. Using conditional Notch1 gene deletion mouse models, we found that inactivation of Notch1 signaling resulted in profound prostatic alterations, including increased tufting, bridging and enhanced epithelial proliferation. Cells within these lesions co-expressed both luminal and basal cell markers, a feature of prostatic epithelial cells in predifferentiation developmental stages. Microarray analysis revealed that the gene expression in a number of genetic networks was altered following Notch1 gene deletion in prostate. Furthermore, expression of Notch1 and its effector Hey-1 gene in human prostate adenocarcinomas were found significantly down-regulated compared to normal control tissues. Taken together, these data suggest that Notch signaling is critical for normal cell proliferation and differentiation in the prostate, and deregulation of this pathway may facilitate prostatic tumorigenesis.

Topics: Amyloid Precursor Protein Secretases; Animals; Animals, Newborn; Aspartic Acid Endopeptidases; Basic Helix-Loop-Helix Transcription Factors; Cell Cycle Proteins; Cell Differentiation; Cell Proliferation; Cell Transformation, Neoplastic; Cells, Cultured; Down-Regulation; Endopeptidases; Epithelial Cells; Humans; Keratins; Male; Mice; Mice, Knockout; Morphogenesis; Oligonucleotide Array Sequence Analysis; Prostate; Protease Inhibitors; Receptor, Notch1; Stem Cells

Cytokeratins are markers of epithelial cell differentiation useful in determining histogenesis for malignancies with an unknown primary. Application of this principle to a single malignancy may identify cancer subtypes with altered developmental programs. Herein, we investigate the relevance of two widely used cytokeratins (CKs), 7 and 20, to subtype pancreas cancer and identify associations with clinical features.. A tissue microarray was constructed using tumor specimens from 103 patients who underwent resection for pancreatic adenocarcinoma with curative intent. A subset of resection specimens was evaluated for pancreatic intraepithelial neoplasia (PanIN) lesions. Tissues were immunostained by using specific anticytokeratin 7 and 20 monoclonal antibodies.. CK 7 and 20 expression was present in 96% and 63% cases of pancreatic adenocarcinoma, respectively. Ubiquitous CK 7 expression precluded further analysis. Tumoral CK 20 expression was not associated with any histopathologic parameter but correlated with worse prognosis when considered as either a dichotomous (P=0.0098) or continuous (P=0.007) variable. In a multivariate model, tumoral CK 20 expression remained a significant independent prognosticator. CK 20 expression was absent in all PanIN lesions from eight resection specimens in which the tumor component was negative for CK 20. In contrast, presence of tumoral CK 20 was highly concordant with its expression in corresponding PanINs.. CK 20 expression defines a subtype of pancreas cancer with important biologic properties. When present, CK 20 expression is an early event in pancreatic carcinogenesis identifiable in precursor lesions. Further studies to identify the underlying genetic changes associated with this altered developmental pathway are warranted.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Cell Transformation, Neoplastic; Female; Gene Expression Profiling; Humans; Keratin-20; Keratin-7; Keratins; Male; Middle Aged; Oligonucleotide Array Sequence Analysis; Pancreatic Neoplasms; Prognosis; Retrospective Studies; Survival Analysis

Oral submucous fibrosis (OSF) is a pre-malignant condition caused by habitual use of areca nut, affecting the oro-pharynx and characterized by progressive fibrosis. Alteration of cytokeratin (CK) expression has been documented in leukoplakia and oral cancer (OC). However, very little is known of CK alterations in OSF. The present study was carried out to characterize the CK profile in OSF and ascertain if this could be used as a surrogate marker for malignant transformation.. Paraffin-embedded tissues of OSF (n = 50), normal (n = 10) and OC (n = 10) were stained with pancytokeratin (PanCK), high molecular weight cytokeratin (HMWCK), CKs 18, 14, 8, 5, 4 and 1 by immunohistochemistry.. Significant difference in the CK staining pattern was seen between normal, OSF and cancer. Significant changes in OSF included increased intensity of staining for PanCK and HMWCK, aberrant expression of CK8 and decreased expression of CKs 5 and 14.. Cytokeratin profile of OSF was significantly different from normals for PanCK, HMWCK, CK8, 5 and 14 suggesting their potential to be used as surrogate markers of malignant transformation. Further studies will help in better defining the nature and clinical implications of these alterations.

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Coloring Agents; Female; Humans; Immunohistochemistry; Keratins; Male; Molecular Weight; Mouth Neoplasms; Oral Submucous Fibrosis; Precancerous Conditions

To determine how different epithelial cell types respond to the same oncogenic stimulation, we have used a modified human keratin 18 gene to conditionally express the polyomavirus middle T antigen (PyMT) oncogene in simple epithelial tissues of transgenic mice. Activation of PyMT expression by transgenic Cre recombinase in mammary epithelial cells resulted in carcinomas in all bitransgenic females. PyMT expression induced by K18-driven Cre in internal epithelial organs resulted in pancreatic acinar metaplasia and ductal dysplasia with remarkable desmoplastic stromal responses in all 25 bitransgenic mice. Hepatoma formation with altered lipid metabolism and gastric adenocarcinoma occurred in 96 and 54% of these mice, respectively. Elevated PyMT RNA expression also correlated with intraepithelial neoplasia in the prostate. Activated Erk2 was found in mammary tumors, pancreatic tissues, and affected livers. Hes1 RNA, a target of Notch signaling that has been implicated downstream of Ras pathway activation, was elevated in pancreatic and liver lesions. The variety of responses of different epithelia to PyMT demonstrates the importance of the differentiated state in interpreting oncogenic signals.

Topics: Animals; Antigens, Polyomavirus Transforming; Basic Helix-Loop-Helix Transcription Factors; Blotting, Southern; Cell Transformation, Neoplastic; Epithelial Cells; Extracellular Signal-Regulated MAP Kinases; Female; Homeodomain Proteins; Humans; Keratin-18; Keratins; Mice; Mice, Transgenic; Oncogenes; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription Factor HES-1

Breast cancer is thought to arise in mammary epithelial stem cells. There is, therefore, a large amount of interest in identifying these cells. The breast is a complex tissue consisting of two epithelial layers (an outer myoepithelial/basal layer and an inner luminal epithelial layer) as well as a large non-epithelial component (fibroblasts, endothelial cells, lymphocytes, adipocytes, neurons and myocytes). The definitive identification of a mammary epithelial stem cell population is critically dependent on its purity. To date, this has been hampered by the lack of suitable markers to separate out the two epithelial layers, and to remove contaminating non-epithelial cells.. Mouse mammary glands were dissociated and stained with CD24. Cells were sorted into separate populations based on CD24 expression and assessed for luminal epithelial and myoepithelial/basal markers by direct fluorescent microscopy and real time PCR. The stem/progenitor potential of these cell populations was assessed in vivo by cleared mammary fat pad transplantation.. Three populations of CD24 expressing cells were identified: CD24Negative, CD24Low and CD24High. Staining of these cells with cytokeratin markers revealed that these populations correspond to non-epithelial, myoepithelial/basal and luminal epithelial cells, respectively. Cell identities were confirmed by quantitative PCR. Cleared mammary fat pad transplantation of these cell populations revealed that extensive mammary fat pad repopulation capacity segregates with the CD24Low cells, whilst CD24High cells have limited repopulation capacity.. Differential staining of mammary epithelial cells for CD24 can be used to simultaneously isolate pure populations of non-epithelial, myoepithelial/basal and luminal epithelial cells. Furthermore, mammary fat pad repopulation capacity is enriched in the CD24Low population. As separation is achieved using a single marker, it will be possible to incorporate additional markers to further subdivide these populations. This will considerably facilitate the further analysis of mammary epithelial subpopulations, whilst ensuring high purity, which is key for understanding mammary epithelial stem cells in normal tissue biology and carcinogenesis.

Topics: Adipose Tissue; Animals; Biomarkers; CD24 Antigen; Cell Transformation, Neoplastic; Epithelial Cells; Female; Keratins; Mammary Glands, Animal; Mice; Microscopy, Fluorescence; Polymerase Chain Reaction; Staining and Labeling; Stem Cells

PTEN tumor suppressor gene failure in ras(Ha)-activated skin carcinogenesis was investigated by mating exon 5 floxed-PTEN (Delta5PTEN) mice to HK1.ras mice that expressed a RU486-inducible cre recombinase (K14.creP). PTEN inactivation in K14.cre/PTEN(flx/flx) keratinocytes resulted in epidermal hyperplasia/hyperkeratosis and novel 12-O-tetradecanoylphorbol-13-acetate (TPA)-promoted papillomas, whereas HK1.ras/K14.cre/PTEN(flx/flx) cohorts displayed a rapid onset of papillomatogenesis due to a synergism of increased AKT activity and extracellular signal-regulated kinase (ERK) elevation. High 5-bromo-4-deoxyuridine labeling in Delta5PTEN papillomas showed that a second promotion mechanism centered on failures in cell cycle control. Elevated cyclin D1 was associated with both HK1.ras/ERK- and Delta5PTEN-mediated AKT signaling, whereas cyclin E2 overexpression seemed dependent on PTEN loss. Spontaneous HK1.ras/Delta5PTEN malignant conversion was rare, whereas TPA promotion resulted in conversion with high frequency. On comparison with all previous HK1.ras carcinomas, such TPA-induced carcinomas expressed atypical retention of keratin K1 and lack of K13, a unique marker profile exhibited by TPA-induced K14.cre/PTEN(flx/flx) papillomas that also lacked endogenous c-ras(Ha) activation. Moreover, in all PTEN-null tumors, levels of ras(Ha)-associated total ERK protein became reduced, whereas phosphorylated ERK and cyclin D1 were lowered in late-stage papillomas returning to elevated levels, alongside increased cyclin E2 expression, in TPA-derived carcinomas. Thus, during early papillomatogenesis, PTEN loss promotes ras(Ha) initiation via elevation of AKT activity and synergistic failures in cyclin regulation. However, in progression, reduced ras(Ha)-associated ERK protein and activity, increased Delta5PTEN-associated cyclin E2 expression, and unique K1/K13 profiles following TPA treatment suggest that PTEN loss, rather than ras(Ha) activation, gives rise to a population of cells with greater malignant potential.

Topics: Animals; Carcinogens; Cell Differentiation; Cell Transformation, Neoplastic; Keratin-13; Keratins; MAP Kinase Signaling System; Mice; Mice, Transgenic; Mifepristone; Oncogene Protein v-akt; Papilloma; PTEN Phosphohydrolase; ras Proteins; Skin Neoplasms; Tetradecanoylphorbol Acetate; Up-Regulation

Screening of head and neck carcinoma patients with the proteomics-based AMIDA technology yielded a set of tumour-associated antigens, including the intermediate filament protein cytokeratin 8 (CK8). The expression pattern and specificity of CK8 was compared with those of the established markers pan-cytokeratins and CK13, and with that of the proliferation marker Ki67. Expression of CK8 correlated positively with malignancies of the head and neck areas. CK8 was not expressed in healthy epithelium, except for some rare cases of cells of the basal layer and laryngeal tissue. In contrast, the vast majority of head and neck squamous cell carcinomas and metastases strongly expressed CK8. Interestingly, CK8 de novo expression correlated with dysplastic areas of oral leukoplakic lesions, while hyperplastic leukoplakia remained CK8-negative but strongly panCK and CK13 positive. Thus, CK8 is an attractive marker molecule for a differentiated diagnosis of leukoplakia and head and neck carcinomas, which possesses notedly improved specificity as compared with panCK and CK13.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Head and Neck Neoplasms; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Leukoplakia, Oral

Metastatic tumors to the ovary are infrequently of urinary tract origin. In approximate descending order of frequency, this subset of secondary ovarian neoplasms includes renal cell carcinoma, transitional cell carcinoma of the urinary bladder, and urachal adenocarcinomas. These tumors usually raise a differential in turn of primary ovarian clear cell, transitional cell, or mucinous carcinomas. Only rare metastatic signet-ring adenocarcinomas of the bladder have shown the features of a Krukenberg tumor. We report the case of a 74-year old woman with bilateral Krukenberg tumors metastatic from a primary renal pelvic transitional cell carcinoma with glandular and signet-ring cell differentiation. This unique case reinforces that tumors with signet-ring cell morphology have a propensity to metastasize to the ovary, and indicates that renal pelvic carcinoma rarely may be the source of Krukenberg tumors.

Topics: Aged; Carcinoma, Signet Ring Cell; Carcinoma, Transitional Cell; Cell Differentiation; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratin-20; Keratin-7; Keratins; Krukenberg Tumor; Ovarian Neoplasms; Prevalence; Urologic Neoplasms

Ultraviolet A (UVA, 315-400 nm), constituting about 95% of ultraviolet irradiation in natural sunlight, represents a major environmental challenge to the skin and is clearly associated with human skin cancer. It has proven difficult to show direct actions of UVA as a carcinogen in human cells. Here, we demonstrate that chronic UVA exposures at environmentally relevant doses in vitro can induce malignant transformation of human keratinocytes associated with acquired apoptotic resistance. As evidence of carcinogenic transformation, UVA-long-treated (24 J/cm(2) once/week for 18 weeks) HaCaT (ULTH) cells showed increased secretion of matrix metalloproteinase (MMP-9), overexpression of keratin 13, altered morphology and anchorage-independent growth. Malignant transformation was established by the production of aggressive squamous cell carcinomas after inoculation of ULTH cells into nude mice (NC(r)-nu). ULTH cells were resistant to apoptosis induced not only by UVA but also by UVB and arsenite, two other human skin carcinogens. ULTH cells also became resistant to apoptosis induced by etoposide, staurosporine and doxorubicin hydrochloride. Elevated phosphorylation of protein kinase B (PKB, also called AKT) and reduced expression of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) were detected in ULTH cells. The resistance of ULTH cells to UVA-induced apoptosis was reversed by either inhibition of phosphatidylinositol 3-kinase (PI-3K) or adenovirus expression of PTEN or dominant negative AKT. These data indicate that UVA has carcinogenic potential in human keratinocytes and that the increased AKT signaling and decreased PTEN expression may contribute to this malignant transformation. Further comparisons between the transformed ULTH and control cells should lead to a better understanding of the mechanism of UVA carcinogenesis and may help identify biomarkers for UVA-induced skin malignancies.

Topics: Animals; Apoptosis; Arsenites; Carcinogenicity Tests; Cell Transformation, Neoplastic; Cells, Cultured; Doxorubicin; Etoposide; Humans; Keratin-13; Keratinocytes; Keratins; Mice; Mice, Nude; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Signal Transduction; Staurosporine; Ultraviolet Rays

Gastrointestinal adenocarcinoma arising in mature cystic teratomas of the ovary is extremely rare. We report a case of well-differentiated intestinal adenocarcinoma arising in a mature cystic teratoma of the ovary in a 77-year-old woman, presenting as acute abdomen with ovarian torsion. An immunohistochemical study revealed expression of CK20 and CK7, and the tumor was also positive for MUC2. The patient had no evidence of disease after 12 months of follow-up.

Topics: Adenocarcinoma; Aged; Biomarkers, Tumor; Cell Transformation, Neoplastic; Female; Humans; Intestinal Neoplasms; Keratin-20; Keratin-7; Keratins; Mucin-2; Mucins; Neoplasms, Multiple Primary; Ovarian Neoplasms; Teratoma; Tomography, X-Ray Computed; Treatment Outcome

The genetic bases underlying esophageal tumorigenesis are poorly understood. Our previous studies have shown that coordinated deletion of the Smad4 and PTEN genes results in accelerated hair loss and skin tumor formation in mice. Herein, we exemplify that the concomitant inactivation of Smad4 and PTEN accelerates spontaneous forestomach carcinogenesis at complete penetrance during the first 2 months of age. All of the forestomach tumors were invasive squamous cell carcinomas (SCCs), which recapitulated the natural history and pathologic features of human esophageal SCCs. A small population of the SCC lesions was accompanied by adenocarcinomas at the adjacent submucosa region in the double mutant mice. The rapid progression of forestomach tumor formation in the Smad4 and PTEN double knockout mice corresponded to a dramatic increase in esophageal and forestomach epithelial proliferation. The decreased expression of p27, p21, and p16 together with the overexpression of cyclin D1 contributed cooperatively to the accelerated forestomach tumorigenesis in the double mutant mice. Our results point strongly to the crucial relevance of synergy between Smad4 and PTEN to suppress forestomach tumorigenesis through the cooperative induction of cell cycle inhibitors.

Topics: Animals; Carcinoma, Squamous Cell; Cell Growth Processes; Cell Transformation, Neoplastic; Cyclin D; Cyclins; Down-Regulation; Esophageal Neoplasms; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Keratins; Mice; Mice, Knockout; Mice, Transgenic; Mutation; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Smad4 Protein; Stomach Neoplasms

The formation of a simple columnar epithelium in the uterus is essential for implantation. Perturbation of this developmental process by exogenous estrogen, such as diethylstilbestrol (DES), results in uterine metaplasia that contributes to infertility. The cellular and molecular mechanism underlying this transformation event is not well understood. Here we use a combination of global gene expression analysis and a knockout mouse model to delineate genetic pathways affected by DES. Global gene expression profiling experiment revealed that neonatal DES treatment alters uterine cell fate, particularly in the luminal epithelium by inducing abnormal differentiation, characterized by the induction of stratified epithelial markers including members of the small proline-rich protein family and epidermal keratins. We show that Msx2, a homeodomain transcription factor, functions downstream of DES and is required for the proper expression of several genes in the uterine epithelium including Wnt7a, PLAP, and K2.16. Finally, Msx2-/- uteri were found to exhibit abnormal water trafficking upon DES exposure, demonstrating the importance of Msx2 in tissue responsiveness to estrogen exposure. Together, these results indicate that developmental exposure to DES can perturb normal uterine development by affecting genetic pathways governing uterine differentiation.

Topics: Alkaline Phosphatase; Animals; Apoptosis; Cell Differentiation; Cell Lineage; Cell Proliferation; Cell Transformation, Neoplastic; Diethylstilbestrol; DNA Primers; DNA-Binding Proteins; DNA, Complementary; Epithelium; Estrogens, Non-Steroidal; Female; Homeodomain Proteins; In Situ Hybridization; Infertility; Keratins; Metaplasia; Mice; Mice, Knockout; Mice, Transgenic; Models, Biological; Mutation; Oligonucleotide Array Sequence Analysis; Pregnancy; Pregnancy, Animal; Prenatal Exposure Delayed Effects; Proto-Oncogene Proteins; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleases; Time Factors; Transcription, Genetic; Up-Regulation; Uterus; Wnt Proteins; Wnt-5a Protein

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epithelial Cells; Female; Humans; Keratins; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Topics: Cell Transformation, Neoplastic; Humans; Keratins; Loss of Heterozygosity; Odontogenic Cysts; Patched Receptors; Receptors, Cell Surface; Signal Transduction; Veratrum Alkaloids

This study aimed to investigate the role of underlying fibroblasts on morphogenesis of in vitro epithelium reconstituted with normal and neoplastic human oral keratinocytes at various stages of malignant transformation. Primary normal human oral keratinocytes (NOKs), early neoplastic/dysplastic human oral keratinocytes (DOK cell line), and neoplastic human oral keratinocytes (PE/CA-PJ 15 cell line) were organotypically grown on top of a collagen type I matrix with or without primary normal human oral fibroblasts. Morphogenesis of the reconstituted epithelia was assessed by histomorphometry, immunohistochemistry (Ki-67, cyclin D1, cytokeratin 13 (CK13), collagen IV, E-cadherin, p53, CD40), and the terminal deoxynucleotidyl transferase-mediated dUTP in situ nick end-labelling method. Reproducible in vitro models of multistage oral carcinogenesis were established. Presence of fibroblasts in the collagen matrix significantly increased cell proliferation in all three models (p<0.05), and induced an invasive pattern of growth in the neoplastic cell lines (p<0.05). In normal, but not in neoplastic oral keratinocytes fibroblasts induced the expression of CD40, and polarized the expression of E-cadherin and p53 to the basal cell layer. In both normal and early neoplastic keratinocytes (DOK cell line), fibroblasts induced the expression of CK13 and collagen IV. In the neoplastic oral keratinocytes (PE/CA-PJ 15 cell line), the presence of underlying fibroblasts did not change the expression of any of the protein markers assessed. This study showed that (1) major steps of oral carcinogenesis can be reproduced in vitro, and (2) the tight control exerted by fibroblasts on epithelial morphogenesis of in vitro reconstituted normal human oral mucosa is gradually lost during neoplastic progression.

Topics: Biomarkers; Cadherins; Carcinoma; Cell Differentiation; Cell Transformation, Neoplastic; Collagen Type IV; Epithelium; Fibroblasts; Humans; Keratinocytes; Keratins; Morphogenesis; Mouth Mucosa; Mouth Neoplasms

The vitamin K analog menadione (K3), capable of both redox cycling and arylating nucleophilic substrates by Michael addition, has been extensively studied as a model stress-inducing quinone in both cell culture and animal model systems. Exposure of keratin 8 (k-8) expressing human breast cancer cells (MCF7, T47D, SKBr3) to K3 (50-100 microM) induced rapid, sustained, and site-specific k-8 serine phosphorylation (pSer73) dependent on signaling by a single mitogen activated protein kinase (MAPK) pathway, MEK1/2. Normal nuclear morphology and k-8 immunofluorescence coupled with the lack of DNA laddering or other features of apoptosis indicated that K3-induced cytotoxicity, evident within 4 h of treatment and delayed but not prevented by MEK1/2 inhibition, was due to a form of stress-activated cell death known as oncosis. Independent of MAPK signaling was the progressive appearance of K3-induced cellular fluorescence, principally nuclear in origin and suggested by in vitro fluorimetry to have been caused by K3 thiol arylation. Imaging by UV transillumination of protein gels containing nuclear extracts from K3-treated cells revealed a prominent 17-kDa band shown to be histone H3 by immunoblotting and mass spectrometry (MS). K3 arylation of histones in vitro followed by electrospray ionization-tandem MS analyses identified the unique Cys110 residue within H3, exposed only in the open chromatin of transcriptionally active genes, as a K3 arylation target. These findings delineate new pathways associated with K3-induced stress and suggest a potentially novel role for H3 Cys110 as a nuclear stress sensor.

Topics: Breast Neoplasms; Cell Line, Tumor; Cell Transformation, Neoplastic; Histones; Humans; Keratin-8; Keratins; Mitogen-Activated Protein Kinases; Phosphorylation; Spectrometry, Fluorescence; Spectrometry, Mass, Electrospray Ionization; Sulfhydryl Compounds; Vitamin K 3

Topics: Cell Transformation, Neoplastic; Cervix Uteri; Endometrial Hyperplasia; Female; Humans; Keratins

Topics: Cell Transformation, Neoplastic; Cervix Uteri; Endometrial Hyperplasia; Female; Humans; Keratins; Uterine Cervical Dysplasia

Cytokeratins (CKs) are the intermediate filament proteins of the epithelium cells, which have become important markers of normal and abnormal cell differentiation. The goal of the present study was to investigate the expression pattern of CK 10, 13, 14 and 16 in oral verrucous carcinoma (OVC) and oral squamous papilloma (OSP).. Formalin-fixed paraffin-embedded sections from eight cases of each lesion were assessed. Immunohistochemistry was carried out using streptoavidin-biotin complex method.. In OVC, CK 10 was expressed in suprabasal to superficial layers whereas in OSP mainly in superficial layer. CK 13 was detected in prickle and superficial cells in most cases of OVC and in suprabasal to superficial cells of OSP. All the cell layers of OVC reacted positively for CK 14 while basal and suprabasal layers of OSP were more pronounced for CK 14. Finally, CK 16 was observed in suprabasal to superficial layer in OVC and the majority cases in OSP showed only superficial reactive cells.. CK 10, 13, 14 and 16 immunohistochemical profile emphasis the biological behavior of the studied lesions and confirm the use of these proteins as markers of differentiation.

Topics: Carcinoma, Verrucous; Cell Proliferation; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Intermediate Filament Proteins; Keratinocytes; Keratins; Mouth Neoplasms; Papilloma

The entity of serrated adenoma of the colorectum was first proposed in 1990, and it was characterized as epithelial neoplasia combining the architectural features of a hyperplastic polyp with the cytological features of an adenoma. Over the past few years, various clinicopathological studies on serrated adenoma have been reported, but its histogenesis remains unclear. Recently the existence of a "serrated neoplasia pathway" leading to malignancy, which is different from the so-called adenoma-carcinoma sequence, has been discussed. Yao et al. reported that hyperplastic polyps and serrated adenomas share a common cell lineage with gastric differentiation. To clarify the existence of the serrated neoplasia pathway, we performed immunohistochemical staining of cytokeratin 7 (CK7) and cytokeratin 20 (CK20), which are commonly used to determine the primary site of a metastatic lesion, and we examined the pattern of CK7/CK20 expression in various colorectal lesions including 44 serrated adenomas, 25 hyperplastic polyps, 20 traditional adenomas, and 48 carcinomas. An obvious difference existed in the pattern of CK7/CK20 expression between the serrated lesions (hyperplastic polyps and serrated adenomas) and others. The majority of serrated adenomas and hyperplastic polyps presented a CK7+/CK20+ pattern, whereas most conventional adenomas and adenocarcinomas expressed CK7-/CK20+. Adenocarcinoma developing in serrated adenoma also presented a CK7+/CK20+ pattern. There are several reports that CK7 is a possible marker of transient dedifferentiation in the gastric carcinogenesis process. Taken together with the present results, a distinct pathway of colorectal carcinogenesis must exist, which is different from the adenoma-carcinoma sequence. CK7 is a possible marker for the serrated neoplasia pathway of colorectal carcinogenesis.

Topics: Adenocarcinoma; Adenoma; Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Cell Transformation, Neoplastic; Colonic Polyps; Colorectal Neoplasms; Female; Gene Expression Profiling; Humans; Immunohistochemistry; Intermediate Filament Proteins; Keratin-20; Keratin-7; Keratins; Male; Middle Aged

Activated oncogenes have been detected in a variety of malignant tumors and altered expressions of certain genes are known to play a functional role in the cancer process. The chemical carcinogen, BP, and the insertion of c-Ha-ras, induced characteristics of transformed phenotypes in a suitable human breast epithelial cell line. Carcinogen-treated and Ha-ras-transfected cells showed a progression of changes in the morphology, anchorage independent growth, invasiveness and tumorigenicity in SCID mice. Tumor growth occurs after a series of molecular events that parallel morphological changes. The aim of this work was to determine the neoplastic phenotypes following treatment with benzo(a)pyrene (BP) and transfection with c-Ha-ras oncogene changes and PCNA, Neu, ErbB-3 and Cytokeratin 18 protein expression in MCF-10F cells, a spontaneously immortalized human breast epithelial cell line. Protein expression was determined by immunofluorescent staining coupled with confocal microscopy. An increased oncoprotein expression in comparison to MCF-10F cells was observed in PCNA, Neu, ErbB-3 and Cytokeratin 18 protein expression in breast epithelial cells transformed with a chemical carcinogen and/or oncogene transfected that are not present in the MCF-10F. This in vitro cancer model can be used as a valuable model in the study of breast carcinogenesis.

Topics: Benzo(a)pyrene; Breast Neoplasms; Carcinogens; Cell Differentiation; Cell Line, Transformed; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Genes, ras; Humans; In Vitro Techniques; Keratins; Microscopy, Confocal; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Microscopy, Fluorescence; Neoplasms; Oncogene Proteins; Phenotype; Proto-Oncogene Proteins p21(ras)

Medullary breast cancer (MBC) is a rare, diagnostically difficult, pathological subtype. Despite being high grade, it has a good prognosis. MBC patients have an excess of BRCA1 germ-line mutation and reliable identification of MBC could help to identify patients at risk of carrying germline BRCA1 mutations or in whom chemotherapy could be avoided. The aim of this study was therefore to improve diagnosis by establishing an MBC protein expression profile using immunohistochemistry (IHC) on tissue-microarrays (TMA). Using a series of 779 breast carcinomas ('EC' set), diagnosed initially as MBC, a double-reading session was carried out by several pathologists on all of the histological material to establish the diagnosis as firmly as possible using a 'medullary score'. Only MBCs with high scores, i.e. typical MBC (TMBC) (n=44) and non-TMBC grade III with no or low scores (n=160), were included in the IHC study. To validate the results obtained on this first set, a control series of TMBC (n=17) and non-MBC grade III cases (n=140) ('IPC' set) was studied. The expression of 18 proteins was studied in the 61 TMBCs and 300 grade III cases from the two sets. The global intra-observer concordance of the first reading for the diagnosis of TMBC was 94%, with almost perfect kappa (kappa) of 0.815. TMBC was characterized by a high degree of basal/myoepithelial differentiation. In multivariate analysis with logistic regression, TMBC was defined by the association of P-cadherin (R=2.29), MIB1 > 50 (R=3.80), ERBB2 negativity (R=2.24) and p53 positivity (RR=1.45).

Topics: Breast Neoplasms; Cadherins; Carcinoma, Basal Cell; Carcinoma, Medullary; Cell Transformation, Neoplastic; Female; Gene Expression Regulation, Neoplastic; Genes, BRCA1; Genes, erbB-2; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Mutation; Neoplasm Proteins; Phenotype; Protein Array Analysis; Tumor Suppressor Protein p53

The keratin filament network is an important part of the cytoskeleton. It is involved in the regulation of shape and viscoelasticity of epithelial cells. The morphology of keratin networks depends on post-translational modifications of keratin monomers. In-vitro studies indicated that network characteristics, such as filament crosslink density, determines the biophysical properties of the filament network. This report presents a quantitative method for the morphological analysis of keratin filament networks. Visualization of filaments was based on prefixation extraction of epithelial cells and scanning electron microscopy (SEM). SEM images were processed by a skeletonization algorithm to obtain a graph structure that represents individual filaments as well as their connections. This method was applied to investigate the effects of transforming growth factor alpha (TGFalpha) on the morphology of keratin networks in pancreatic cancer cells. TGFalpha contributes to pancreatic cancer progression and activates signalling pathways phosphorylating keratin monomers. Using this new method, a significant alteration to the keratin network morphology could be detected in response to TGFalpha.

Topics: Cell Transformation, Neoplastic; Humans; Intermediate Filaments; Keratins; Microscopy, Electron, Scanning; Tumor Cells, Cultured

The interplay among nucleotide excision repair, cell-cycle regulation, and apoptosis in the UV-exposed epidermis is extremely important to avoid mutations and malignant transformation. In Xpc(-/-) mice deficient in global genome nucleotide excision repair (GGR), a cell-cycle arrest of epidermal cells in late S-phase [with near-double normal diploid (4N) DNA content] was observed 48-72 h after UV exposure. This arrest resolved without apoptosis (96-168 h). We surmised that these arrested keratinocytes with persistent DNA damage were removed by epidermal turnover. In vivo BrdUrd pulse-chase labeling (>17 h after UV exposure) showed that DNA replication after UV exposure was resumed in Xpc(-/-) mice, but it did not reveal any evidence of retained BrdUrd-labeled S-phase cells in the basal layer of the epidermis at 72 h. Interestingly, by this time a maximum number of cytokeratin 10-negative and cytokeratin 5-positive cells had appeared in the suprabasal epidermal cell layers of UV-exposed Xpc(-/-) mice. Accumulation of these "basal cell"-like keratinocytes in the suprabasal layers was clearly aberrant and was not observed in WT and heterozygous mice. Flow cytometric analyses of single-cell suspensions from UV-exposed Xpc(-/-) epidermis further showed that the "near-4N" arrested cells retained cytokeratin 5 and lacked cytokeratin 10. Hence, we conclude that the arrested near-4N cells became detached from the basal layer without entering a proper differentiation program and were indeed subsequently lost through the epidermal turnover. This expulsion apparently constitutes an alternative route, different from in situ apoptosis, to eliminate DNA-damaged arrested cells from the epidermis.

Topics: Animals; Apoptosis; Bromodeoxyuridine; Cell Differentiation; Cell Transformation, Neoplastic; DNA; DNA Damage; DNA Repair; DNA-Binding Proteins; Epidermis; Flow Cytometry; Heterozygote; Immunohistochemistry; Keratinocytes; Keratins; Mice; Mice, Transgenic; Models, Biological; Mutation; Time Factors; Transgenes; Ultraviolet Rays

To clarify whether hepatocellular carcinoma (HCC) originates from hepatic progenitor cells and whether there is any correlation with the clinicopathologic factors of HCC, we reviewed 217 resected HCC specimens.. Immunohistochemical examination of cytokeratin (CK) 7, CK19, CD34, and CD117 (c-KIT) was performed. Overexpression of CK7 and CK19 indicates differentiation from cholangiocellular and hepatic progenitor cells, while overexpression of CD34 and CD117 indicates hepatic stem cells. Fresh specimens were obtained from 20 HCC patients for mutation of the c-KIT gene.. CK7, CK19, and CD117 were positive in 41, 9.7, and 0.9% of the HCC specimens, respectively, and CD34 was never positive. None of the fresh HCC specimens demonstrated a c-KIT mutation. CK19 positivity was significantly correlated with a positive hepatitis B core antibody, and with poor survival outcome, and tended to correlate with poor histologic differentiation.. These results suggest that: (i) about 10% of HCCs with typical histologic features originate from an intermediate hepatic progenitor cell, such as the canal of Hering and oval cells in the rat, or acquire the characteristics of cholangiocellular epithelium by metaplasia; (ii) HCC with typical histologic features rarely originates from hepatic stem cells, and (iii) patients with CK19-positive HCC have a poor prognosis.

Topics: Analysis of Variance; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Chi-Square Distribution; Female; Hepatocytes; Humans; Immunohistochemistry; Keratins; Liver Neoplasms; Male; Middle Aged; Prognosis; Risk Factors; Statistics, Nonparametric; Stem Cells; Survival Rate

Topics: Adenolymphoma; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Keratins; Ki-67 Antigen; Male; Metaplasia; Middle Aged; Parotid Neoplasms

We present a case of adenocarcinoma developing at the vesicocutaneous edge of a vesicostomy, 40 years after it was created, in a patient who underwent cadaveric kidney transplant. Although transitional and squamous cell carcinoma of a vesicostomy have been reported, to our knowledge, the presence of adenocarcinoma at the vesicostomy edge has not been reported previously.

Topics: Abnormalities, Multiple; Adenocarcinoma; Adult; Biomarkers, Tumor; Cell Transformation, Neoplastic; Cystostomy; Dermatologic Surgical Procedures; Female; Humans; Immunosuppression Therapy; Intermediate Filament Proteins; Keratin-20; Keratin-7; Keratins; Kidney Failure, Chronic; Kidney Transplantation; Neoplasm Proteins; Postoperative Complications; Surgical Stomas; Time Factors; Urinary Bladder Neoplasms; Urinary Bladder, Neurogenic; Urogenital Abnormalities; Urothelium

The cytokeratin (CK) pair 8 and 18 is normally expressed in all simple epithelia. This pair is not normally seen in stratified epithelial cells. Squamous cell carcinomas derived from stratified epithelia show anomalous expression of this CK pair. It is not known whether CKs 8 and 18 in any way contribute to the malignant phenotype of these cells. We used an immortalised, nontransformed human foetal buccal mucosa (FBM) cell line that expresses significantly higher amounts of CK18 compared to CK8. FBM cells were transfected with the full-length CK8 gene to study the role of CKs 8 and 18 in malignant transformation. Clones with higher expression of CK8 compared to untransfected FBM cells were studied for changes in their phenotypic characteristics. Immunofluorescence studies using antibodies to CKs 8 and 18 revealed well-decorated filaments throughout the cytoplasm in CK8 gene-transfected cells vs. untransfected FBM cells. Transmission images showed that FBM cells were isolated while transfected cells were in groups of well-spread cells with cellular projections. Transfected cells were independent of growth supplement requirements and showed anchorage-independent growth in soft agar assay and significantly reduced doubling time compared to nontransfected FBM cells. DNA flow-cytometric studies revealed increased DNA content and prolonged S phase in transfected clones vs. FBM cells. Injection of cells s.c. obtained from soft agar colonies developed from 2 of the clones resulted in tumour formation at the site of injection. In both cases, lung metastasis was also seen. Thus, in conclusion, it appears that increased expression of CK8 in some way changes the phenotypic characteristics of stratified epithelial cells, resulting in malignant transformation.

Topics: Animals; Carcinoma, Squamous Cell; Cell Cycle; Cell Division; Cell Line; Cell Transformation, Neoplastic; DNA; Epithelial Cells; Flow Cytometry; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Humans; Keratins; Mice; Mice, Nude; Mouth Mucosa; Mouth Neoplasms; Neoplasms, Experimental; Phenotype; Transfection; Transplantation, Heterologous

In this study, we examined the distribution of heparanase protein in 75 esophageal squamous cell carcinomas by immunohistochemistry and analyzed the relationship between heparanase expression and clinicopathological characteristics. In situ hybridization showed that the mRNA expression pattern of heparanase was similar to that of the protein, suggesting that increased expression of the heparanase protein at the invasive front was caused by an increase of heparanase mRNA in tumor cells. Heparanase expression correlated significantly with depth of tumor invasion, lymph node metastasis, tumor node metastasis (TNM) stage and lymphatic invasion. Overexpression of heparanase in esophageal cancers was also associated with poor survival. In addition to its localization in the cytoplasm and cell membrane, heparanase was also identified in the nuclei of normal epithelial and tumor cells by immunohistochemistry. Furthermore, nuclear heparanase was detected in nuclear extract of cancer cell lines by Western blot and immunohistochemistry. Examination of the role of nuclear heparanase in cell proliferation and differentiation by double immunostaining for proliferating cell nuclear antigen (PCNA) and cytokeratin 10 (CK10) showed significant relationship between nuclear heparanase expression and differentiation (heparanase vs CK10), but not for proliferative state of esophageal cancer cells (heparanase vs PCNA). Our results suggest that cytoplasmic heparanase appears to be a useful prognostic marker in patients with esophageal cancer and that nuclear heparanase protein may play a role in differentiation. Inhibition of heparanase activity may be effective in the control of esophageal tumor invasion and metastasis.

Topics: Adult; Aged; Biomarkers, Tumor; Blotting, Western; Carcinoma, Squamous Cell; Cell Differentiation; Cell Division; Cell Line, Tumor; Cell Transformation, Neoplastic; Esophageal Neoplasms; Female; Fluorescent Antibody Technique, Indirect; Glucuronidase; Humans; Immunoenzyme Techniques; In Situ Hybridization; Keratins; Male; Middle Aged; Prognosis; Proliferating Cell Nuclear Antigen; RNA, Messenger; Survival Rate

Keratins K8 and K18 are the major components of the intermediate filament cytoskeleton of simple epithelia. Increased levels of these keratins have been associated with invasive growth and progression to malignancy in different types of human and murine epithelial tumors (including skin tumors), and even in tumors from nonepithelial origin. However, it has not yet clarified whether K8/K18 expression in tumors is cause or consequence of malignancy. Given the increasing incidence of epidermal cancer in humans (40% of all tumors diagnosed), we generated a mouse model to examine the role of simple epithelium keratins in the establishment and progression of human skin cancer. Transgenic mice expressing human K8 in the epidermis showed severe epidermal and hair follicle dysplasia with concomitant alteration in epidermal differentiation markers. The severity of the skin phenotype of these transgenic mice increases with age, leading to areas of preneoplastic transformation. Skin carcinogenesis assays showed a dramatic increase in the progression of papillomas toward malignancy in transgenic animals. These results support the idea that K8 alters the epidermal cell differentiation, favors the neoplastic transformation of cells, and is ultimately responsible of the invasive behavior of transformed epidermal cells leading of conversion of benign to malignant tumors.

Topics: Aging; Animals; Animals, Newborn; Cell Differentiation; Cell Transformation, Neoplastic; Disease Progression; Epidermis; Hair Follicle; Humans; Keratins; Mice; Mice, Transgenic; Precancerous Conditions; Skin Abnormalities; Skin Neoplasms; Transgenes

We report a 75-year-old male with anaplastic carcinoma in an extrathyroid area. Thyroid remained unchanged. The patient is alive without incident of tumor recurrence at 3.5 years after total resection and at 5 years after initial symptom. The tumor developed between the sternocleidomastoid muscle and common carotid artery, and was completely separated from the thyroid. The tumor location was consistent with a branchial cyst. The tumor consisted of two parts; an upper solid tumor and a deep cystic tumor. The former showed anaplastic carcinoma with osteoclast-like giant cells. The latter was consistent with thyroid papillary carcinoma. The center was intermingled with these two carcinomas. Anaplastic carcinoma cells were positive for vimentin and papillary carcinoma cells were positive for keratin, thyroglobulin, and thyroid transcription factor-1. These results remain insufficient to find any conclusions concerning the tumor nature; either ectopic thyroid carcinoma arising from a branchial cyst or occult thyroid carcinoma metastasis. This is rare case in which thyroid anaplastic carcinoma transformed from papillary carcinoma in an extrathyroid area.

Topics: Aged; Branchioma; Carcinoma; Carcinoma, Papillary; Cell Transformation, Neoplastic; Diagnosis, Differential; Humans; Keratins; Male; Nuclear Proteins; Osteoclasts; Thyroglobulin; Thyroid Neoplasms; Thyroid Nuclear Factor 1; Tomography, X-Ray Computed; Transcription Factors; Vimentin

In recent studies, Böcker and colleagues described a population of cells in paraffin wax sections of normal human breast that express cytokeratins (CK) 5/6 without expression of CK8/18 or smooth muscle actin (SMA). They proposed that these represent stem cells that give rise to differentiated luminal and myoepithelial cells. The data have been used to generate a model for breast cancer progression and classification with associated implications for management of pre-invasive disease. In this study, the expression of CK5/6, CK8/18, and SMA was investigated using multiple immunofluorescence on matched pairs of paraffin wax-embedded and frozen breast specimens. The staining patterns reported previously in antigen-retrieved paraffin wax-embedded sections were confirmed but no CK5/6-only cells were found in frozen sections of normal breast. There were cells with low levels of CK8/18 expression in frozen sections that may correspond to the CK8/18 'negative' cells seen in paraffin wax sections. This study brings into question the previously described profile of breast 'stem cells' based on CK5/6 staining and hence the breast cancer progression model and classification based on this phenotype.

Topics: Actins; Breast; Breast Neoplasms; Cell Transformation, Neoplastic; Disease Progression; Epithelial Cells; Female; Fluorescent Antibody Technique; Humans; Immunohistochemistry; Keratin-18; Keratin-5; Keratin-8; Keratins; Muscle, Smooth; Neoplasm Proteins; Phenotype; Stem Cells

To assess cytokeratin-17 (CK17) as an immunohistochemical marker for squamous cell carcinoma of the larynx, we stained 63 tissue samples from 63 consecutive patients who were believed or suspected to have squamous cell carcinoma of the larynx for CK17 and analyzed them by computerized histomorphometry. The mean staining intensity for CK17 was significantly stronger (p < .01) in cancerous cells, dysplasia, and normal epithelium proximal to the tumor than in distal normal epithelium and polyps. The percentage of stained area, within samples taken from a single patient, was significantly higher in malignancy and dysplasia as compared to distal normal epithelium and in malignancy as compared to dysplasia and proximal normal epithelium (p < .001). The integrated optical density was significantly higher in the malignant epithelium, dysplasia, polyps, and proximal normal epithelium than in distal normal epithelium (p < .0001). We conclude that CK17 is a highly sensitive and specific immunohistochemical marker for premalignant and malignant transformation in the larynx. Further investigation is warranted in order to assess the role of CK17 in determining safe resection borders.

Topics: Aged; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Laryngeal Neoplasms; Male; Middle Aged; Neoplasm Proteins; ROC Curve; Sensitivity and Specificity

The aim of this study was to determine the effects of exogenous expression of the catalytic subunit of telomerase (hTERT) on the lifespan, growth characteristics, and tumorigenicity of normal human ovarian surface epithelial (OSE) cells.. Low-passage primary cultures of normal human OSE cells were transfected with hTERT and the resulting cell lines were characterized.. The ectopic expression of hTERT stabilized the telomeres of the OSE cultures above 8 kb. The hTERT-transfected OSE cell lines grew beyond the normal lifespan seen in OSE cells and propagated in culture for more than 40 passages before senescing. Moreover, the hTERT-transfected cells demonstrated extensive proliferative capacity as evidenced by their ability to continuously grow even when seeded at low dilutions. The morphologic features and normal differentiation patterns seen in normal OSE cells were likewise retained by the hTERT-transfected cells. In addition, the cultures remained responsive to physiologic concentrations of epidermal growth factor and transforming growth factor-beta. Changes associated with neoplastic transformation like anchorage-independent growth, tumorigencity and karyotypic instability were not observed.. We were able to show that the ectopic expression of hTERT in normal human OSE: 1) resulted in cultures with greater growth potential and longer lifespan and 2) did not induce a transformed phenotype previously seen in viral oncogene-transfected OSE cells. The established cell lines would not only provide sufficient material for comprehensive studies to investigate the normal physiology of OSE cells, but could also help in the understanding of the early steps of ovarian carcinogenesis.

Topics: Animals; Cell Differentiation; Cell Division; Cell Survival; Cell Transformation, Neoplastic; Cells, Cultured; DNA-Binding Proteins; Epidermal Growth Factor; Epithelial Cells; Female; Humans; Immunohistochemistry; Keratins; Mice; Mice, SCID; Ovarian Neoplasms; Ovary; Telomerase; Transfection; Transforming Growth Factor beta; Tumor Cells, Cultured

Topics: Aged; Biomarkers, Tumor; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Keratins; Male; Mucin-1; Sebaceous Glands; Skin Neoplasms

The current study was undertaken to analyse growth and differentiation-related functions of normal keratinocytes (NOK) and an SV40T-immortalized keratinocyte line (SVpgC2a) from buccal mucosa, viewing the latter cell line as a model of a dysplastic epithelium. Morphological and immunohistochemical assessments of organotypic epithelia generated from 10 or 17 d of culture showed three- to five-fold higher apoptotic and proliferative activity in SVpgC2a relative to NOK. Conditions with or without serum (up to 10%) did not significantly influence these parameters in NOK whereas serum supported proliferation of SVpgC2a. Both cell types showed basal expression of collagen IV and laminin 1, indicating basal lamina, as well as vimentin, indicating an activated, proliferative state. Reduced expression of keratin, including the non-keratinizing marker K13, was seen in SVpgC2a. Assessment of proliferative monolayer cultures by microarray showed that NOK transcribed tissue-specific keratins, but also the epidermal keratin K2a, several simple epithelial keratins and low levels of hair keratins. SVpgC2a transcribed keratins seen in epithelial dysplasia, and K2a and hair keratins, albeit at low level. Overall, the results implied aberrant apoptosis, proliferation and keratin expression in the immortalized state of SVpgC2a. Comparison of NOK and SVpgC2a under identical culture conditions may serve to model the progression from a normal to a pre-neoplastic state of buccal epithelium.

Topics: Apoptosis; Cell Culture Techniques; Cell Differentiation; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Cells, Cultured; Collagen Type IV; Culture Media, Serum-Free; Humans; Immunoenzyme Techniques; Keratinocytes; Keratins; Laminin; Mouth Mucosa; Oligonucleotide Array Sequence Analysis; Organ Specificity; RNA, Messenger; Vimentin

Recent genetic investigations support the idea that basal cell carcinoma (BCC) is trichoblastic carcinoma. However, it is generally thought that clear cell basal cell carcinoma is a result of degeneration rather than tricholemmal differentiation.. We report a case of BCC, with clear cell components, that developed within seborrheic keratosis, with histopathological and immunohistochemical findings.. The clear cell components in the present case showed the following four characteristics: (i) at the periphery of the aggregations, columnar clear cells were aligned in a palisade along a well-defined basement membrane; (ii) the nuclei of the columnar clear cells were at the pole opposite the basement membrane; (iii) the clear cells contained glycogen; (iv) in the aggregations with clear cell components, there was diffuse positive staining for cytokeratin 7 (CK7) (OV/TLR/30), but only the inner region stained positive for CK17. These four characteristics are comparable to those of the lower portion of normal outer root sheath. In addition, the BCC in the present case was partly composed of squamous cells that contained glycogen and were selectively positive for CK17 - features similar to those of squamous cells in normal outer root sheath.. Some clear cell BCCs are simply the result of degenerative change, but other clear cell BCCs may be the result of tricholemmal (at the lower portion) differentiation.

Topics: Biomarkers, Tumor; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Hair Diseases; Hair Follicle; Humans; Immunoenzyme Techniques; Keratin-7; Keratins; Keratosis, Seborrheic; Male; Middle Aged; Skin Neoplasms

While squamous cell carcinoma and pseudocarcinomatous hyperplasia have been documented as pre-existing lesions in cases of reactive eccrine syringofibroadenoma (ESFA), to the best of our knowledge carcinoma occurring in a solitary ESFA has not yet been reported. We present one such case in a 91-year-old female who had a dome-shaped, reddish tumor on the extensor side of the left forearm.. We review the histopathological, immunophenotypical and ultrastructural findings of this tumor, including the keratin expression profile.. Histopathologically, long, branching, anastomosing, thin and thick strands of small cuboidal epithelial cells were extending from the surface epidermis into the dermis. In the center of the tumor, there were irregular-shaped nests of atypical tumor cells invading downward into the dermis. Ultrastructurally, duct-like lumina lined with cuboidal tumor cells were present in the epithelial cords. From these findings, the present case was diagnosed as solitary eccrine syringofibroadenocarcinoma (ESFAC). Keratin expression studies revealed that cells of the thick strands, except for the luminal and basal cells, were positive for differentiation-specific keratins, keratins 1 and 10, and that cells of the thin strands were positive for keratins 5 and 14.. Histopathological, immunophenotypical and ultrastructural evidence, as well as the pattern of keratin expression, suggest differentiation of the present malignant tumor towards the eccrine dermal duct. This case is the first reported case of ESFAC as far as we know.

Topics: Aged; Aged, 80 and over; Biomarkers, Tumor; Carcinoma; Cell Transformation, Neoplastic; Eccrine Glands; Female; Fibroadenoma; Humans; Immunohistochemistry; Keratins; Sweat Gland Neoplasms; Syringoma; Treatment Outcome

PMC42-LA cells display an epithelial phenotype: the cells congregate into pavement epithelial sheets in which E-cadherin and beta-catenin are localized at cell-cell borders. They abundantly express cytokeratins, although 5% to 10% of the cells also express the mesenchymal marker vimentin. Stimulation of PMC42-LA cells with epidermal growth factor (EGF) leads to epithelio-mesenchymal transition-like changes including up-regulation of vimentin and down-regulation of E-cadherin. Vimentin expression is seen in virtually all cells, and this increase is abrogated by treatment of cells with an EGF receptor antagonist. The expression of the mesenchyme-associated extracellular matrix molecules fibronectin and chondroitin sulfate proteoglycan also increase in the presence of EGF. PMC42-LA cells adhere rapidly to collagen I, collagen IV, and laminin-1 substrates and markedly more slowly to fibronectin and vitronectin. EGF increases the speed of cell adhesion to most of these extracellular matrix molecules without altering the order of adhesive preference. EGF also caused a time-dependent increase in the motility of PMC42-LA cells, commensurate with the degree of vimentin staining. The increase in motility was at least partly chemokinetic, because it was evident both with and without chemoattractive stimuli. Although E-cadherin staining at cell-cell junctions disappeared in response to EGF, beta-catenin persisted at the cell periphery. Further analysis revealed that N-cadherin was present at the cell-cell junctions of untreated cells and that expression was increased after EGF treatment. N- and E-cadherin are not usually coexpressed in human carcinoma cell lines but can be coexpressed in embryonic tissues, and this may signify an epithelial cell population prone to epithelio-mesenchymal-like responses.

Topics: Actins; Breast Neoplasms; Cadherins; Carcinoma; Cell Adhesion; Cell Transformation, Neoplastic; Chemotaxis; Dose-Response Relationship, Drug; Epidermal Growth Factor; Extracellular Matrix Proteins; Keratins; Neoplasm Proteins; Tumor Cells, Cultured; Vimentin; Vinculin

We have developed a series of novel mammary epithelial cell lines from tumors arising in strain 129 mice, with the ultimate goal of evaluating the role of host factors in the development of bone metastases. Mammary tumors were induced in mice with subcutaneously implanted medroxyprogesterone acetate (MPA) pellets followed by administration of DMBA by oral gavage. Mammary tumor development was efficient in the 129 strain and was independent of osteopontin (OPN) expression. Epithelial cell lines were isolated from these tumors; surprisingly, these cells did not form tumors upon inoculation into the mammary fat pad of syngeneic mice, even when MPA was present. One OPN-deficient cell line was selected for further study; full transformation of these cells required expression of both polyoma middle T and activated ras. These doubly transfected cells, 1029 GP+Er3, grew in soft agar, and formed hormone-independent tumors efficiently in the mammary fat pad that spontaneously metastasized to several soft tissue sites but not to the bone. Derivatives of these cells were isolated from tumors arising in the fat pad and from a lung metastasis (r3T and r3L, respectively): these cells formed tumors more rapidly in the fat pad than the parental GP+Er3 cells. Upon left ventricle injection, the r3T and r3L cells formed osteolytic bone metastases in 129 mice, with few metastases seen in other organs. These tumors filled the marrow cavity, and caused extensive destruction of both cortical and trabecular bone. Intriguingly, in an alternative syngeneic host, (129xC57B1/6) F1, osteolytic bone metastases were not seen on x-ray; instead extensive liver metastasis was present in these mice, indicating that genetic factors in these two strains regulate tumor cell homing and distribution during metastasis. These cell lines provide an important new tool in the study of bone metastasis, particularly in elucidating the role of host factors in the development of these lesions, as the 129 mouse strain is frequently used for genetic manipulations in the mouse.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Antigens, Polyomavirus Transforming; Antineoplastic Agents, Hormonal; Bone Neoplasms; Carcinogens; Cell Transformation, Neoplastic; Epithelial Cells; Female; Genes, ras; Heart Ventricles; Humans; Keratins; Liver Neoplasms, Experimental; Male; Mammary Neoplasms, Experimental; Medroxyprogesterone Acetate; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Neoplastic Cells, Circulating; Osteolysis; Osteopontin; Retroviridae; Sialoglycoproteins; Transfection; Tumor Cells, Cultured

A frequent genetic alteration found in premalignant stages of pancreatic adenocarcinoma is K-ras oncogene point mutation. The mechanistic basis for the inability of K-ras mutation to transform pancreatic ductal cells is unclear, although cooperating events with p16 inactivation, p53 mutation, and SMAD 4 mutation are recognized to be necessary. We have generated a novel mouse model in which the cytokeratin 19 promoter, specifically active in pancreatic ductal cells but not other cell types of the pancreas, is fused to mutant K-ras. This is of direct relevance to human pancreatic cancer because premalignant lesions are found specifically in ductal cells. There is dramatic periductal lymphocytic infiltration in the pancreata of transgenic mice, predominantly CD4+ T lymphocytes, which may act as an adaptive immune response to activated ras-mediated signaling. In addition, gene array analysis reveals an induction of N-cadherin in transgenic mice pancreatic ductal cells, the significance of which relates to promotion of cell adhesion and deterrence of cell migration. Apart from these important biological considerations, there is parallel activity of the cytokeratin 19 promoter in the stem cell region of the gastric epithelium, namely in mucous neck cells. Activated K-ras in this context causes mucous neck cell hyperplasia, a precursor to gastric adenocarcinoma. There is concomitant parietal cell decrease, which is a key step toward gastric adenocarcinoma. Taken together, we have defined how mutant K-ras signaling modulates important molecular events in the initiating events of pancreatic and gastric carcinogenesis.

Topics: Adenocarcinoma; Animals; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Gastric Mucosa; Genes, ras; Humans; Hyperplasia; Keratins; Lymphocytes, Tumor-Infiltrating; Mice; Mice, Transgenic; Mutation; Pancreatic Neoplasms; Precancerous Conditions; Promoter Regions, Genetic; Stomach Neoplasms; Transfection; Tumor Cells, Cultured

Expression of cytokeratin 7 (CK7) and cytokeratin 20 (CK20) helps to establish the origin of biliary and metastatic carcinomas. We investigated the expression of CK7 and CK20 in inflammatory, metaplastic and neoplastic conditions of the bile ducts, and evaluated possible relationships between the CK expression pattern and extrahepatic bile duct/gallbladder carcinomas (EBDCs) or intrahepatic bile duct carcinomas (IBDCs). We used immunohistochemistry for the investigation of 48 formalin-fixed, paraffin-embedded specimens grouped as: A) lithiasic or inflamed surgically resected extrahepatic bile ducts/gallbladders: all were CK7+/CK20+; B) percutaneous liver biopsies from patients with chronic hepatitis C primary biliary cirrhosis and primary sclerosing cholangitis: all were CK7+/CK20-; C) EBDCs: all were CK7+/CK20+, except for two cases which were CK7-/CK20-; D) IBDCs: all were CK7+/CK20-, except for one case showing CK20 positivity. Metaplastic changes were seen only among specimens in groups A and C: in these cases, CK20 was either focally or diffusely expressed. Our study suggests that the expression of cytokeratins under specific stimuli can be different from normal tissues, and that sometimes CK20 expression can be related to and precede the occurrence of metaplastic alterations.

Topics: Bile Duct Diseases; Bile Duct Neoplasms; Bile Ducts, Extrahepatic; Bile Ducts, Intrahepatic; Carcinoma; Cell Transformation, Neoplastic; Gallbladder Diseases; Gallbladder Neoplasms; Gene Expression Profiling; Humans; Immunohistochemistry; Intermediate Filament Proteins; Keratin-20; Keratin-7; Keratins

Tumor promoters such as the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) are proinflammatory agents, and their mechanism of action in epithelial carcinogenesis has been linked to the release of IL-1 alpha and the induction of chronic inflammation in skin. To test the role of IL-1 alpha and inflammation in models of cutaneous carcinogenesis, we used our previously described FVB/N transgenic mice overexpressing 17-kDa IL-1 alpha in the epidermis under the keratin 14 (K14) promoter. Strikingly, the K14/IL-1 alpha mice were completely resistant to papilloma and carcinoma formation induced by a two-stage DMBA/TPA protocol, while littermate controls developed both tumor types. K14/IL-1 alpha mice crossed with the highly sensitive TG.AC mice, constitutively expressing mutant Ha-Ras, also failed to develop papillomas or carcinomas. When the K14/IL-1 alpha transgene was bred onto a recombinase-activating gene-2-deficient background, the resistance persisted, indicating that innate, but not acquired, mechanisms may be involved in the resistance to the initiation/promotion model. As an alternative approach, a complete carcinogenesis protocol using repetitive application of DMBA alone was applied. Surprisingly, although the IL-1 alpha mice still did not develop papillomas, they did develop carcinomas de novo at an accelerated rate compared with controls. We conclude that constitutive IL-1 alpha expression rendered FVB mice completely resistant to carcinomas that required evolution from prior papillomas, but facilitated carcinomas that did not evolve from papillomas, as in the complete carcinogenesis protocol. Thus, the role of IL-1 alpha and, by extension that of other proinflammatory factors, in epithelial carcinogenesis are more complex than previously appreciated. These mice may provide a mechanism to investigate the validity of these models of human skin tumorigenesis.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; DNA-Binding Proteins; Epidermis; Female; Humans; Immunity, Innate; Interleukin-1; Keratin-14; Keratins; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Nuclear Proteins; Papilloma; Skin Neoplasms; Tetradecanoylphorbol Acetate; Transgenes

Human papillomaviruses (HPVs) are the causative agent of warts. Infections with high-risk HPVs are associated with anogenital and head and neck cancers. One of the viral genes responsible for HPV's oncogenic activity is E6. Mice expressing the HPV-16 E6 protein in their epidermis (K14E6(WT)) develop epithelial hyperplasia and squamous carcinomas. Numerous cellular proteins interact with E6, some of which can be grouped based on common amino acid motifs in their E6-binding domains. One such group, the PDZ partners, including hDLG, hSCRIBBLE, MUPP1, and MAGI, bind to the carboxy-terminal four amino acids of E6 through their PDZ domains. E6's interaction with the PDZ partners leads to their degradation. Additionally, E6's binding to PDZ proteins has been correlated with its ability to transform baby rat kidney cells in tissue culture and to confer tumorigenicity onto cells in xenograft experiments. To address whether the ability of E6 to bind PDZ domain partners is necessary for E6 to confer epithelial hyperproliferation in vivo, we generated transgenic mice that express in stratified squamous epithelia a mutant of E6 lacking the last six amino acids at its carboxyl terminus, E6(Delta 146-151), from the human keratin 14 (K14) promoter. The K14E6(Delta 146-151) mice exhibit a radiation response similar to that of the K14E6(WT) mice, demonstrating that this protein, as predicted, retains an ability to inactivate p53. However, the K14E6(Delta 146-151) mice fail to display epithelial hyperplasia. These results indicate that an interaction of E6 with PDZ partners is necessary for its induction of epithelial hyperplasia.

Topics: Adaptor Proteins, Signal Transducing; Animals; Carrier Proteins; Cell Transformation, Neoplastic; Discs Large Homolog 1 Protein; DNA Damage; Guanylate Kinases; Humans; Hyperplasia; Keratin-14; Keratins; Ligands; Membrane Proteins; Mice; Mice, Transgenic; Oncogene Proteins, Viral; Papillomaviridae; Proliferating Cell Nuclear Antigen; Proteins; Rats; Repressor Proteins; Skin Neoplasms

We report here that human esophageal keratinocyte stem cells are characterized by the expression of the low-affinity neurotrophin receptor p75(NTR) and differentially expressed cell adhesion molecules, the beta1 and beta4 integrins. The candidate stem cells could be fractionated from keratinocytes as a minor cell subset by means of immunocytochemical cell sorting based on the different levels of expression of these cell surface molecules. Flow cytometric analysis revealed that this minor cell subset retained a relatively slow-cycling phenotype in vitro. These cells expressed low levels of involucrin and cytokeratin 13, indicating that the p75(NTR)-positive cell subset is immature relative to the other predominant subpopulations coexpressing beta1 integrin at higher levels. The p75(NTR)-positive cell subset was crucial for achieving longevity and the greatest output of keratinocytes comprising all distinguishable subpopulations in vitro. This process was associated with self-renewal and self-amplification of the p75(NTR)-positive cell subset. These findings strongly implicate p75(NTR) as a stem cell marker, which will be valuable for prospectively investigating stem cell regulation in association with different biological processes including neoplastic transformation of regenerative epithelia.

Topics: Blotting, Western; Carcinoma, Squamous Cell; Cell Adhesion; Cell Cycle; Cell Division; Cell Membrane; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Esophagus; Flow Cytometry; Humans; Immunohistochemistry; Integrin beta1; Integrin beta4; Keratinocytes; Keratins; Neoplasms; Phenotype; Propidium; Protein Precursors; Receptor, Nerve Growth Factor; Receptors, Nerve Growth Factor; Reverse Transcriptase Polymerase Chain Reaction; Stem Cells; Subcellular Fractions; Time Factors

Accumulating evidence indicates a decisive role for the adjacent stroma in tumour growth and dissemination. However, it is not clear how far altered differentiation such as expression of aberrant keratins and vimentin, common in invasive human carcinomas, may reflect intrinsic cell properties or a response to the tumour environment. We have addressed this by transplanting benign and malignant human HaCaT-ras keratinocytes, seeded on collagen matrix, onto nude mice. Initially, epithelia derived from benign and malignant cells, being separated from host stroma by collagen, were poorly organized and exhibited the same differentiation markers, as identified by immunofluorescence and in situ hybridization. Epidermal basal and suprabasal keratins were expressed persistently even upon contact with newly formed stroma and malignant cell invasion. In contrast, non-epidermal keratins (K4/K13, K8/18, K19), which were similarly synthesized by benign and malignant cells in culture and in early transplants, were differentially regulated with increasing stromal vicinity. While both proteins and mRNAs were downregulated in benign epithelia, the malignant, invasive tumour cells continuously expressed these non-epidermal keratins throughout (K19), suprabasally (K4/13) or at invasive sites (K8/18). Furthermore, the mesenchymal protein vimentin was expressed de novo in invasive areas confronting tumour stroma. Thus, atypical tissue markers, similarly synthesized in isolated cells in vitro, are downregulated in benign but maintained and upregulated in malignant epithelia. This is presumably caused by the neighbouring stroma being permanently activated by malignant epithelia.

Topics: Animals; Antigens, Differentiation; Cell Transformation, Neoplastic; Down-Regulation; Epidermis; Fluorescent Antibody Technique; Humans; In Situ Hybridization; Keratinocytes; Keratins; Mice; Neoplasm Invasiveness; Neoplasm Proteins; RNA, Messenger; RNA, Neoplasm; Skin Neoplasms; Tumor Cells, Cultured; Vimentin

We have developed a nontumorigenic epithelial cell line, DP-153, from the dorsal prostate of a Lobund/Wistar rat treated with N-methyl-N-nitrosourea and testosterone propionate. DP-153 cells express cytokeratins 5 and 14, but not cytokeratin 18, consistent with a basal epithelial cell phenotype. Similar to the nontumorigenic NRP-152 prostatic cell line, DP-153 cells do not form tumors in athymic mice and retain many of the properties of normal prostatic cells. They express prostatic acid phosphatase and androgen receptors and require several mitogens (epidermal growth factor, insulin, dexamethasone, and cholera toxin) for sustained growth in culture under serum-containing conditions. DP-153 cells are also growth-stimulated by keratinocyte growth factor and basic fibroblast growth factor and growth-inhibited by all-trans-retinoic acid, 1,25-dihydroxyvitamin D(3), and transforming growth factor (TGF)-beta1. We demonstrate that expression of dominant-negative TGF-beta receptor type II by retroviral transduction of DP-153 cells leads to complete loss of TGF-beta1-induced growth inhibition. When transplanted s.c. in athymic mice, DP-153 cells expressing dominant-negative TGF-beta receptor type II form tumors as early as 4 weeks, in contrast to the vector control and parental cell line, which do not form tumors even 8 months after transplantation, supporting the observation that TGF-beta functions as a tumor suppressor in these cells. Our data further support that DP-153 is a suitable cell line for analysis of normal prostatic growth and carcinogenesis.

Topics: Animals; Calcitriol; Cell Division; Cell Transformation, Neoplastic; Growth Substances; Isoenzymes; Karyotyping; Keratins; Male; Mice; Mice, Nude; Neoplasm Transplantation; Prostatic Neoplasms; Protein Serine-Threonine Kinases; Rats; Rats, Wistar; Receptor, Transforming Growth Factor-beta Type II; Receptors, Androgen; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta; Transplantation, Heterologous; Tretinoin; Tumor Cells, Cultured

Adenocarcinomas of nonsalivary origin represent approximately 10% to 20% of all sinonasal malignancies and are characterized by varying histopathologic features and uncertain histogenesis. To better understand the histogenesis and phenotypic heterogeneity of these tumors, we performed immunohistochemical analyses for cytokeratin (CK) 7 and CK20 on 12 primary sinonasal adenocarcinomas (SNACs) representing the histopathologic spectrum of these tumors, adjacent normal mucosa, and 2 metastatic adenocarcinomas from colonic primaries. The demographic and clinicopathologic characteristics of our cohort were similar to those in previously published series. Our results indicate that histologically normal respiratory-type epithelium and submucosal seromucous glands show restricted reactivity to CK7. Epithelial metaplasia of surface epithelium associated with enteric SNACs was accompanied by a conversion from CK7 positivity to CK20 positivity. All primary enteric-type carcinomas and the 2 colonic metastases were reactive to CK20, but all nonenteric-type tumors were negative for CK20 (P=0.003) and positive for CK7. In some of the enteric types, coexpression of CK7 and CK20 was noted. We conclude that (1) nonenteric-type (seromucinous) adenocarcinoma may originate directly from surface respiratory-type epithelium or from seromucous glands, (2) metaplastic transformation of surface respiratory to enteric-type epithelium precedes the development of enteric adenocarcinoma, and (3) coordinate analyses of CK7 and CK20 reactivity may aid the differential diagnosis of adenocarcinoma in the sinonasal tract.

Topics: Adenocarcinoma; Aged; Aged, 80 and over; Biomarkers, Tumor; Cell Transformation, Neoplastic; Colonic Neoplasms; Diagnosis, Differential; Humans; Immunohistochemistry; Intermediate Filament Proteins; Keratin-20; Keratin-7; Keratins; Male; Middle Aged; Nose Neoplasms; Paranasal Sinus Diseases; Phenotype; Respiratory Mucosa

The Wnt/beta-catenin signaling pathway controls cell fate and neoplastic transformation. Expression of an endogenous stabilized beta-catenin (DeltaE3 beta-catenin) in mammary epithelium leads to the transdifferentiation into epidermis- and pilar-like structures. Signaling molecules in the canonical Wnt pathway upstream from beta-catenin induce glandular tumors but it is not clear whether they also cause squamous transdifferentiation. To address this question we have now investigated mammary epithelium from transgenic mice that express activating molecules of the Wnt pathway: Wnt10b, Int2/Fgf3, CK2alpha, DeltaE3 beta-catenin, Cyclin D1, and dominant negative (dn) GSK3beta. Cytokeratin 5 (CK5), which is expressed in both mammary myoepithelium and epidermis, and the epidermis-specific CK1 and CK6 were used as differentiation markers. Extensive squamous metaplasias and widespread expression of CK1 and CK6 were observed in DeltaE3 beta-catenin transgenic mammary tissue. Wnt10b and Int2 transgenes also induced squamous metaplasias, but expression of CK1 and CK6 was sporadic. While CK5 expression in Wnt10b transgenic tissue was still confined to the lining cell layer, its expression in Int2 transgenic tissue was completely disorganized. In contrast, cytokeratin expression in CK2alpha, dnGSK3beta and Cyclin D1 transgenic mammary tissues was similar to that in DeltaE3 beta-catenin tissue. In support of transdifferentiation, expression of hard keratins specific for hair and nails was observed in pilar tumors. These results demonstrate that the activation of Wnt signaling components in mammary epithelium induces not only glandular tumors but also squamous differentiation, possibly by activating LEF-1, which is expressed in normal mammary epithelium.

Topics: Animals; beta Catenin; Breast; Calcium-Calmodulin-Dependent Protein Kinases; Cell Transformation, Neoplastic; Cyclin D1; Cytoskeletal Proteins; DNA-Binding Proteins; Female; Fibroblast Growth Factor 3; Fibroblast Growth Factors; Glycogen Synthase Kinase 3; Immunoenzyme Techniques; Keratins; Lymphoid Enhancer-Binding Factor 1; Mammary Neoplasms, Experimental; Mice; Mice, Transgenic; Proto-Oncogene Proteins; Signal Transduction; Trans-Activators; Transcription Factors; Up-Regulation; Wnt Proteins

Multidirectional differentiation in colorectal carcinomas is a rare phenomenon. Four cases are reported herein, and their clinical and pathologic characteristics are discussed. Two men and 2 women between the ages of 56 and 76 years who presented with abdominal symptoms are included in this report. Two tumors were located in the right colon, one in the splenic flexure, and one in the descending colon. Distant metastases were evident at presentation in 3 of 4 cases. Histologically, two tumors exhibited neuroendocrine and glandular differentiation; the third tumor was an adenocarcinoma with a sarcomatous component and the fourth tumor showed 3 lines of differentiation (glandular, squamous, and sarcomatoid). In all tumors evaluated, areas of adenocarcinomas were positive for low-molecular weight cytokeratin (CAM 5.2) and mucicarmine, but negative for high-molecular weight cytokeratin (AE3). The squamous cell component was AE3 positive and CAM 5.2 negative. The neuroendocrine component was highlighted by neuroendocrine markers and the sarcomatoid component revealed smooth muscle differentiation. All tumors (except one mucinous tumor) were negative for cytokeratin-20 staining. One patient was on supportive care for terminal metastatic carcinoma, and 2 patients were being treated with adjuvant chemotherapy at the time of this report. Colon carcinoma with multidirectional differentiation is a rare event and may originate from stem cells within the gastrointestinal mucosa, and/or represent the convergence of multiple tumors arising at the same site. This type of tumor should be considered in the differential diagnosis of a bowel biopsy with multiple histopathologic variants.

Topics: Adenocarcinoma; Aged; Carcinoma, Squamous Cell; Carmine; Cell Transformation, Neoplastic; Colorectal Neoplasms; Coloring Agents; Female; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Neoplasm Metastasis

The stability of cytokeratin (CK) protein during tumor transformation in human hepatocellular carcinoma (HCC) was studied with molecular approach previously. The results demonstrated that the CK was modulated in human HCC. Besides this, three low molecular weight CK molecules (named HCC CK) were found. It indicated that these HCC CKs are undergone modulation from human hepatocyte CK18. However, there were many differences between the CK18 and HCC CK. First, the antigenecity of HCC CK had been changed since they could not be recognized by CAM5.2 antibody on Western blot. Second, the sequences of N-terminal residues of HCC CK were matched with those of the N-terminal residues of human histone. In this study, we confirmed that the HCC CK was actually to be histones because they reacted to anti-histone antibody on Western blot. Furthermore, we found that the histones of human HCC had been changed during the process of tumor transformation since they could be co-immunoprecipitated with CK18 and could be detected by Western blot while this phenomenon did not happen in the normal liver tissue. We also found that not all histones change in human HCC. Only H3 was detected on Western blot while H1, H2A, H2B, and H4 were not detected in human HCCs.

Topics: Blotting, Western; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Electrophoresis, Polyacrylamide Gel; Histones; Humans; Keratins; Liver Neoplasms; Precipitin Tests

Cadmium is a ubiquitous environmental human carcinogen. Epidemiological and animal studies have suggested its carcinogenic potential on the prostate. In the present study, non-tumorigenic human prostate epithelial cells (pRNS-1-1) immortalized by simian papovavirus (SV40) were transformed after repeated exposures to cadmium. Such transformants showed morphological alterations, anchorage-independent growth in soft agar, and formed tumors when transplanted into SCID mice. The tumors were characterized histologically as poorly-differentiated adenocarcinomas, expressing prostate-specific antigen (PSA), androgen receptor (AR), prostate stem cell antigen (PSCA), NKX3.1 and cytokeratin 8 (CK8). These findings provide evidence of malignant transformation of human prostate epithelial cells exposed to this environmentally important chemical.

Topics: Adenocarcinoma; Animals; Cadmium; Cell Transformation, Neoplastic; Epithelial Cells; Humans; Karyotyping; Keratins; Male; Mice; Mice, SCID; Prostate-Specific Antigen; Prostatic Neoplasms; Receptors, Androgen; Reverse Transcriptase Polymerase Chain Reaction; Simian virus 40; Time Factors; Tumor Cells, Cultured

Liver regeneration after partial hepatectomy results primarily from the simple division of mature hepatocytes. However, during embryonic and fetal development or in circumstances under which postnatal hepatocytes are injured, organ regeneration is believed to occur from a compartment of epithelial liver stem or progenitor cells with biliary and hepatocytic bipotentiality. The ability to identify, isolate, and transplant epithelial liver stem cells from fetal liver would greatly facilitate the treatment of hepatic diseases currently requiring orthotopic liver transplantation. Here we report the identification and immortalization by retrovirus-mediated transfer of the simian virus 40 large T antigen gene of primate fetal epithelial liver cells with a dual hepatocytic biliary phenotype. These cells grow indefinitely in vitro and express the liver epithelial cell markers cytokeratins 8/18, the hepatocyte-specific markers albumin and alpha-fetoprotein, and the biliary-specific markers cytokeratins 7 and 19. Bipotentiality of gene expression was confirmed by clonal analysis initiated from single cells. Endogenous telomerase also is expressed constitutively. After orthotopic transplantation via the portal vein, approximately 50% of the injected cells integrated into the liver parenchyma of athymic mice without tumorigenicity. Three weeks after transplantation, cells having seeded in the liver parenchyma expressed both albumin and alpha-fetoprotein but had lost expression of cytokeratin 19. These results provide strong evidence for the existence of a bipotent epithelial liver stem cell in nonhuman primates. This unlimited source of donor cells also should enable the establishment of a model of allogenic liver cell transplantation in a large animal closely related to humans and shed light on important questions related to liver organogenesis and differentiation.

Topics: Albumins; alpha-Fetoproteins; Animals; Biomarkers; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Cell Transplantation; Cells, Cultured; Chimera; Clone Cells; Embryonic and Fetal Development; Epithelial Cells; Fetus; Gene Expression Regulation, Developmental; Keratins; Liver; Liver Regeneration; Macaca fascicularis; Mice; Mice, Nude; Stem Cells; Telomerase; Transplantation, Heterologous

Regular expansion of heterogeneous populations of epithelial cells, including the luminal phenotype, was achieved from small biopsies of human breast tumours and cutaneous metastases by optimized feeder layer technique based on irradiated NIH 3T3 cells. Forty-one out of 47 primary tumour specimens and all three cutaneous metastases grew successfully for two to 10 passages in vitro. The main phenotypes of cultured cells and their changes in subcultures were characterized using immunocytochemistry and phase contrast microscopy (in few cases also time-lapse recording). In the majority of cultured cell populations a fraction of cells positive for keratin 19 (K19+), typical for the luminal phenotype, was detected. This is the cell type from which breast carcinoma is supposed to arise. While in cultures derived from benign lesions only basic phenotypes of luminal and myoepithelial cells were found, in cultures derived from malignant tumours unusual phenotypes of epithelial cells, in their majority K19+, were detected. The growth properties of cells from six benign and seven malignant samples were analyzed in detail. In the analyzed cell populations the culture lifetime - related to the number of colony-forming cells varied for cells from malignant tumours between 21 and 51 and from benign tumours between 22 and 40 cell generations. The total number of passages achieved was three to seven for malignant or four to nine for benign cultures. In spite of negative results of tumourigenicity testing in immunologically compromised Nu/nu mice the potential to culture apparently neoplastic cells was indicated by positive immunostaining for the p53 oncoprotein (seven of 23 tested malignant cases), the src oncoprotein (five of eight), and overexpression of the c-erbB-2 protein (five of 26). This was further confirmed by successful cultivation of malignant cells from cutaneous metastases. Two of the three metastasis-derived cultures were nearly homogeneously positive for K19 while the third was almost negative. The results proved the optimized feeder layer technique to be useful for regular yielding of large amounts of epithelial cells from small tumour biopsies and for supporting the majority of cell phenotypes present in the original tumour. Therefore, it appeared to be a promising tool for further analysis of interactions between luminal and myoepithelial cells in the development of human breast carcinoma and for the study of individual tumours.

Topics: 3T3 Cells; Adult; Aged; Animals; Breast; Breast Neoplasms; Carcinoma, Ductal, Breast; Case-Control Studies; Cell Count; Cell Transformation, Neoplastic; Cells, Cultured; Coculture Techniques; Female; Humans; Immunohistochemistry; Keratins; Mice; Microscopy, Phase-Contrast; Middle Aged; Phenotype; Skin Neoplasms; Tumor Cells, Cultured

A case of esophageal basaloid carcinoma with marked myoepithelial differentiation in a 60-year-old man is reported. The tumor arose as an exophytic mass, measuring 65 x 60 mm, in the middle thoracic esophagus. Approximately two-thirds of the tumor surface was covered with non-cancerous esophageal epithelium. The depth of tumor invasion was limited to the submucosal layer. Histologically, about 70% of the tumor contained a typical basaloid carcinoma component and about 30% contained glandular and intercalated duct-like components with distinct epithelial and myoepithelial differentiation. The tumor presented no component of distinct squamous cell carcinoma, but a small portion of cribriform-like structure, which is typical of adenoid cystic carcinoma, was visible. The inner epithelium composing the intercalated duct-like structure showed immunohistochemical positivity for cytokeratin 14, and the outer epithelium lining adjacent to the stroma showed positivity for alpha-smooth muscle actin. These findings supported epithelial/myoepithelial differentiation. To our knowledge, our case is the first patient with an esophageal basaloid carcinoma showing marked myoepithelial differentiation.

Topics: Actins; Biomarkers, Tumor; Carcinoma, Transitional Cell; Cell Transformation, Neoplastic; Esophageal Neoplasms; Humans; Keratin-14; Keratins; Male; Middle Aged; Myoepithelioma; Neoplasm Staging; Treatment Outcome

A 54-year-old male had a dome-shaped and skin-colored nodule on his nose. Histopathologically, we diagnosed this neoplasm as a low-grade sebaceous carcinoma rather than a sebaceoma based on the scanning magnification and cytology. This low-grade sebaceous carcinoma was associated with glandular structures. We regarded the glandular structures as those of apocrine glandular differentiation based on 1) the histopathologic features of the glandular structures formed by columnar luminal cells with evidence of decapitation secretion; 2) the expression of cytokeratin (CK) 19, CK8, CK8/18, and CK7 in the luminal cells; 3) the positive reaction of carcinoembryonic antigen and epithelial membrane antigen on the luminal surface and in the cytoplasm of the luminal cells; and 4) the common embryologic origin of the folliculosebaceous-apocrine unit. We found CK15 expression in undifferentiated cells within the mantles of normal hair follicles, suggesting that sebaceous stem cells might exist in mantles as follicular stem cells exist in bulge areas. Pluripotent stem cells in the folliculosebaceous-apocrine unit can give rise to follicular stem cells, sebaceous stem cells, and apocrine stem cells. Our patient's neoplasm showed apocrine glandular differentiation and partial immunohistochemical positivity for CK15 in the neoplastic aggregations. We believe this neoplasm originated from pluripotent stem cells destined to become sebaceous stem cells or from sebaceous stem cells, which also have the ability to differentiate within apocrine glands.

Topics: Adenocarcinoma, Sebaceous; Apocrine Glands; Biomarkers, Tumor; Cell Transformation, Neoplastic; Humans; Immunoenzyme Techniques; Keratins; Male; Middle Aged; Neoplasm Proteins; Nose; Sebaceous Gland Neoplasms; Stem Cells

To determine whether Brenner tumors and transitional cell carcinomas (TCCs) of the ovary are urothelial in type, the immunoprofiles of 14 Brenner tumors, including three malignant examples, and eight ovarian TCCs were compared with those of Walthard nests, urothelium, 12 urinary bladder TCCs and 17 ovarian adenocarcinomas (serous, endometrioid, mucinous, and undifferentiated type). The immunohistochemical stains used included those for cytokeratins CKs 5/6, CK7, CK8, CK13, and CK20, vimentin, CA125, and the specific urothelial differentiation marker uroplakin III. CK7 and CK8 were broadly expressed in most tumors of ovary and bladder examined, while vimentin was focally present in some ovarian TCCs and adenocarcinomas. As in normal and neoplastic bladder urothelium, urothelial markers, including uroplakin III, CK13, and CK20, were detected in the epithelial nests of Brenner tumors. Brenner tumor cells also expressed uroplakins Ia and II. CA125 was observed focally in some Brenner tumors. In contrast, TCCs of the ovary and Walthard nests lacked uroplakins and were essentially negative for CK20 and CK13 but quite strongly expressed CA125. This immunophenotype closely resembled that found in ovarian adenocarcinomas. Thus, it appears that the only true urothelial-type ovarian neoplasm is the Brenner tumor, whereas ovarian TCC most likely represents a poorly differentiated adenocarcinoma with a morphologic transitional cell pattern. These results may explain the controversies as expressed in the recent literature concerning TCC of the ovary and establish its place among the ovarian adenocarcinomas of müllerian type.

Topics: Adenocarcinoma; Brenner Tumor; Carcinoma, Transitional Cell; Cell Transformation, Neoplastic; Female; Humans; Keratins; Membrane Glycoproteins; Ovarian Cysts; Ovarian Neoplasms; Urinary Bladder Neoplasms; Uroplakin III; Urothelium

The cytoskeleton plays important roles in cell function and is therefore implicated in the pathogenesis of many human liver diseases, including malignant tumors. The stability of cytokeratin proteins during tumor transformation in human hepatocellular carcinoma has been studied with a molecular approach previously. The results demonstrate that the cytokeratin is modulated in human hepatocellular carcinoma. Besides this, three low molecular weight cytokeratin molecules (named HCC CK) are found. This indicates that these HCC CKs have undergone modulation from the human hepatocyte cytokeratin 18. We also checked the cytokeratin profile of the human hepatoma cell line PLC/PRF/5 with the same methods to ensure the HCC CK molecules are produced by modulation but not protein degradation. The stability of cytokeratin molecules was studied by a different approach. The cytokeratin compositions of human liver cells (Chang cell line) were analysed under the effects of microtubule-disrupting drug (colchicine) by SDS-PAGE, Western blot, immunoprecipitation using a commercially available monoclonal anti-cytokeratin 18 antibody and immunofluorescent staining. Within 1 h of treatment, the microtubule began to collapse and the filamentous structure was shortening. The microtubule had almost collapsed and became fragmented to form a lattice-like network after 24 h of treatment. The cytokeratin was modulated after long-term (24 h) treatment of colchicine, and the molecular weight became 14 kD and the antigenicity was lost. The stability of cytokeratin molecules was related to the intact microtubule network, after disruption of the microtubule the cytokeratin would be modulated. The intact microtubule network was a stabilizing factor of cytokeratin 18 in human liver cells.

Topics: Blotting, Western; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Colchicine; Electrophoresis, Polyacrylamide Gel; Fluorescent Antibody Technique, Indirect; Humans; Keratins; Liver; Liver Neoplasms; Microscopy, Phase-Contrast; Microtubules; Precipitin Tests; Time Factors; Tumor Cells, Cultured

Cytokeratin (CK) expression was studied in 22 sinonasal inverted papillomas. Columnar (respiratory) epithelium in inverted papillomas abundantly expressed CK7, CK8, CK18, and CK19. Immunoreactivity for CK5/14 and CK17 was found in basal and parabasal/suprabasal cells. Transitional (cuboidal) and squamous epithelium in inverted papillomas comparably expressed CK7, CK8, CK18, and CK19. In addition CK13 was found in subluminal and surface cells. Immunoreactivity for CK5/14 and CK17 involved all layers of the epithelium. In nonpapillomatous nasal mucosa adjacent to inverted papillomas, CK expression in columnar (respiratory) epithelium exactly matched the findings in inverted papillomas. Transitional (cuboidal) and squamous epithelium in nonpaillomatous mucosa were negative for CK7, CK8, CK18, and CK19. CK13 was expressed in subluminal and surface cells. Immunoreactivity for CK5/14 and CK17 was restricted to basal and parabasal/suprabasal cells. Conclusively, transitional (cuboidal) and squamous epithelium in inverted papillomas but not in the adjacent mucosa coexpress CKs typical for columnar and squamous differentiation.

Topics: Adult; Aged; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Nasal Mucosa; Papilloma, Inverted; Paranasal Sinus Neoplasms

Transduction of human papillomavirus type 16 (HPV16) E6/E7 into primary culture of human esophageal keratinocytes using a recombinant adenovirus prolonged the life-span, while untreated cells senesced within 14-16 population doublings (PDLs). Up-regulation of telomerase activity and acquisition of serum-resistant growth were observed in the esophageal keratinocytes with extended life-span between 50 and 100 PDLs, and drastically increased after 100 PDLs. A keratinocyte sample with a polymorphism of Pro/Pro at codon 72 of p53 showed resistance to HPV16 E6/E7-induced life-span-extension and immortalization, in contrast to others with p53 polymorphisms of Arg/Arg or Arg/Pro, which did not. The high efficiency of E6/E7-induction by adenovirus vector also revealed the M1 and M2 stages of keratinocyte immortalization first described in this report.

Topics: Adenoviridae; Calcium; Cell Transformation, Neoplastic; Cell Transformation, Viral; Colony-Forming Units Assay; DNA Primers; Epithelial Cells; Esophagus; Genetic Vectors; Humans; In Situ Hybridization; In Vitro Techniques; Keratins; Microscopy, Phase-Contrast; Oncogene Proteins, Viral; Papillomaviridae; Papillomavirus E7 Proteins; Repressor Proteins; Telomere; Transfection; Tumor Suppressor Protein p53; Up-Regulation

We describe an insulinoma of the pancreas in a 56-year-old patient, which showed insular-ductular differentiation in its liver metastasis. Although the primary tumor was uniformly endocrine in nature with insulin production, the metastasis contained two distinct cell types in organoid arrangement. One cell type was insulin-positive and was arranged in islet-like structures; the other was insulin-negative but distinctly pan-cytokeratin and cytokeratin 7 positive and arranged in ducts. In the primary tumor and the metastasis, the tumor cells were surrounded by a desmoplastic stroma. As to the histogenesis of the tumor and its metastasis, we discuss the following possibilities: (1) the tumor cells might derive from a common stem cell that matures into two phenotypically different cell lines, resembling the situation in embryogenesis and (2) one tumor cell type originates from the other by transdifferentiation (metaplasia). We conclude that the parallel occurrence of endocrine and ductal differentiation supports the concept that, under certain conditions, islet cells and ductular cells may also originate from islets and that mixed endocrine/exocrine pancreatic tumors do not necessarily arise from totipotent duct cells but might also have a primary endocrine cell origin.

Topics: Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Cell Transformation, Neoplastic; Humans; Insulin; Insulinoma; Keratin-7; Keratins; Liver Neoplasms; Male; Middle Aged; Neoplastic Stem Cells; Pancreas; Pancreatic Neoplasms

Ultraviolet B irradiation initiates and promotes skin cancers, photo-aging, and immune suppression. In order to elucidate the effect of these processes at the level of gene expression, we used cDNA microarray technology to examine the effect of ultraviolet B irradiation on 588 cancer-related genes in human keratinocytes at 1, 6, and 24 h post-irradiation with a mildly cytotoxic dose of ultraviolet B (170 mJ/cm(2)). The viability of the irradiated keratinocytes was 75% at 24 h post-irradiation. Various cytokeratins and transcription factors were up-regulated within 1 h post-irradiation. After 6 h, expression of a variety of genes related to growth regulation (e.g. p21(WAF1), notch 4, and smoothened), apoptosis (e.g. caspase 10, hTRIP, and CRAF1), DNA repair (ERCC1, XRCC1), cytokines (e.g. IL-6, IL-13, TGF-beta, and endothelin 2), and cell adhesion (e.g. RhoE, and RhoGDI) were altered in human keratinocytes. These data suggest the changes in a cascade of gene expression in human keratinocytes occurring within 24 h after UVB exposure. Although the roles of these cellular genes after UVB-irradiation remain to be elucidated, microarray analysis may provide a new view of gene expression in epidermal keratinocytes following UVB exposure.

Topics: Apoptosis; Cell Survival; Cell Transformation, Neoplastic; Cytokines; DNA Repair; DNA, Complementary; Dose-Response Relationship, Radiation; Endothelins; Gene Expression Regulation; Growth Substances; Humans; Keratinocytes; Keratins; Oligonucleotide Array Sequence Analysis; RNA, Messenger; Transcription Factors; Transcription, Genetic; Ultraviolet Rays

We determined whether treatment of metastatic prostate cancer cells with doxorubicin (DOX) and interferon-alpha (IFN-alpha) prevented the emergence of highly undifferentiated tumor cells.. The state of cell differentiation was determined by analysis of prostate-specific antigen (PSA), E-cadherin, keratin, and vimentin.. Human prostate cancer LNCaP-LN3 cells growing in culture as multicell spheroids expressed higher levels of E-cadherin and E-cadherin-associated beta-catenin than LNCaP-LN3 cells growing as monolayers. Treatment of cells with DOX downregulated PSA, E-cadherin, and keratin, and upregulated expression of vimentin and vascular endothelial growth factor (VEGF) mRNA. While treatment of cells with IFN-alpha did not alter gene expression, the addition of IFN-alpha to cultures treated with DOX produced synergistic toxicity and abrogated the changes in gene expression observed in cells treated with DOX alone.. Treatment with IFN-alpha and DOX should be further explored as a therapeutic strategy for androgen-insensitive prostate cancer.

Topics: Androgens; Antineoplastic Agents; Cadherins; Cell Differentiation; Cell Transformation, Neoplastic; Doxorubicin; Drug Interactions; Drug Resistance, Neoplasm; Gene Expression Regulation, Neoplastic; Humans; Interferon-alpha; Keratins; Male; Prostate-Specific Antigen; Prostatic Neoplasms; Tumor Cells, Cultured; Up-Regulation; Vimentin

The development of prostate cancer through a multistep process of carcinogenesis may have a long latent period of 20-30 years. It is possible that progression to a malignant state could be blocked or reversed during this time. This study focuses on the ability of the synthetic retinoid, N-(4-hydroxyphenyl)-retinamide (4-HPR), to reverse changes associated with malignant transformation and tumor progression, towards a normal phenotype. To examine the responsiveness of cells at different steps of prostate carcinogenesis, three immortalized, but non-tumorigenic (RWPE-1, WPE1-7 and WPE1-10), and one human prostate carcinoma cell line (DU-145), were used. The effects of 4-HPR on cell proliferation, expression of intermediate filament proteins cytokeratin 18 and vimentin, and tumor suppressor proteins p53 and pRb were examined by immunostaining and compared. Results show that 4-HPR caused inhibition of growth in all cell lines in a dose-dependent manner. 4-HPR induced an increase in staining for cytokeratin 18, a marker of differentiation for prostate epithelial cells. While all cell lines showed strong immunostaining for vimentin, treatment with 4-HPR for 8 days caused a marked decrease in staining for vimentin in all cell lines. In an in vitro assay, 4-HPR also caused inhibition of invasion by DU-145 cells in a dose-dependent manner. Furthermore, 4-HPR treatment was effective in significantly decreasing the abnormal nuclear staining for the tumor suppressor proteins p53 and pRb. Because 4-HPR decreased invasion-associated vimentin expression, inhibited invasion, and normalized p53 and pRb immunostaining, we propose that 4-HPR may be an effective agent for secondary and tertiary prevention, i.e. promotion and progression stages, respectively, of prostate cancer.

Topics: Anticarcinogenic Agents; Cell Count; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Dose-Response Relationship, Drug; Fenretinide; Humans; Immunoenzyme Techniques; Keratins; Male; Neoplasm Invasiveness; Phenotype; Prostate; Prostatic Neoplasms; Retinoblastoma Protein; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Vimentin

Cancer presumably arises from stem cells, preserved in an undifferentiated status since fetal development, or from a dedifferentiation of mature cells that return into a fetal phenotype with the potential for proliferation and renewal. Dedifferentiation in this context could represent a transient phase, passed through by cells, before they switch to redifferentiation, metaplasia or neoplasia. Cytokeratin-7 (CK7) is present in fetal, largely absent in normal adult, and transiently neoexpressed in metaplastic and neoplastic epithelial cells of the stomach according to previous observations. CK7 neoexpression in the stomach could, hence, define a fetal-like, dedifferentiated, cellular phenotype during the development of metaplasia and neoplasia. To test this hypothesis, we investigated CK7 expressions in fetal stomachs, non-neoplastic control stomachs, and neoplastic stomachs exhibiting metaplasia, intraepithelial neoplasia, and early cancer. Proliferation and beta-catenin expression of CK7-positive cells were also evaluated. The chronology of CK7 expression was studied during the experimental gastritis-cancer sequence in Mongolian gerbils. Our results show that metaplastic and neoplastic changes in the gastritis-cancer sequence are related to dedifferentiated epithelial cells which are defined by CK7 expression and can phenotypically be linked to fetal cells at the start of gastric pit development. The dedifferentiated cells exhibit a low proliferation and beta-catenin accumulation, similar to stem cells. Thus, the "stem cell" and "dedifferentiation" hypotheses for cancer origin could complement one another, and dedifferentiation-redifferentiation processes might be decisive for carcinogenesis in the stomach.

Topics: Adenocarcinoma; Adolescent; Adult; Aged; Aged, 80 and over; Animals; Carcinoma in Situ; Cell Transformation, Neoplastic; Child; Disease Models, Animal; Epithelial Cells; Female; Fetus; Gastric Mucosa; Gastritis; Gerbillinae; Gestational Age; Helicobacter Infections; Helicobacter pylori; Humans; Keratin-7; Keratins; Male; Metaplasia; Middle Aged; Stomach; Stomach Neoplasms

Topics: Animals; Carcinoma, Basal Cell; Cattle; Cell Transformation, Neoplastic; Epidermis; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Genetic Predisposition to Disease; Hedgehog Proteins; Keratinocytes; Keratins; Kruppel-Like Transcription Factors; Mice; Mice, Transgenic; Neoplasm Proteins; Neoplasms, Multiple Primary; Promoter Regions, Genetic; Proteins; RNA, Messenger; RNA, Neoplasm; Signal Transduction; Skin Neoplasms; Trans-Activators; Transcription Factors; Zinc Finger Protein Gli2

Disturbances in the regulation of cell proliferation and differentiation play an important role in the formation of neoplastic lesions. Consequently, abnormalities in apoptosis regulation may contribute to this process. Expression of a neoepitope on cytokeratin 18, unmasked by an early caspase cleavage event and recognized by the novel monoclonal antibody M30, is an indicator of early epithelial cell apoptosis. The purpose of this study was to evaluate the quantitative relation among apoptosis (M30), cell persistence (bcl-2), and proliferation (S-phase fraction; SPF) in malignant and benign endometrium.. Using multiparameter DNA flow cytometry on 54 formalin-fixed paraffin-embedded samples from benign (proliferative, secretory, inactive, and hyperplastic endometrium) and malignant (grades 1-3 endometrial adenocarcinoma) endometrial tissue, bcl-2 expression and M30 reactivity were assessed together with the SPF in the cytokeratin-positive epithelial cells.. Benign cyclic endometrium showed a relatively high bcl-2 expression and low M30 reactivity in the proliferative phase whereas in the secretory phase this relation was inverse. In endometrial hyperplasia the expression of bcl-2 was increased compared to that in secretory and postmenopausal endometrium, but still below the level of proliferative samples. The expression of M30 also increased compared to normal proliferative endometrium but did not reach the level of endometrium in the secretory phase of the menstrual cycle. In cancer the expression of bcl-2 decreased with the progression of differentiation grade. For M30 expression this relation was inverse. Overall there was a significant increase of M30 reactivity in cancerous compared to hyperplasia and normal cyclic endometrium.. Transition of endometrial epithelium from hyperplasia to cancer seems to involve both increased apoptosis and decreased bcl-2 expression. Flow cytometric evaluation of M30 and bcl-2 expression levels, with the SPF, in currettage specimens from postmenopausal patients complaining of bleeding provides a quantitative assessment of endometrial apoptosis, anti-apoptosis, and proliferation. Further studies are needed to determine the relationship among these three processes as indicators of the biological behavior of gynecological tumors.

Topics: Adult; Aged; Aged, 80 and over; Apoptosis; Biomarkers, Tumor; Cell Transformation, Neoplastic; Endometrial Hyperplasia; Endometrial Neoplasms; Female; Flow Cytometry; Humans; Keratins; Middle Aged; Precancerous Conditions; Proto-Oncogene Proteins c-bcl-2; S Phase

A unique series of epidermal cell lines representing different stages of malignant transformation were spontaneously derived from a single adult immunosuppressed individual. Four keratinocyte lines (PM1-4) established from forehead skin are here compared with 4 squamous cell carcinoma (SCC) lines (MET1-4) derived respectively from a primary cutaneous tumour, two local recurrences and a distant metastasis of invasive SCC. Despite altered growth properties, the PM lines retained many features of normal keratinocytes including keratin phenotype, differentiation capacity and non-tumorigenicity in athymic mice. In contrast, from early passage, the MET lines displayed markedly reduced growth requirements, abnormal differentiation, aberrant K18 expression and tumorigenicity in athymic mice. The abnormal keratin profile of individual MET lines closely reflected the keratin phenotype of the tumour of origin. Although unusual HPV types were identified in the original tissue, there was no evidence of persistent virus within any cell line and it appears that HPV is not critical for maintenance of the immortal phenotype. The PM lines were distinctly different from invasive SCC lines and are likely to be useful for studies of mutations important early in neoplastic progression. The SCC series represent primary, recurrent and metastatic carcinoma. Availability of such a series from the same individual will facilitate genetic analysis of the malignant process.

Topics: Adaptation, Physiological; Adult; Animals; Carcinogenicity Tests; Carcinoma, Squamous Cell; Cell Differentiation; Cell Division; Cell Line; Cell Transformation, Neoplastic; Epidermis; Face; Humans; Keratinocytes; Keratins; Male; Mice; Mice, Nude; Neoplasm Staging; Neoplasm Transplantation; Papillomaviridae; Skin Neoplasms

To investigate the molecular basis of oncogenesis induced by the v-jun oncogene of avian sarcoma virus 17 (ASV17), we developed a conditional cell transformation system in which transcription of the ASV17 v-jun allele is controlled by a doxycycline-sensitive transactivator (tTA) or a reverse (doxycycline-dependent) transactivator (rtTA), respectively. Permanent cell lines of quail embryo fibroblasts conditionally transformed by a doxycycline-controlled v-jun allele revert to the normal phenotype within 3 days and lose their ability to grow in soft agar, strictly dependent on the addition or removal of the drug, respectively. The reverted cells are rapidly retransformed on conditional activation of v-jun. While full-level synthesis of v-jun mRNA and v-Jun protein in these cells is established within 2 and 14 h, respectively, after switching to the permissive conditions, the first morphological alterations are observed after 24 h, and as early as 2 days later the morphology has changed entirely from flat cells resembling normal fibroblasts to spindle-shaped fusiform cells showing a typical jun-transformed phenotype. Kinetic expression analysis revealed that transcriptional activation of the direct jun target gene BKJ precisely coincides with the establishment of full-level v-Jun protein synthesis. Furthermore, we have analyzed the expression of a novel candidate v-jun target gene, termed JAC, which shows no sequence homology to known genes. Similar to BKJ, JAC is specifically activated in jun-transformed fibroblasts, and induction of JAC is tightly linked to the conditional expression of oncogenic v-Jun. These results demonstrate the high stringency of the doxycycline-controlled v-jun expression system, and they also indicate that expression of v-jun in these cells is indispensable for enhanced proliferation, cell transformation, and the induction of specific expression patterns of downstream target genes.

Topics: Alleles; Animals; Avian Sarcoma Viruses; Cell Division; Cell Line; Cell Size; Cell Transformation, Neoplastic; Coturnix; Doxycycline; Fibroblasts; Gene Expression Regulation, Neoplastic; Genes, jun; Genetic Vectors; Keratins; Kinetics; Oncogene Protein p65(gag-jun); Phenotype; RNA, Messenger; Transcriptional Activation; Transfection; Tumor Stem Cell Assay

The telomere and the telomerase in human esophageal cancer are not yet completely understood. The regulatory mechanism of telomerase activity and telomere dynamics has drawn considerable attention. It is generally assumed that when telomerase has been activated, no further telomere shortening should ensue; however, a much more complex pattern of telomere dynamics may exist in telomerase-positive cancer cells. A novel human esophageal cancer cell line (KAN-ES) was established and characterized. Using KAN-ES and its serially passaged subclones up to the 55th generation, we determined the alteration of telomere length (TRF), telomerase activity (TA), telomerase RNA expression (hTR), population doubling time, karyotype, and cytokeratin 14 expression during the process of establishing a cancer cell line. We found that the TRF was maintained between 4.0 and 5.0 kb during the serial passages, despite sustained high TA (assessed by an in vitro TRAP assay). No close relationships were found among TRF, TA, and hTR expression. TA and telomere dynamics were not associated with cellular growth ability and differentiation. However, the number of population doublings showed significant correlations with both the TA and doubling times. In conclusion, these dissociations between telomere dynamics and TA support the existence of additional controls on TRF in cancer cells. KAN-ES and its restored subclones should prove a valuable resource for esophageal cancer research.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Division; Cell Transformation, Neoplastic; Esophageal Neoplasms; Gene Expression Regulation, Neoplastic; Humans; Keratins; Male; Middle Aged; RNA, Messenger; Telomerase; Telomere; Tumor Cells, Cultured

The erbB family of receptor tyrosine kinases, which consists of the epidermal growth factor receptor (EGFr/erbB1), erbB2 (neu), erbB3 and erbB4, has been shown to be important for both normal development as well as neoplasia. The expression of rat erbB2 was targeted to the basal layer of mouse epidermis with the bovine keratin 5 promoter. Overexpression of wild type rat erbB2 in the basal layer of epidermis led to alopecia, follicular hyperplasia and sebaceous gland enlargement as well as hyperplasia of the interfollicular epidermis. Spontaneous papillomas, some of which converted to squamous cell carcinomas, arose in homozygous erbB2 transgenic mice as early as 6 weeks of age with >90% incidence by 6 months. Analysis of several proliferation/differentiation markers indicated that erbB2 overexpression led to epidermal hyperproliferation and a possible delay in epidermal differentiation. Transgenic mice were also hypersensitive to the proliferative effects of the skin tumor promoter, 12-0-tetradecanoylphorbol-13-acetate (TPA) and were more sensitive to two-stage carcinogenesis. Elevations in EGFr and erbB2 protein as well as erbB2:EGFr and erbB2:erbB3 heterodimers were observed in skin of the erbB2 transgenic mice. Phosphotyrosine levels of the EGFr, erbB2 and erbB3 proteins were also elevated. These results indicate an important role for erbB2 signaling in epidermal growth, development and neoplasia. Oncogene (2000) 19, 4243 - 4254

Topics: Animals; Carcinogens; Carcinoma, Squamous Cell; Cattle; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Cocarcinogenesis; Dimerization; Disease Progression; Epidermis; ErbB Receptors; Female; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Genes, erbB-2; Genes, ras; Genes, Synthetic; Hyperplasia; Keratins; Male; Mice; Mice, Inbred ICR; Mice, Transgenic; Neoplasm Proteins; Papilloma; Phosphorylation; Promoter Regions, Genetic; Protein Processing, Post-Translational; Rats; Receptor, ErbB-2; Receptor, ErbB-3; Recombinant Fusion Proteins; Signal Transduction; Skin Neoplasms; Tetradecanoylphorbol Acetate; Transgenes

Diploid tumour cells regularly continue to progress after the development of aneuploid cell populations in head and neck squamous cell carcinomas. The coexistence of aneuploid clones with their diploid progenitor cells provides a unique opportunity to study the order of appearance of p53 mutation and aneuploidy in the same tumour. Multiparameter flow cytometry was therefore applied to 22 oral squamous cell carcinomas to simultaneously assess cellular DNA content and p53 protein expression on a single-cell basis. Concurrent measurements of cytokeratin expression served to identify tumour cells of epithelial origin. One of 5 diploid and 2 of 17 aneuploid carcinomas were p53-negative. For 15 p53-positive aneuploid tumours, overexpression of p53 protein was identified for the aneuploid clones as well as for coexisting diploid tumour cell populations in 14 cases. On the understanding that coexisting diploid and aneuploid tumour cell populations have a common clonal origin, these results provide evidence that aneuploid tumour clones typically develop from p53-deficient diploid progenitor cells. Loss of wild-type p53 function may therefore contribute to the development of aneuploidy in head and neck cancer.

Topics: Aneuploidy; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Clone Cells; Cohort Studies; Diploidy; Disease Progression; DNA, Neoplasm; Flow Cytometry; Genes, p53; Humans; Keratins; Mouth Neoplasms; Neoplasm Proteins; Neoplastic Stem Cells; Pharyngeal Neoplasms; Tumor Suppressor Protein p53

9-cis beta-Carotene was extracted from a commercial extract of the algae Dunaliella salina (Betatene), and its actions on proliferation and gene expression were examined in murine 10T1/2 cells and human HaCaT keratinocytes. The 9-cis isomer was less active than all-trans beta-carotene in reducing proliferation and in upregulating expression of connexin 43 in 10T1/2 cells. However, it had comparable ability to suppress carcinogen-induced neoplastic transformation. When tested in HaCaT cells in organotypic culture, it was less active in inducing connexin 43 expression and suppressing expression of keratin K1. In this assay the all-trans isomer was highly active at 10(-6) M, whereas 10(-5) M 9-cis beta-carotene was required to produce a comparable effect. Only small reductions in expression of the basal keratin 5 were seen. All-trans and 9-cis retinoic acids, potential metabolites of beta-carotene isomers, were studied in the same systems. In contrast to the carotenoids, the 9-cis isomer of retinoic acid was approximately 10-fold more active in suppressing neoplastic transformation and inducing connexin 43 expression in both cell types than the all-trans isomer. The retinoic acid isomers were about equipotent in suppressing K1 expression. Cellular levels of 9-cis beta-carotene were approximately 3.5-fold lower than levels of all-trans beta-carotene, suggesting that part, but not all, of this decreased activity of the 9-cis isomer was due to decreased cell uptake. Thus 9-cis beta-carotene is consistently less active than the all-trans isomer; that 9-cis retinoic acid is, in general, much more potent than the all-trans isomer suggests little or no conversion from the carotenoid to the retinoid under these culture conditions.

Topics: Animals; beta Carotene; Cell Division; Cell Line; Cell Transformation, Neoplastic; Chlorophyta; Chromatography, High Pressure Liquid; Connexin 43; Gene Expression; Humans; Immunoblotting; Isomerism; Keratinocytes; Keratins; Kinetics; Mice; Tumor Cells, Cultured

The modulation of cytokeratin 18 during tumor transformation in hepatoma had been previously recognized through a series of biochemical and immunological approaches. Expression of cytokeratin 18 in transitional cell carcinoma comparing with hepatoma was investigated using the hepatoma transformation model. CK18 related molecules were found. In the present study, we design various epithelial cancers with the same model. CK18 related molecules were all evident. Therefore, we suggest that CK18 related proteins would play an important role in tumorigenesis of epithelial cancers.

Topics: Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Female; Humans; Keratins; Liver Neoplasms; Male; Molecular Weight; Neoplasm Proteins; Neoplasms, Glandular and Epithelial

To clarify the correlation between multidirectional differentiation and aggressiveness of endometrial adenocarcinomas, we assessed both proliferative activities (PA) using Ki-67 expression and squamous and/or endocrine differentiation. We divided 51 adenocarcinomas into 22 adenocarcinomas with typical squamous differentiation (>/=10% of tumor cells, typical SQ) classified into 10 adenoacanthomas (AA) and 12 adenosquamous carcinomas (AS), 17 adenocarcinomas with focal squamous differentiation (<10% of tumor cells), and 12 typical adenocarcinomas without morphological squamous differentiation (pure AC), according to the new WHO classification. Paraffin-embedded sections were stained using monoclonal antibodies against high-molecular-weight keratins (HMWK) to recognize squamous cells, chromogranin A to recognize endocrine cells, and Ki-67 antigen to recognize proliferating cells. Both AA and AS exhibited lower PA than pure AC. Typical SQ exhibited lower PA than pure AC. This difference was also significant after selecting only grade 1 or stage I/II cases. AA exhibited lower PA than AS and also after selecting only grade 1 or stage I/II cases. PA of adenocarcinoma with the expression of HMWK in >/=30% of tumor cells was lower than those without HMWK. PA of adenocarcinoma with the expression of chromogranin A in >/=10% of tumor cells was lower than those without chromogranin A. These differences were also significant after selecting only grade 1 or stage I/II cases. Squamous and/or endocrine differentiation is a good marker for a reduction of PA. Endometrial adenocarcinomas with multidirectional differentiation exhibited lower PA and were likely to be more mature than those with monodirectional differentiation.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Carcinoma, Adenosquamous; Cell Transformation, Neoplastic; Chromogranin A; Chromogranins; Endometrial Neoplasms; Female; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Metaplasia; Middle Aged; Neoplasm Invasiveness

Comparative investigations of odontogenic cells in normally forming teeth and tumors may provide insights into the mechanisms of the differentiation process. The present study is devoted to late phenotypic markers of ameloblast and odontoblast cells, i.e., proteins involved in biomineralization. The in situ expression of amelogenins, keratins, collagens type III and IV, vimentin, fibronectin, osteonectin, and osteocalcin was performed on normal and tumor odontogenic human cells. The pattern of protein expression showed some similarities between ameloblasts and odontoblasts present in normally developing human teeth and cells present in neoplastic tissues of ameloblastic fibroma, ameloblastic fibro-odontomas, and complex odontomas. Amelogenins (for ameloblasts) and osteocalcin (for odontoblasts) were detected in cells with well-organized enamel and dentin, respectively. In contrast, "mixed" cells located in epithelial zones of mixed odontogenic tumors co-expressed amelogenins and osteocalcin, as shown by immunostaining. The presence of osteocalcin transcripts was also demonstrated by in situ hybridization in these cells. Keratins and vimentin were detected in the same epithelial zones. Tumor epithelial cells were associated with various amounts of polymorphic matrix (amelogenin- and osteocalcin-immunoreactive), depending on the types of mixed tumors. No osteocalcin labeling was found in epithelial tumors. This study confirms that the differentiation of normal and tumor odontogenic cells is accompanied by the expression of some common molecules. Furthermore, the gene products present in normal mesenchymal cells were also shown in odontogenic tumor epithelium. These data may be related to a tumor-specific overexpression of the corresponding genes transcribed at an undetectable level during normal development and/or to an epithelial-mesenchymal transition proposed to occur during normal root formation. A plausible explanation for the results is that the odontogenic tumor epithelial cells are recapitulating genetic programs expressed during normal odontogenesis, but the tumor cells demonstrate abnormal expression patterns for these genes.

Topics: Ameloblastoma; Amelogenin; Cell Differentiation; Cell Polarity; Cell Transformation, Neoplastic; Dental Enamel Proteins; Epithelial Cells; Extracellular Matrix Proteins; Gene Expression Regulation, Developmental; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization; Infant, Newborn; Keratins; Odontogenesis; Odontogenic Tumors; Odontoma; Osteocalcin; Osteonectin; Tumor Cells, Cultured; Vimentin

Ovarian carcinomas are thought to arise in the ovarian surface epithelium (OSE). Although this tissue forms a simple epithelial covering on the ovarian surface, OSE cells exhibit some mesenchymal characteristics and contain little or no E-cadherin. However, E-cadherin is present in metaplastic OSE cells that resemble the more complex epithelia of the oviduct, endometrium and endocervix, and in primary epithelial ovarian carcinomas. To determine whether E-cadherin was a cause or consequence of OSE metaplasia, we expressed this cell-adhesion molecule in simian virus 40-immortalized OSE cells. In these cells the exogenous E-cadherin, all three catenins, and F-actin localized at sites of cell-cell contact, indicating the formation of functional adherens junctions. Unlike the parent OSE cell line, which had undergone a typical mesenchymal transformation in culture, E-cadherin-expressing cells contained cytokeratins and the tight-junction protein occludin. They also formed cobblestone monolayers in two-dimensional culture and simple epithelia in three-dimensional culture that produced CA125 and shed it into the culture medium. CA125 is a normal epithelial-differentiation product of the oviduct, endometrium, and endocervix, but not of normal OSE. It is also a tumor antigen that is produced by ovarian neoplasms and by metaplastic OSE. Thus, E-cadherin restored some normal characteristics of OSE, such as keratin, and it also induced epithelial-differentiation markers associated with weakly preneoplastic, metaplastic OSE and OSE-derived primary carcinomas. The results suggest an unexpected role for E-cadherin in ovarian neoplastic progression.

Topics: Actins; Cadherins; Cell Differentiation; Cell Line, Transformed; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Epithelial Cells; Female; Humans; Keratins; Mesoderm; Metaplasia; Ovary; Simian virus 40

The exact pathogenesis for prostate cancer is not known. Progress made in prostate cancer research has been slow, largely due to the lack of suitable in vitro models. Here, we report our work on the immortalization of a human prostate epithelial cell line and show that it can be used as a model to study prostate tumorigenesis.. Replication-defective retrovirus harboring the human papillomavirus (HPV) type 16 E6 and E7 open reading frames was used to infect primary human prostate epithelial cells. Polymerase chain reaction, followed by Southern hybridization for the HPV 16 E6/E7, Western blot for prostatic acid phosphatase, telomeric repeat amplification protocol assay for telomerase activity, two-dimensional gels for cytokeratins, and cytogenetic analysis were undertaken to characterized the infected cells.. The retrovirus-infected cell line, HPr-1, continued to grow in culture for more than 80 successive passages. Normal primary cells failed to proliferate after passage 6. HPr-1 cells bore close resemblance to normal primary prostate epithelial cells, both morphologically and biochemically. However, they possessed telomerase activity and proliferated indefinitely. Cytogenetic analysis of HPr-1 cells revealed a human male karyotype with clonal abnormalities and the appearance of multiple double minutes.. The HPr-1 cells expressed prostatic acid phosphatase and cytokeratins K8 and K18, proving that they were prostate epithelial cells. They were benign in nude mice tumor formation and soft agar colony formation assay. The HPr-1 cell line is an in vitro representation of early prostate neoplastic progression.

Topics: Acid Phosphatase; Adolescent; Animals; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Chromosome Aberrations; Epithelial Cells; Genetic Vectors; Humans; Karyotyping; Keratins; Male; Mice; Mice, Nude; Oncogene Proteins, Viral; Open Reading Frames; Papillomaviridae; Papillomavirus E7 Proteins; Prostate; Repressor Proteins; Retroviridae; Telomerase; Transplantation, Heterologous; X Chromosome; Y Chromosome

Carcinogenesis involves the accumulation of genetic changes within a single cell. Tumor promotion functions in the initial clonal expansion of an initiated cell but is generally not considered to influence later stages. To investigate whether tumor promotion can influence later stages of carcinogenesis we developed a two-hit 7, 12-dimethylbenz[a]anthracene (D) protocol designed to enrich for keratinocytes that contain at least two D-induced genetic alterations. FVB/N mice were initiated with D and promoted with 12-O-tetradecanoylphorbol-13-acetate (T) or treated with acetone (A) vehicle for 6 weeks. At 7 weeks after the start of promotion, but before visible papilloma development, groups of mice were treated with a second dose of D or A and 1 week later T promotion was resumed. D/T/A/T mice developed 2.8 papillomas/mouse and D/A/D/T mice demonstrated an additive tumor response and developed 5.8 papillomas/mouse. Importantly, D/T/D/T mice developed 12.4 papillomas/mouse, thereby demonstrating a synergistic tumor response compared with D/A/D/T and D/T/A/T mice. D/T/D/T papillomas exhibited increases in suprabasal S phase cells and keratin 13 expression when compared with D/T/A/T papillomas. D/T/D/T mice developed squamous cell carcinomas (SCCs) 10 weeks earlier than D/T/A/T mice and demonstrated a 96% malignancy incidence and 1.71 SCC/mouse compared with D/T/A/T mice, which demonstrated a 28% malignancy incidence and 0.32 SCC/mouse. Greater than 90% of D/T/A/T and D/T/D/T papillomas and SCCs contained mutant Ha-ras, while a normal Ha-ras allele persisted in all cases, indicating that a gene other than the remaining normal allele of Ha-ras was a target gene for the second D hit. These data demonstrate that: (i) promotion between the first and second hits has a profound outcome on carcinogenesis, presumably by increasing the probability that a second hit will occur in a previously initiated cell; (ii) continued promotion after the second hit is required for full expression of malignancy; (iii) the classic initiation-promotion protocol can be extended to a multihit, multistage model.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Acetone; Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cocarcinogenesis; DNA Mutational Analysis; DNA, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Genes, ras; Keratinocytes; Keratins; Mice; Models, Biological; Neoplasm Proteins; Papilloma; Polymerase Chain Reaction; Polymorphism, Single-Stranded Conformational; S Phase; Skin Neoplasms; Tetradecanoylphorbol Acetate

Disruption of the retinoblastoma (RB) tumor suppressor pathway is a common and important event in breast carcinogenesis. To examine the role of the retinoblastoma protein (pRB) in this process, we created human mammary epithelial cells (HMEC) deficient for pRB by infecting primary outgrowth from breast organoids with the human papillomavirus type 16 (HPV16) E7 gene. HPV16 E7 binds to and inactivates pRB and also causes a significant down-regulation of the protein. Culturing normal HMEC in a reconstituted basement membrane (rBM) provides a correct environment and signaling cues for the formation of differentiated, acini-like structures. When cultured in this rBM, HMEC+E7 were found to respond morphologically as normal HMEC and form acinar structures. In contrast to normal HMEC, many of the cells within the HMEC+E7 structures were not growth arrested, as determined by a 5-bromo-2'-deoxyuridine incorporation assay. pRB deficiency did not affect polarization of these structures, as indicated by the normal localization of the cell-cell adhesion marker E-cadherin and the basal deposition of a collagen IV membrane. However, in HMEC+E7 acini, we were unable to detect by immunofluorescence microscopy the milk protein lactoferrin or cytokeratin 19, both markers of differentiation expressed in the normal HMEC structures. These data suggest that loss of RB in vivo would compromise differentiation, predisposing these cells to future tumor-promoting actions.

Topics: Breast; Cadherins; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Collagen; Epithelial Cells; Extracellular Matrix; Humans; Keratins; Lactoferrin; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Retinoblastoma Protein; Transduction, Genetic

To evaluate cell proliferative activity and expression of cytokeratins (CKs) and epithelial membrane antigen (EMA) in intraductal papillary-mucinous neoplasm of the pancreas (IPNP).. We examined cell proliferative activity in normal pancreatic ducts, IPNP, and invasive ductal adenocarcinoma of the pancreas by immunohistochemistry for proliferating cell nuclear antigen (PCNA) and Ki67 antigen. Expression of CKs and EMA was also examined immunohistochemically.. In normal pancreas (n = 5), PCNA- or Ki67-positive ductal epithelia were not found. Cytokeratins (polyclonal, CAM5.2, CK-7, CK-8, CK-18, and CK-19) were expressed in the ducts and ductules but not in the acinus, and EMA expression was noted in the acinus but rarely in the ducts. In IPNP (n = 9) and invasive ductal adenocarcinoma (n = 6) of the pancreas, the overall PCNA-labeling index (PCNA-LI) was 3.2 +/- 4.1 and 16.0 +/- 7.2, respectively, and overall Ki67-LI was 2.2 +/- 2.6 and 14.5 +/- 6.3, respectively. In IPNP, the PCNA-LI and Ki67-LI were low in adenoma areas (PCNA-LI = 0.9 +/- 0.7), intermediate in dysplastic areas (PCNA-LI = 3.7 +/- 2.4), and rather high in carcinoma in situ areas (PCNA-LI = 11.5 +/- 8.4). Both CKs and EMA were noted in tumor cells.. The data suggest that cell proliferative activity is low in IPNP compared to invasive ductal adenocarcinoma, that cell proliferative activity increases with the grade of cell atypia in IPNP, and that CK expression is not changed during the neoplastic change, but EMA is newly expressed or overexpressed during the neoplastic change.

Topics: Aged; Carcinoma, Ductal, Breast; Carcinoma, Intraductal, Noninfiltrating; Cell Division; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Ki-67 Antigen; Male; Middle Aged; Mucin-1; Pancreatic Neoplasms; Proliferating Cell Nuclear Antigen

A cell line, designated ISOS-1, was established from a tumor formed by transplantation of a human angiosarcoma into mice with severe combined immunodeficiency (SCID). The cells showed endothelial properties, based on the uptake of Dil-Ac-LDL and binding of UEA-I/GSA-I lectins, but were negative for CD11b and Pan Cytokeratin. However, the cells lost differentiated characteristics such as expression of von Willebrand factor, contact inhibition growth and tube formation activity. These findings indicate that ISOS-1 is a poorly-differentiated endothelial cell line. At the 81st passage, all of the cells were positive for H-2Dd in various intensity, but not HLA-ABC. The metaphase chromosomes consistently showed a characteristic mouse, but not human, telocentric form. Furthermore, this cell line produced fatal tumor growth in SCID mice and also in BALB/c mice. These results suggest that ISOS-1 is a murine-phenotypic angiosarcoma cell line.

Topics: Animals; CD11 Antigens; Cell Division; Cell Transformation, Neoplastic; Chromosomes; Female; H-2 Antigens; Hemangiosarcoma; Histocompatibility Antigens Class I; Humans; Immunohistochemistry; Keratins; Lectins; Lipoproteins, LDL; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; Mice, Inbred Strains; Mice, SCID; Neoplasm Transplantation; Phenotype; Skin Neoplasms; Tumor Cells, Cultured; von Willebrand Factor

Tubulopapillary hidradenoma is a benign sweat gland tumor that appears as a well-defined, superficially located dermal nodule. It combines ductal as well as apocrine and eccrine glandular differentiation. Microscopically, the tumor is composed of tubular structures that characteristically show intraluminal non-villous papillary projections and a peripheral myoepithelial cell layer. A tumor that is histologically and immunohistochemically identical to tubulopapillary hidradenoma occurred in the mandible of a 73-year-old man and resulted in considerable diagnostic difficulty. The neoplasm developed in a mandibular cyst and recurred 5 years after initial enucleation. This is the first report of a central (intraosseous) sweat gland adenoma of the mandible. The differential diagnosis and possible histogenesis are discussed.

Topics: Actins; Adenoma, Sweat Gland; Aged; Apocrine Glands; Bone Cysts; Cell Nucleolus; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Diagnosis, Differential; Eccrine Glands; Epithelial Cells; Follow-Up Studies; Humans; Keratins; Male; Mandibular Neoplasms; Muscle, Smooth; Neoplasm Recurrence, Local; Vimentin

A rare case of primary squamous cell carcinoma surrounding Stensen's duct in a 75-year-old man is presented. The tumor was a relatively well-defined, hard, subcutaneous mass, measuring 18 x 14 x 9 mm and situated in the right cheek. Microscopic examination of an excisional biopsy specimen revealed tumor cells showing squamous differentiation, a papillary growth pattern, and ductal structures with comedo necrosis. Immunohistochemical studies showed positive reactivity for KL-1 (cytokeratin, monoclonal), epithelial membrane antigen, and carcinoembryonic antigen in some tumor cells. The origin of the tumor was thought to be the accessory parotid gland duct epithelium.

Topics: Aged; Carcinoembryonic Antigen; Carcinoma, Squamous Cell; Cell Differentiation; Cell Nucleolus; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Epithelium; Humans; Keratins; Male; Mucin-1; Necrosis; Parotid Gland; Parotid Neoplasms; Salivary Ducts

Renal cell carcinoma (RCC) arising in acquired cystic kidney disease (ACKD) is considered to be a tumor of low malignant potential, compared with classic RCC. The aim of the present study was to identify any significant differences in the antigenic profiles or tumor cell proliferative activity of ACKD-associated RCC and classic RCC that might be responsible for differences in their biologic behavior. We studied the immunohistochemical profiles and proliferative activity of 12 classic RCCs and 5 ACKD-associated RCCs with markers of proximal tubules (Leu M1, alpha-1 antitrypsin, CAM 5.2), markers of distal tubules (Arachis hypogaea lectin, AE1/AE3, epithelial membrane antigen [EMAJ, CAM 5.2), vimentin, and proliferating cell nuclear antigen (PCNA). We performed proliferation analysis with the CAS 200 image analysis system. For each case, 8 to 20 fields of tumor tissue in the areas of maximal PCNA staining were quantitated, and the percentage of PCNA-positive nuclear area for each individual tumor was calculated. All of the five ACKD-associated RCCs expressed AE1/AE3, EMA, and CAM 5.2 in more than 50% of the tumor cells. Arachis hypogaea lectin was significantly expressed in three of the five ACKD-associated RCCs. Leu M1 and alpha-1 antitrypsin reacted with fewer than 10% of the tumor cells in all of the five ACKD-associated RCCs. In contrast, the 12 classic RCCs showed expression of CAM 5.2 in 11 cases, alpha-1 antitrypsin in 10 cases, Leu M1 in 9, EMA in 8, and AE1/AE3 in 3 cases in more than 50% of the tumor cells and a totally negative reaction with Arachis hypogaea lectin in 8 cases, EMA in 4, AE1/AE3 in 4, and vimentin in 5 cases. Although coexpression of proximal and distal tubule markers was seen in some cases of RCC in either category, there was uniform and strong staining for distal tubule markers in ACKD-associated RCC and for proximal tubule markers in classic RCC. The mean percentage of PCNA-positive nuclear area for the ACKD-associated RCCs (2.41%) was significantly (P < .05) less than that of the classic RCCs (21.42%). The differences in expression of proximal and distal tubule markers and proliferative activity might be responsible for the differences in the biologic behavior of ACKD-associated RCC and classic RCC.

Topics: Adult; Aged; Aged, 80 and over; alpha 1-Antitrypsin; Analysis of Variance; Biomarkers, Tumor; Carcinoma, Renal Cell; Cell Nucleus; Cell Transformation, Neoplastic; Female; Humans; Image Processing, Computer-Assisted; Immunohistochemistry; Keratins; Kidney Diseases, Cystic; Kidney Neoplasms; Kidney Tubules; Lewis X Antigen; Male; Middle Aged; Mucin-1; Peanut Agglutinin; Proliferating Cell Nuclear Antigen; Vimentin

Our earlier studies demonstrated neoplastic transformation of SV40-immortalized neonatal human prostate epithelial cells (267B1) by fractionated doses of ionizing radiation or by introduction of v-ki-ras oncogene. X-ray-treated 267B1 cells represent three different stages of neoplastic progression: nontumorigenic F3-SAC cells that acquired morphological changes and anchorage independence when treated with 2 x 2 Gy of X-rays; malignantly transformed 267B1-XR and 267B1-SXR cells that received 2-Gy doses to a total of 30 Gy. We also reported alterations in cell size, morphology, actin stress fibers, and levels of actin-binding proteins in these transformed human prostate cells.. We analyzed intermediate filament-nuclear matrix (IF-NM) protein expression in the various 267B1 cells as a consequence of neoplastic progression by two-dimensional gel electrophoresis and immunofluorescence.. Our present study revealed that the 267B1 cells experienced progressive changes in their intermediate filament protein composition during the process of neoplastic conversion, achieved either by X-rays or by ras-oncogene. In particular, we observed a stepwise downregulation of cytokeratin-19 in these in vitro transformed 267B1 cells.. Our data suggest that loss of expression of cytokeratin-19 accompanied the morphological alterations associated with in vitro neoplastic transformation of SV40-immortalized prostate epithelial cells.

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Disease Progression; Epithelial Cells; Humans; In Vitro Techniques; Intermediate Filament Proteins; Keratins; Male; Nuclear Matrix; Nuclear Proteins; Prostatic Neoplasms; Tumor Cells, Cultured

The applicability of the experimental skin carcinogenesis model for studies of tumor development was examined by exposing the skin of various mouse strains to different chemical carcinogens and UV radiation regimens, in order to analyze the development and progression of the neoplastic process and the role of differentiation markers such as keratins. In tumor-sensitive hairy NMRI mouse skin, the chemical carcinogen, 7,12-dimethylbenz(a)-anthracene (DMBA) induced an abnormal epidermal cell differentiation and structural irregularities associated with an altered keratin expression, as well as numerous papillomas and squamous cell carcinomas. A suboptimal dose of UVB irradiation increased the number of DMBA-induced benign squamous neoplasms. Low doses of benzo(a)pyrene resulted in mild epidermal alterations, but only in one tumor. High doses of UVB induced a large number of undifferentiated spindle cell tumors with few keratinpositive cells in NMRI mice, similar though fewer tumors in hairy, heavily pigmented C57BL/6 mice, numerous papillomas and squamous cell carcinomas in hairless hr/hr mice but only two papillomas in hairy, moderately pigmented DBA/2 mice while UVA exposure produced only two papillomas in hairless SKH-1 mice. In conclusion, the extent and type of skin tumor development depended upon the induction regimen: physical, chemical, dose and duration, as well as on the skin structure: pigmentation and adnexal development, all of which have to be taken into account when relating experimental results to human conditions.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Biomarkers, Tumor; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Disease Models, Animal; Female; Keratins; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Inbred Strains; Skin Neoplasms

Somatic mutations of the K-ras oncogene play an important role in colorectal carcinogenesis. We determined whether rat colon epithelial cells could be transformed by introducing retroviruses carrying the activated human K-ras oncogene alone. Primary epithelial cells from the rat distal colon were infected with retroviruses carrying wild-type and two types of activated K-ras (asp and val at codon 12) cDNAs. Cells infected with the wild-type K-ras virus showed no change in morphology and died within 3 weeks, whereas the activated K-ras virus-infected cells underwent morphological changes within 3 days and continued to proliferate. From these cells, several cell lines were subsequently established. Epithelial cells transformed by activated K-ras formed colonies in soft agar culture and tumors in athymic nude mice. Multiple copies of human K-ras genes and large amounts of K-ras mRNAs and proteins were found in the transformed cells. These data suggest that overexpression of activated K-ras transforms rat colon epithelial cells.

Topics: Animals; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Colon; Colonic Neoplasms; Epithelial Cells; Female; Genes, ras; Humans; Keratins; Mice; Mice, Nude; Neoplasm Transplantation; ras Proteins; Rats; Rats, Inbred F344; Retroviridae; RNA, Messenger; Transfection

This study aimed to ascertain whether cathepsin D expression could be related to the stage of differentiation of oral tumors.. Human oral biopsies of 10 squamous cell carcinomas and of the corresponding perilesional normal tissues were used. The tumors had all been clinically graded as advanced stage but nonmetastatic; five were classified histopathologically as poorly differentiated.. The gene expression of cathepsin D and keratin K13 in the biopsies was measured by reverse transcription polymerase chain reaction. Ratios of tumor-to-control readings helped compensate for sample variability.. Keratin K13, as a suprabasal cell marker, tended to confirm the histological grading of the tumors (but was not otherwise useful in distinguishing tumors from normal tissue). Substantial overexpression of cathepsin D was found in the poorly differentiated tumors.. Cathepsin D overexpression is considered a prognostic indicator of metastasis. In this sample, it was also associated with dedifferentiation. Cathepsin D might serve as a valuable gauge in clinical exploration of the connection between dedifferentiation and metastasis.

Topics: Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cathepsin D; Cell Transformation, Neoplastic; Female; Humans; Keratins; Male; Middle Aged; Mouth Neoplasms; Polymerase Chain Reaction; Prognosis; RNA-Directed DNA Polymerase; RNA, Messenger

The presence of specific keratins can be of diagnostic value for studying normal and neoplastic epithelium of the vulva. The aim of the present study was to investigate normal, preneoplastic and neoplastic epithelium of the vulva. Keratins 5, 6 and 18, identified by a polyspecific anti-human CK antibody (clone LP 34, DAKO), and the keratin subtypes 7, 10, 14, 18, 19 and 20 of normal, dysplastic and malignant vulval epithelium (paraffin-embedded sections) were detected by immunohistochemical APAAP staining. Keratins 5, 6, and 18 (clone LP 34) and keratin subtype 10 are expressed in the upper third of the normal vulval epithelium. In mild and moderate intraepithelial neoplasia only a few cells express these keratins. In patients with severe intraepithelial neoplasia (VIN III) the expression of these keratins seems to be associated with recurrence of the disease. In biopsy specimens of patients without recurrence we find positive results for keratins 5, 6 and 18 (clone LP 34) and keratin 10. If patients have a recurrence of the disease, expression of these keratins is only diffuse or is absent. The expression of these keratin subtypes in vulval carcinomas is mostly seen in differentiated cells. There was no association between recurrence and keratin pattern. We have not found any other expression of the tested keratin subtypes in VIN and in vulval carcinoma.

Topics: Biomarkers, Tumor; Biopsy; Carcinoma in Situ; Cell Transformation, Neoplastic; Epithelium; Female; Humans; Immunoenzyme Techniques; Keratins; Neoplasm Recurrence, Local; Precancerous Conditions; Prognosis; Vulva; Vulvar Neoplasms

Juvenile pleomorphic adenoma of the parotid gland represents an extremely rare tumour entity and is comparable to congenital tumours of the salivary glands concerning its embryonal structure. The clinical detection of the tumour in a 7-year-old girl does not exclude that the tumour had developed either earlier or immediately after the birth. The high cellularity and the evidence of primitive epithelial and myoepithelial cellular structures do not justify its classification as a malignant tumour. However the presence of embryonal tissue structures associated with the end of the third month of embryogenesis is characterized by more solid cell formations in partly verticulate arrangement. The absence of further differentiation into lobular structures and differentiated duct or acinic cell formations may be due to cell arrest. Differential diagnosis of juvenile pleomorphic adenoma must distinguished it from other congenital salivary gland tumours (e.g. congenital basal cell adenoma, hybrid basal cell adenoma-adenoid cystic carcinoma, sialoblastoma, salivary gland anlage tumour.

Topics: Adenoma, Pleomorphic; Biomarkers, Tumor; Cell Transformation, Neoplastic; Child; Diagnosis, Differential; Epithelium; Female; Humans; Keratins; Neoplasms, Germ Cell and Embryonal; Parotid Gland; Parotid Neoplasms

The BKJ gene was originally identified based on its specific transcriptional activation in jun-transformed avian fibroblasts. We now show that BKJ is a direct transcriptional target of the AP-1 transcription factor components Jun and Fos. The complete structural organization of the quail BKJ gene was determined by nucleotide sequence analysis and transcriptional mapping. The gene contains three exons with the coding region confined to the third exon. A major mRNA species of 0.8 kb and a minor one of 1.3 kb are produced by variable usage of two transcriptional initiation sites. The BKJ promoter region contains two authentic AP-1 binding sites. By transactivation of reporter gene constructs and direct binding of Jun recombinant protein, the proximal AP-1 element was shown to be essential for BKJ promoter activation. Using polyclonal antiserum directed against recombinant BKJ protein, the activation of BKJ in jun-transformed avian fibroblasts was also demonstrated at the protein level. BKJ is a novel gene related to the avian beta-keratin gene family whose members display highly specific expression patterns during embryogenesis and epidermal development. Activation of BKJ in fibroblasts by retroviral or deregulated cellular jun or fos alleles may contribute to cell transformation.

Topics: Amino Acid Sequence; Animals; Base Sequence; Cell Transformation, Neoplastic; Chick Embryo; Cloning, Molecular; Coturnix; Fibroblasts; Gene Expression Regulation, Neoplastic; Genes, jun; Keratins; Molecular Sequence Data; Oncogene Protein p65(gag-jun); Promoter Regions, Genetic; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Proto-Oncogene Proteins c-myc; RNA, Messenger; Transcription Factor AP-1; Transcriptional Activation; Transfection

Squamous carcinoma of the larynx arises from pre-existing lesions, the so-called "preneoplastic lesions". Hyperplastic lesions represent a part of their spectrum, from both clinical and biological points of view. On morphologic grounds, the most characteristic feature with prognostic value in the evaluation of preneoplastic lesions is dysplasia. It is not only nuclear alterations that are seen in the process of malignant transformation, the cytoplasmic pattern of cytokeratins changes through neoplastic progression, with a progressive reduction of the molecular weight of the produced species. Dysplasia also associates with gross alterations of the DNA content. This is in agreement with our finding of alterations of genes participating in the control of the cell cycle, p53 and p21(WAF1/cip1). p53 overexpression is detected in non-invasive squamous lesions (even in the absence of obvious dysplasia) and p21(WAF1/cip1) shows a dramatic change in the pattern of expression in dysplastic epithelium compared with the normal. However, not all genes participating in the control of the cell cycle are altered in early lesions. Overexpression of cyclin D1, a common phenomenon in advanced carcinomas, is not likely to participate in the early phases of neoplastic development.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cyclin D1; Cyclin-Dependent Kinases; Cyclins; DNA, Neoplasm; Epithelium; Genes, p53; Humans; Hyperplasia; Keratins; Laryngeal Diseases; Laryngeal Mucosa; Laryngeal Neoplasms; Oncogene Proteins; Precancerous Conditions

The cytokeratin 18 related molecules of human hepatocellular carcinoma have been previously recognized through a series of biochemical and immunological approaches. It is suggested that these molecules undergo modulation from human hepatocyte cytokeratin 18. To prove whether these molecules are produced by modulation or protein degradation, we checked the cytokeratin profile of human hepatoma cell line PLC/PRF/5 with the methods used before. These results revealed that the PLC cells have the same cytokeratin 18 related molecules as human hepatocellular carcinoma tissue. The gene expression of the cytokeratin 18 in non-tumor liver tissues, hepatocellular carcinoma and PLC/PRF/5 cells were investigated. First, the mRNAs of non-tumor liver tissues, hepatocellular carcinoma tissues and PLC/PRF/5 cells were collected by the acid guanidinium thiocyanate phenol chloroform method. After transcription into cDNA by reverse transcriptase polymerase chain reaction, the cDNAs of each specimen were amplified by PCR and then digested by SmaI and BamHI restriction enzymes. The digested cDNA fragments were electrophoresed in agarose gel and the base pairs were found to be the same in length between neoplastic and non-neoplastic hepatocytes.

Topics: Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Humans; Keratins; Liver Neoplasms; Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured

The differential expression of cytokeratins in epithelial or squamous cells has been demonstrated to be altered during the process of carcinogenesis. This altered expression of cytokeratins (CKs) may be closely related with epithelial differentiation and may remain stable in malignant tumors. In the present study an analysis using two monoclonal antibodies, CK 8.12 antibody specific for CK type 13 and 16 and CK 8.60 antibody specific for CK type 1, 10 and 11 was done in different grades of lesions in the uterine cervix. Changes from the normal expression pattern were seen in high grade Squamous intraepithelial lesions (SIL) (CIN-2/3) and invasive squamous cell carcinoma (SCC). No conspicuous difference in the staining expression between normal/benign cervical tissue and low grade SIL (CIN-I) was evident. Statistical analysis also revealed a significant correlation between the expression of these CK types to the differentiation status of the cervical lesions analyzed. Alterations in the expression of these CKs can be correlated to the differentiation pathway which may be deregulated during cervical carcinogenesis. The findings of the present study suggest that the expression of CK types 13 and 16 and 1, 10 and 11 using CK 8.12 and CK 8.60 antibodies respectively may serve as markers of differentiation in cervical squamous neoplasms.

Topics: Adult; Analysis of Variance; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Cervix Uteri; Female; Humans; Immunochemistry; Keratins; Middle Aged; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Growth factor-independent proliferation and loss of response to differentiation factors are believed to be critical elements in carcinogenesis. We have developed an in vitro model of human prostatic carcinogenesis by the introduction of SV40 DNA into normal prostatic epithelial cells to create a transformed, immortal cell line, pRNS-1-1. This non-tumorigenic cell line responded similarly to normal prostatic epithelial cells to most growth- and differentiation-regulatory factors, with the notable exception of loss of response to the inhibitory factor 1,25-dihydroxyvitamin D3. In this study, we describe the introduction of the ras oncogene into pRNS-1-1 cells to create a tumorigenic cell line, pRNS-1-1/ras. In addition to an attenuated response to 1,25-dihydroxyvitamin D3, these cells also became unresponsive to retinoic acid and gained the ability to undergo clonal proliferation in the absence of epidermal growth factor (EGF). EGF-independent growth could not be linked to the production of autocrine transforming growth factor-alpha, but instead was likely due to sustained signaling by the ras oncogene, bypassing ligand-activation of the EGF receptor. Ligand-independent proliferation, coupled with the loss of response to the growth-inhibitory and differentiation agent retinoic acid, may be important elements in the conversion of human prostatic epithelial cells to tumorigenicity.

Topics: Animals; Calcitriol; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Growth Factor; Epithelium; Fibroblast Growth Factor 1; Genes, ras; Humans; Keratins; Male; Mice; Mice, Nude; Neoplasm Transplantation; Prostate; Somatomedins; Transforming Growth Factor alpha; Tretinoin; Tumor Necrosis Factor-alpha

Carcinosarcoma is a rare neoplasm that displays morphological features of both an adenocarcinoma and a sarcoma. The question is whether two tumors co-exist or whether the two morphological aspects represent sequential steps in tumor progression. We report a case of carcinosarcoma of the caecum in a young female. To characterize the two tumor cell populations and to gain insight into the pathogenesis of the lesion, we conducted immunohistochemical and ultrastructural analyses of the tumor. The biphasic aspect of the tumor showed an admixture of carcinoma and spindle-cell sarcomatoid areas. Both adenocarcinoma and sarcomatous cells were positive for cytokeratins. Vimentin was undetectable in the epithelial portion, but many of the sarcomatous cells stained for vimentin. Electron microscopic analyses of the sarcomatous portion revealed budding of "retroviral particles" from the rough endoplasmic reticulum cisternae. Our data support the contention that "carcinosarcoma" is a part of a single clinicopathological continuum with "spindle-cell carcinoma", the former being the biphasic expression of the neoplasia, the latter the monophasic expression; the presence of productive retroviral infection in the sarcomatous cells could constitute one of the additional support in tumor progression from the carcinomatous to the sarcomatous phase.

Topics: Adenocarcinoma; Adult; Biomarkers, Tumor; Carcinosarcoma; Cell Transformation, Neoplastic; Colonic Neoplasms; Disease Progression; Endoplasmic Reticulum, Rough; Fatal Outcome; Female; Humans; Keratins; Neoplasm Proteins; Organelles; Retroviridae; Sarcoma; Vimentin

The immunohistochemical expression patterns of cytokeratin polypeptides and vimentin were investigated in normal epithelia and squamous cell carcinomas of the larynx with special emphasis on tumor grading. During malignant transformation of epithelial cells, the cytokeratin expression patterns changed, depending on the differentiation grade of the carcinomas. In low-grade carcinomas, the expression patterns were close to those of the normal epithelium. With increasing tumor grade, there was decreased expression of stratification cytokeratins and increased expression of basal cell, simple cell and hyperproliferation-related cytokeratins. Increasing tumor grade was also associated with the expression of vimentin, a cytoskeletal protein of mesenchymal cells. No relationship was found between vimentin expression and the presence of lymph-node metastases.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Division; Cell Transformation, Neoplastic; Epithelium; Humans; Keratins; Laryngeal Neoplasms; Larynx; Lymphatic Metastasis; Neoplasm Staging; Prognosis; Reference Values; Vimentin

Our experiments were designed to identify initial biochemical and biological changes that occur during pancreatic carcinogenesis. TAKA-1, an immortal hamster pancreatic ductal cell line, was treated in vitro for up to 11 weeks with the pancreatic carcinogen N-nitorosobis(2-oxopropyl)amine (BOP). These treated cells were designated TAKA-1 + BOP. The growth of TAKA-1 and TAKA-1 + BOP cell lines was investigated in soft agar and in hamsters intradermally. The resulting tumor from TAKA-1 + BOP was re-cultured in vitro and designated TAKA-1 + BOP-T. Mutation of c-K-ras and p53 oncogenes, chromosomal changes, expression of transforming growth factor alpha (TGF-alpha) and epidermal growth factor (EGF) receptor and several biochemical markers were examined in all cell lines. TAKA-1 + BOP but not TAKA-1 cells grew in soft agar and produced an invasive tumor in vivo. However, there were no differences in cell growth rate, DNA flow cytometry, or immunohistochemical findings between the non-transformed and transformed cells. TAKA-1, TAKA-1 + BOP and TAKA-1 + BOP-T cells all expressed mRNA of TGF-alpha and EGF receptor in a comparable pattern. DNA sequence analysis following polymerase chain reaction showed that neither TAKA-1 nor TAKA-1 + BOP cells has a mutation of c-K-ras or p53. Karyotype analysis demonstrated that TAKA-1 + BOP cells had more chromosomal abnormalities compared with TAKA-1 cells. Mutation of c-K-ras and p53 was not essential for carcinogenesis in hamster pancreatic ductal cells in vitro. In conclusion, immortality of the TAKA-1 cells caused expression of TGF-alpha to the same extent as in malignant cells. Chromosomal and ultrastructural patterns were the only differences detected between the non-transformed and BOP-transformed cells.

Topics: Amino Acid Sequence; Animals; Base Sequence; Carcinogens; Cell Division; Cell Line; Cell Transformation, Neoplastic; Cricetinae; Epidermal Growth Factor; ErbB Receptors; Genes, p53; Genes, ras; Karyotyping; Keratins; Kinetics; Mice; Microscopy, Electron; Molecular Sequence Data; Mutagenesis; Nitrosamines; Pancreatic Ducts; Pancreatic Neoplasms; Rats; Sequence Alignment; Transforming Growth Factor alpha

Progressive loss of the differentiated phenotype and communication with stroma accompanies the transition of nonmalignant rat prostate epithelial cells to anaplastic, malignant tumors. Here we show that cell surface expression of the fibroblast growth factor receptor 2 (FGFR2) tyrosine kinase is reduced in malignant tumor cell populations (type II) and undetectable at the mRNA level in 30% of cells. This is in addition to the irreversible loss by splice switching of the FGFR2 ectodomain that abrogates response to FGF-7 and homologues from the stroma. One hundred % of type II malignant cells express FGFR1, which is normally expressed in the stroma. Expression of the FGFR1 kinase in premalignant type I tumor epithelial cells by transfection accelerated progression to the malignant phenotype. In contrast to the FGFR2 kinase fused to the ectodomain of FGFR1, the FGFR1 kinase failed initially to support a mitogenic response to FGF-2 in type I tumor cells. However, the FGFR1-transfected cells acquired a mitogenic response after extensive proliferation of the cell population. Resident FGFR2 and ectopic FGFR1 appeared to be partitioned in the type I cells, because neither full-length nor truncated isoforms of FGFR1 affected the mitogenic response of the other. Restoration of the FGFR2IIIb kinase to malignant cells expressing FGFR1 depressed tumor growth rates, restored responsiveness to stromal cells, and restored epithelial cell differentiation. These observations reveal that homologous FGFR1 and FGFR2 kinases play very different roles in cell growth and differentiation and in development and support of the malignant phenotype.

Topics: Animals; Cell Differentiation; Cell Division; Cell Line; Cell Transformation, Neoplastic; Disease Progression; DNA Primers; Epithelial Cells; Fibroblast Growth Factor 2; Keratins; Male; Polymerase Chain Reaction; Precancerous Conditions; Prostate; Prostatic Neoplasms; Rats; Receptor Protein-Tyrosine Kinases; Receptor, Fibroblast Growth Factor, Type 1; Receptor, Fibroblast Growth Factor, Type 2; Receptors, Fibroblast Growth Factor; Recombinant Fusion Proteins; RNA Splicing; Transfection; Tumor Cells, Cultured

Matrix metalloproteinases (MMPs) regulate ductal morphogenesis, apoptosis, and neoplastic progression in mammary epithelial cells. To elucidate the direct effects of MMPs on mammary epithelium, we generated functionally normal cells expressing an inducible autoactivating stromelysin-1 (SL-1) transgene. Induction of SL-1 expression resulted in cleavage of E-cadherin, and triggered progressive phenotypic conversion characterized by disappearance of E-cadherin and catenins from cell-cell contacts, downregulation of cytokeratins, upregulation of vimentin, induction of keratinocyte growth factor expression and activation, and upregulation of endogenous MMPs. Cells expressing SL-1 were unable to undergo lactogenic differentiation and became invasive. Once initiated, this phenotypic conversion was essentially stable, and progressed even in the absence of continued SL-1 expression. These observations demonstrate that inappropriate expression of SL-1 initiates a cascade of events that may represent a coordinated program leading to loss of the differentiated epithelial phenotype and gain of some characteristics of tumor cells. Our data provide novel insights into how MMPs function in development and neoplastic conversion.

Topics: Animals; beta Catenin; Cadherins; Cell Differentiation; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Epithelial Cells; Extracellular Matrix; Fibroblast Growth Factor 10; Fibroblast Growth Factor 7; Fibroblast Growth Factors; Growth Substances; Keratins; Mammary Glands, Animal; Matrix Metalloproteinase 3; Mesoderm; Mice; Phenotype; Rats; Trans-Activators; Tumor Cells, Cultured; Vimentin

The carcinogen 7,12-dimethylbenz(a)anthracene (DMBA) has been used to induce oral carcinogenesis of the hamster buccal mucosa in an experimental model that exhibits many of the genetic, biochemical and pathological features of human oral squamous cell carcinoma. To complement this in vivo process we have established an in vitro transformation procedure that involved the treatment of normal hamster oral mucosal keratinocytes (NHKs) with DMBA. Uptake of DMBA in NHKs was verified by observing autofluorescence of DMBA in the oral mucosal cells. Treatment doses ranged from 5, 50 and 200 ng and the NHKs were generally treated with DMBA for 1-14 days. The 200 ng dose proved to be toxic to these cells. The 5 and 50 ng treatments were found to maintain the viability of the NHKs and demonstrate anchorage-independent agarose growth, producing 18 and 40 colonies, respectively, after 14 days of treatment. Characterisation assays included determinations for cellular growth through plating efficiency, counting of cell colony number, and 3H-thymidine incorporation. Differentiation was ascertained by counting cornified cells, specification of either high or low molecular weight keratins, the percentage of cells expressing gamma glutamyl-transpeptidase (GGT), the level of p53 expression, and a determination of cell cycle. After 24 h the plating efficiency of the NHKs was found to be slightly increased following treatment with a 5 or 50 ng dose of DMBA compared to the untreated NHKs. After 14 days of incubation these doses also enhanced the number of colonies formed by the NHKs (e.g. plating efficiency). In contrast, the number of cornified cells was reduced in these colonies, while immunohistochemistry disclosed an increase in the number of NHKs expressing low molecular weight keratins, significantly lower levels of high molecular weight keratins and high levels for GGT. Flow cytometric analysis verified an increase in p53 expression (e.g. p53 wild type, 19% and p53 mutant, 66%). Cell cycle analysis of NHKs treated with DMBA (5 ng) demonstrated a shift in the number of cells in S phase (17.2%) and G2 + mitosis (11.0%). Cells from this DMBA treatment group were injected into syngeneic hamster recipient buccal pouches (10 x 10(6) cells/0.25 ml). Squamous carcinomas grew in four of six hamster buccal pouches as determined by histopathological analysis. The in vitro assay system will enhance our ability to define the genetic and molecular changes related to chemical carcinogen

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Culture Techniques; Cell Cycle; Cell Transformation, Neoplastic; Cheek; Cricetinae; Dose-Response Relationship, Drug; Keratinocytes; Keratins; Mesocricetus; Mouth Mucosa; Mouth Neoplasms; Neoplasm Transplantation

We describe the case of a 73-year-old patient with gastric adenocarcinoma composed of histologically well differentiated glandular areas, extensive rhabdoid zones and regions depicting a transition between these two constituents. The rhabdoid component showed typical features such as abundant eosinophilic cytoplasm, eccentric nuclei, prominent nucleoli, intense positive immunohistochemical cytoplasmic reaction for vimentin and less evident immunohistochemical for cytokeratin and epithelial membrane antigen (EMA). Our findings strongly suggest that the rhabdoid areas probably represent a phenotypic variant of a gastric adenocarcinoma, otherwise fairly well differentiated, a combination that to the best of our knowledge has not been previously reported.

Topics: Actins; Adenocarcinoma; Aged; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; Desmin; Female; Humans; Immunohistochemistry; Keratins; Mucin-1; Rhabdoid Tumor; Stomach Neoplasms; Vimentin

Cloned v-raf, v-raf/v-myc, and spontaneously transformed rat liver epithelial (RLE) cell lines were examined for meastatic capability in nude mice, using the LacZ gene as a marker for quantitation of micrometastases. Six cloned lines (R3611-T lines) derived from nude mouse xenografts of the v-raf transformed R3611-3 cells displayed variable metastatic capabilities. Three of six subcutaneously inoculated R3611-TlacZ lines produced spontaneous lung metastasis in nude mice. One of the lines, R3611-T2lacZ was highly efficient at metastatic conversion and produced more lung colonies than a faster growing v-raf/v-myc-transformed RJ2-14lacZ line. The spontaneously transformed RLElacZ line (C4T) was nonmetastatic, although it produced larger subcutaneous tumors than the metastatic R3611-T2lacZ line. Metastatic conversion correlated with upregulation of urokinase-type plasminogen activator receptor RNA expression and downregulation of plasminogen activator inhibitor-1, collagen alpha1 (I), and cytokeratin 14 (K14) RNA expression. These findings indicate that proteolytic activities associated with plasminogen activation play a role in the metastatic development in this model. Decreased production of extracellular proteins and cytoskeletal changes associated with lack of K14 expression are also likely to have contributed to the metastatic conversion of the RLE transformants.

Topics: Animals; Cell Transformation, Neoplastic; Clone Cells; Collagen; Down-Regulation; Epithelial Cells; Gene Expression Regulation, Neoplastic; Genes, myc; Keratin-14; Keratins; Liver; Liver Neoplasms, Experimental; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Oncogene Proteins v-raf; Phenotype; Plasminogen Activator Inhibitor 1; Rats; Rats, Inbred F344; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Retroviridae Proteins, Oncogenic; RNA, Messenger

We report the malignant transformation of human prostate epithelial cells (267B1) after multiple exposures to ionizing radiation. Carcinogenic progression of cells from immortal growth to anchorage-independent growth in soft agar to tumorigenicity in athymic mice resulted after a cumulative X-ray dose of 30 Gy. The tumors were characterized histologically as poorly differentiated adenocarcinomas, expressed prostate-specific antigen, and stained positive for keratin. No p53 or ras mutations were observed. Numerous chromosomal defects were noted on karyotypes after radiation exposure. However, chromosome 3 and 8 translocations were observed predominantly in the tumor outgrowths. These findings provide the first evidence of malignant transformation of human prostate epithelial cells exposed to ionizing radiation.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Epithelium; Humans; Karyotyping; Keratins; Male; Mice; Mice, Nude; Neoplasm Transplantation; Prostate; Prostate-Specific Antigen

Human papillomaviruses (HPVs) and cigarette smoking are epidemiologically associated with cervical cancer. We recently found that HEN-16 and HEN-16-2 HPV type 16-immortalized endocervical cells form tumors after treatment with cigarette smoke condensate and derived 2 tumor cell line cultures, HEN-16T and HEN-16-2T, respectively. Here, we examine the molecular pathologic effect of tumorigenesis. HEN-16T and HEN-16-2T exhibit unchanged status and expression of integrated HPV 16 DNA. However, the expression of the cytokeratin CK7 and CK13 endocervical cell markers is more homogeneous in monolayer and organotypic raft cultures after tumorigenesis. For the effect of retinoic acid on monolayers for growth inhibition, HEN-16T were significantly less sensitive than the normal and immortalized non-tumorigenic cells. HEN-16-2T were completely resistant. Moreover, the rafts from both tumorigenic cell line cultures were resistant to retinoic acid and continued to display thick rafts and homogeneous severe dysplasia/carcinoma in situ. In contrast, the non-malignant HEN-16 and HEN-16-2 rafts were thinner, and treatment with retinoic acid blocked the formation of severe dysplasia, reconstructing an epithelium resembling that of the normal endocervix. Our results support the significance of non-viral factors in the mechanism by which cigarette smoking induces tumorigenesis in the late stages of HPV-initiated progression to cervical cancer. Importantly, our data indicate that the sensitivity to retinoic acid of the HPV-containing endocervical cells is lost following tumorigenesis in vitro and possibly in women.

Topics: Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Cell Transformation, Viral; DNA, Viral; Drug Resistance; Female; Humans; Keratins; Keratolytic Agents; Papillomaviridae; Tretinoin; Uterine Cervical Neoplasms

High-risk human papillomaviruses (HPVs), including type 16, have been identified as factors in cervical carcinogenesis. However, the presence and expression of the virus per se appear to be insufficient for carcinogenesis. Rather, cofactors most likely are necessary in addition to viral gene expression to initiate neoplasia. One candidate cofactor is prolonged exposure to sex hormones. To examine the possible effects of estrogen on HPV-associated neoplasia, we treated transgenic mice expressing the oncogenes of HPV16 under control of the human keratin-14 promoter (K14-HPV16 transgenic mice) and nontransgenic control mice with slow release pellets of 17beta-estradiol. Squamous carcinomas developed in a multistage pathway exclusively in the vagina and cervix of K14-HPV16 transgenic mice. Estrogen-induced carcinogenesis was accompanied by an incremental increase in the incidence and distribution of proliferating cells solely within the cervical and vaginal squamous epithelium of K14-HPV16 mice. Expression of the HPV transgenes in untreated transgenic mice was detectable only during estrus; estrogen treatment resulted in transgene expression that was persistent but not further upregulated, remaining at low levels at all stages of carcinogenesis. The data demonstrate a novel mechanism of synergistic cooperation between chronic estrogen exposure and the oncogenes of HPV16 that coordinates squamous carcinogenesis in the female reproductive tract of K14-HPV16 transgenic mice.

Topics: Animals; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Chloramphenicol O-Acetyltransferase; Delayed-Action Preparations; Estradiol; Female; Genes, Viral; Humans; Immunohistochemistry; In Situ Hybridization; Keratins; Mice; Mice, Transgenic; Oncogenes; Papillomaviridae; Proliferating Cell Nuclear Antigen; Promoter Regions, Genetic; RNA, Messenger; Uterine Cervical Neoplasms; Vaginal Neoplasms

To directly examine a multistage carcinogenesis model and the role of UV light in human tissues, we grafted human skin onto mice with severe combined immunodeficiency disease. We found that the maximum dose of UV radiation in the B range (UVB; 280-320 nm) tolerated by these grafts was 500 J/m2 three times weekly. One hundred fifty-one grafted mice were then randomized and observed for a median of 9 months in five groups: no treatment, chemical initiation alone, UVB as a complete carcinogen, initiation plus UVB promotion, and initiation plus UVB and phorbol ester promotion. Actinic damage and squamous atypia were found in grafts of all groups receiving UV treatment; unequivocal human squamous carcinomas developed in two of these. Species origin was verified by human-specific bisbenzimide staining and in situ hybridization for human-specific Alu segment. Overall basal proliferation, measured immunohistologically, was reduced in UV-treated grafts, but foci of hyperproliferation were seen in conjunction with the dedifferentiated expression of cytokeratins 1, 10 and 5, 8. Murine tumors also developed frequently, confirming the biological relevance of the carcinogenic strategies tested. These findings demonstrate that development of malignant human tumors can be experimentally accelerated in human tissue.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Antigens, Neoplasm; Carcinogens; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Humans; Keratins; Ki-67 Antigen; Mice; Mice, SCID; Neoplasm Proteins; Nuclear Proteins; Papilloma; Skin Neoplasms; Skin Transplantation; Tetradecanoylphorbol Acetate; Transplantation, Heterologous; Ultraviolet Rays

Investigations were carried out designed to isolate and biologically characterize the subpopulation of cells within the Syrian hamster embryo (SHE) cell population which are sensitive to morphological transformation (MT). Biological cloning of MT-sensitive cells demonstrated that within the complex SHE cell population, MT-sensitive cells comprise approximately 27% of the clonally plateable cellular population. Importantly, MT-sensitive cells display MT rates of approximately 7% following benzo[a]pyrene exposure, a rate which falls to <1% with additional passage of the cells, indicating that the ability to undergo MT is a transient phenomenon. Biological characterization of the clonal MT-sensitive cells indicates that these cells are relatively undifferentiated, since they express both cytokeratins and vimentin and respond to a variety of stem cell growth and differentiation factors, although the majority appear to be committed progenitor cells, since they express either mesenchymal or epithelial cell characteristics. Together these data demonstrate that MT-sensitive cells comprise a subpopulation of the cells within the complex SHE cell population, that the ability to undergo MT is a transient phenomenon and that MT-sensitive cells are relatively undifferentiated committed progenitor stem-like cells, all of which gives rise to the hypothesis that MT is an alteration in the cellular differentiation state.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Cricetinae; Embryo, Mammalian; Female; Growth Substances; Keratins; Mesocricetus; Pregnancy; Vimentin

Cytokeratins are subunit proteins of epithelial cell intermediate filaments, which are genetically determined. Because epithelia have their own characteristic cytokeratin profile, this may reveal the origin of the epithelium. The cytokeratin profile of Barrett's oesophagus, complicating severe gastro-oesophageal reflux disease, was determined in 35 consecutive patients and in 10 normal controls in order to provide insight into the origin of Barrett's epithelium. Immunostaining of frozen sections showed abundant immunoactivity for cytokeratin (CK) 13, which is characteristic of squamous epithelia, including that of the oesophagus, but is not present in the simple columnar epithelium of the cardia. On the other hand, the latter epithelium expresses mainly CK 8, 18 and 19, also found in Barrett's epithelium. The presence of CK 13 in Barrett's epithelium may indicate its origin from the squamous oesophageal epithelium and not from the proximal migration of columnar epithelial cells of the gastric cardia.

Topics: Adult; Aged; Aged, 80 and over; Barrett Esophagus; Cell Transformation, Neoplastic; Culture Techniques; Endothelium; Female; Fluorescent Antibody Technique, Indirect; Humans; Keratins; Male; Middle Aged

Previously we described (Dong et al., 1990) a nuclear protein (mol. wt. 112 kD) which is expressed abundantly in hepatoma cells and also in hepatocyte cells committed to carcinogenesis. In this report, we further characterize its chemical properties and cellular localization in normal and hepatoma cells. 112 kD hepatoma-associate nonhistone protein is not a cytokeratin-related protein as described by Fukuda et al. (1991). Protein purification experiments revealed that 112 kD protein is a dimer of 56 kD polypeptide present in normal rat liver nuclei. Intranuclear distribution pattern indicated that 112 kD nonhistone protein localizes exclusively in hepatoma nuclear matrix. The data from this study suggest that dimerization of 56 kD nonhistone protein is involved in nuclear matrix reorganization during neoplastic transformation.

Topics: Animals; Cell Transformation, Neoplastic; Female; Histones; Immunoblotting; Keratins; Liver; Liver Neoplasms, Experimental; Molecular Weight; Neoplasm Proteins; Nuclear Matrix; Nuclear Proteins; Rats; Rats, Inbred BUF

By using the subtractive hybridization method, two complementary DNA clones differently expressed in rat normal esophageal epithelium and squamous cell carcinoma induced by administration of precursors of N-nitrososarcosine ethyl ester were isolated. A rat homologue of the human 50-kDa type I cytokeratin 14 was cloned for the first time and shown to be expressed preferentially in squamous cell papillomas and carcinomas, whereas it was weakly expressed or absent in normal squamous epithelial cells and in hyperplastic lesions. A rat homologue of the mouse 57-kDa type II cytokeratin showed strong expression in both normal and tumor tissues. These results are well consistent with the reported alteration of keratin subspecies in human esophageal cancers, therefore, encouraging us to use this experimental system as a model for human esophageal carcinogenesis.

Topics: Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; DNA, Complementary; Esophageal Diseases; Esophageal Neoplasms; Esophagus; Humans; Hyperplasia; In Situ Hybridization; Keratins; Mice; Neoplasm Proteins; Nitrosamines; Papilloma; Precancerous Conditions; Rats; Rats, Wistar; Recombinant Proteins; Subtraction Technique

Reproducible multi-stage progression to invasive squamous carcinoma of the epidermis has been achieved in transgenic mice expressing the HPV16 early-region genes, including the E6/E7 oncogenes, under the control of the human keratin-14 promoter/enhancer. Although 100% of K14-HPV16 transgenic animals develop hyperplastic and/or dysplastic lesions in several inbred backgrounds, including C57BL/6, BALB/c, and SSIN/SENCAR, only mice backcrossed into the FVB/n background progress to malignant squamous cell carcinomas of two pathological grades, well differentiated and moderate/poorly differentiated (WDSC or MPDSC, respectively), each displaying characteristic patterns of malignant behavior. WDSCs typically arise within the epidermis of the ear and invade deeply into the underlying dermis but fail to metastasize, whereas MPDSCs develop on the chest and truncal skin and invariably metastasize to regional lymph nodes. The transition to the malignant state, in 21% of FVB/n transgenic mice, is characterized by alteration of the repertoire of keratin intermediate filament proteins expressed within neoplastic epidermis, such that WDSCs maintain expression of keratins common to terminally differentiating stratified keratinocytes (K10), whereas MPDSCs are distinguished from WDSCs by activation of embryonic and mucosal keratins (K13, K8, and K19). Precursor hyperplastic and dysplastic lesions are characterized by a progressively increased proliferative index, striking morphological alterations in keratinocyte cell-cell and cell-matrix interactions, and extensive remodeling of the underlying dermal stroma. Remarkably, this extensive stromal remodeling, which may facilitate both angiogenesis and eventual tumor cell invasion, develops early at the dysplastic stage in all animals well before malignant conversion.

Topics: Animals; Carcinoma, Squamous Cell; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Disease Susceptibility; DNA-Binding Proteins; Keratins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred SENCAR; Mice, Transgenic; Neoplasm Invasiveness; Oncogene Proteins, Viral; Oncogenes; Papillomaviridae; Papillomavirus E7 Proteins; Transcription Factors

HC11 cells are a model for mammary epithelial cell differentiation. Following treatment with the lactogenic hormones glucocorticoids, insulin and prolactin the HC11 cells synthesize milk proteins. Stereological analysis at the ultrastructural level suggested that lysosomal biogenesis was activated following lactogenic hormone treatment of HC11 cells. Differentiation was also accompanied by an increase in the cellular content of tri- and tetra-antennary oligosaccharides, which were reactive with isolectin L4 from Phaseolus vulgaris (L-PHA). The lysosomal-associated membrane glycoproteins LAMP-1 and LAMP-2 are the major carriers of this glycosylation pattern. An analysis of LAMP-1 and LAMP-2 expression levels showed that there was a dramatic increase in LAMP-1 following lactogenic hormone treatment of HC11 cells. The control of LAMP-1 expression is mainly post-transcriptional since the level of LAMP-1 RNA is not affected by lactogenic hormones. Stereological analysis also showed an increase in intermediate filament control of differentiated cells. Analysis of the cytokeratins expressed in differentiated cells suggests that HC11 cells have characteristics of a mammary-specific stem cell. Increase in lysosomal vesicles and their contents might play a role in intra- and extra-cellular remodeling, which is characteristic of cell differentiation.

Topics: Animals; Antigens, CD; Biomarkers; Cell Differentiation; Cell Nucleus; Cell Transformation, Neoplastic; Cells, Cultured; Cytoplasm; Epithelial Cells; Genes, ras; Glycoproteins; Glycosylation; Keratins; Lysosomal Membrane Proteins; Lysosomes; Mammary Glands, Animal; Membrane Glycoproteins; Mice; Phytohemagglutinins

Tumor of the follicular infundibulum is a rare proliferation of cells and its histogenesis or differentiation at the morphological level has been the subject of some controversy. In recent years cytokeratins have been recognized as important markers of epithelial differentiation, and of late new retrieval methods have meant it is possible to detect them in formalin-fixed and paraffin-embedded tissue. Four patients were studied, the ages ranging between the 2nd and 7th decade. The tumors were all located in the head and neck and in one case the lesion developed in a pre-existing sebaceous nevus. The morphological investigation revealed a flat, proliferation of polygonal, pale eosinophilic cells connected to epidermis or follicular infundibulum and with a centrally located nucleus. In addition, a few ductal structures resembling sebaceous ducts were seen and in one case a hair germ papilla and a follicular papilla was noted. Immunohistochemical investigations with antibodies against cytokeratins revealed differentiation comparable to that in the fetal follicular isthmus and, in one case, also differentiation in keeping with the fetal follicular infundibulum.

Topics: Adult; Aged; Biomarkers, Tumor; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Cytoplasm; Female; Hair Follicle; Hamartoma; Head and Neck Neoplasms; Humans; Keratinocytes; Keratins; Male; Middle Aged; Precancerous Conditions; Sebaceous Glands; Skin Neoplasms

A hormone-dependent, clonal carcinoma cell line, designated RM22-F5, was derived from a malignant mammary mixed tumor spontaneously arising in an outbred old female Wistar rat. These cells expressed keratin and desmosomal protein and formed epithelial monolayers in a growth factor and hormone-supplemented medium (LHC-8) containing 10% fetal bovine serum (E-type cells). Cells subcultured for 6 to 8 passages in RPMI 1640 medium containing 10% fetal bovine serum without supplements appeared to be fibroblastic and expressed vimentin (F-type cells). The shift to a fibroblast-like morphology was associated with a more malignant phenotype which included rapid, hormone-independent growth and invasive sarcoma-like character in nude mice. F-type cells were no longer able to express their original epithelial phenotype in LHC-8 medium. Cytogenetic analysis revealed that both E- and F-type cells had essentially the same karyotype. Analysis of PCR-amplified DNA further demonstrated a point mutation of the H-ras-1 gene at codon 12 and loss of the normal H-ras-1 allele in both cell types. Genetic tagging of E-type cells with the neomycin-resistance gene resulted in the generation of F-type cells with neomycin resistance in RPMI 1640 medium, suggesting that F-type cells are a malignant variant of E-type cells arising during in vitro culture. Somatic cell fusion between E- and F-type cells revealed that with most hybrid clones tested, the fibroblast-like phenotype was greatly suppressed. These results suggest that an irreversible phenotypic transition, representative of tumor progression from hormone-dependent adenocarcinoma to more malignant hormone-independent spindle carcinoma cells, is a recessive event and may involve loss of a suppressor function.

Topics: Adenocarcinoma; Animals; Base Sequence; Carcinoma; Cell Division; Cell Fusion; Cell Transformation, Neoplastic; Culture Media; Desmosomes; Female; Flow Cytometry; Genes, ras; Keratins; Mammary Neoplasms, Animal; Mice; Mice, Nude; Molecular Sequence Data; Neomycin; Neoplasm Invasiveness; Neoplasms, Hormone-Dependent; Phenotype; Rats; Rats, Wistar; Transfection; Tumor Cells, Cultured; Vimentin

To assess the synergistic effect of growth and transcription factor deregulation on carcinogenesis in vivo, mating experiments were performed between transgenic mice expressing human TGF alpha or v-fos exclusively in the epidermis by means of a human keratin K1-based targeting vector (HK1.fos, HK1.TGF alpha and HK1.fos/alpha). While HK1.TGF alpha mice exhibited mild epidermal hyperplasia resulting in a wrinkled appearance, this hyperplasia was significantly increased in HK1.fos/alpha mice which also exhibited a novel opalescent and peeling skin phenotype. HK1.fos/alpha keratinocyte differentiation was considerably deregulated with cornified cells appearing in the granular layer, granular cells in the spinous layer and a sixfold increase in BrdU labeling over normal. In addition, hyperplastic HK1.fos/alpha epidermis exhibited aberrant loricrin, filaggrin and novel K13 expression associated with v-fos expression. Unlike adult HK1.TGF alpha controls, hyperplasia persisted in HK1.fos/alpha adults which also rapidly developed autonomous squamous cell papillomas. These results demonstrate that v-fos and TGF alpha over-expression can cooperate to reprogram keratinocyte differentiation and elicit the early stages of neoplasia. Moreover, TGF alpha over-expression appeared to play an early, initiating role in HK1.fos/alpha papilloma etiology, and a promotion role in the accelerated appearance of v-fos wound-associated preneoplastic phenotypes. However, the stable persistence of HK1.fos/alpha papillomas for up to 12 months, suggests that additional events are required for malignant conversion.

Topics: Animals; Base Sequence; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Filaggrin Proteins; Genetic Vectors; Keratins; Mice; Mice, Transgenic; Microscopy, Electron; Molecular Sequence Data; Oncogene Proteins v-fos; Papilloma; Skin; Skin Neoplasms; Transforming Growth Factor alpha

An electron microscopic, immunocytochemical, and enzyme cytochemical analysis of the previously established oval cell lines OC/CDE 6 and OC/CDE 22 was performed to characterize the phenotype and differentiation patterns of long-term cultures of oval cells. It was found that alpha-fetoprotein, albumin, and cytokeratin 19 are present in all cultured cells. This indicates that oval cells constitute a population of immature cells expressing features of the antigenic phenotype of both the hepatocyte and bile ductular cell lineages. An electron microscopic examination revealed a gradual alteration in the ultrastructure of oval cells toward hepatocyte-like cells. The majority of the oval cells were positive for glucose-6-phosphatase activity. A particularly striking observation was that oval cells were heterogeneous in terms of peroxisome content. Only about 50% of the oval cells had peroxisomes in the cytoplasm, these cells probably being part of the hepatocyte lineage. The other cultured cells did not reveal catalase activity and probably represented cells committed to the bile ductular cell lineage. An addition of clofibrate to the culture medium resulted in a marked peroxisome proliferation in all oval cells, indicating that oval cells might be able to change their differentiation pathway depending on environmental influence toward the hepatocyte lineage. It is most intriguing that in oval cells with abundant cytoplasm peroxisome proliferation was accompanied by proliferation of the smooth endoplasmic reticulum (this is a morphological marker of mature hepatocytes). Taken together, our findings suggest that within the oval cell lines OC/CDE 6 and OC/CDE 22 cells undergoing a morphological and functional differentiation along the hepatocyte and bile ductular cell lineages are present.

Topics: 3,3'-Diaminobenzidine; Albumins; alpha-Fetoproteins; Animals; Antigens, Neoplasm; Cell Differentiation; Cell Transformation, Neoplastic; Choline Deficiency; Clofibrate; Ethionine; Glucose-6-Phosphatase; Immunohistochemistry; Keratins; Liver; Liver Neoplasms, Experimental; Methylnitronitrosoguanidine; Microscopy, Electron; Phenotype; Rats; Rats, Sprague-Dawley; Staining and Labeling; Vimentin

Topics: Aged; Aged, 80 and over; Breast Neoplasms; Carcinosarcoma; Cell Transformation, Neoplastic; Female; Humans; Keratins; Papilloma, Intraductal; S100 Proteins

To determine the effect of retinoic acid on the development of severe dysplasia or carcinoma in situ from endocervical cells containing human papillomavirus (HPV) type 16.. Two independent lines of HPV 16-immortalized endocervical cells were reconstructed into two squamous epithelial tissues using the organotypic raft culture system to examine the differentiated phenotype. The effect of retinoic acid on dysplastic morphology of differentiation of the epithelia was examined by light microscopy of stained sections and electron microscopy. The endocervical cell type cytokeratin expression pattern was determined by indirect immunofluorescence using specific monoclonal antibodies. Ribonucleic acid expression of the HPV 16 E7 oncogene was examined by in situ hybridization.. Untreated HPV 16-immortalized endocervical cells were reconstructed into squamous dysplastic lesions resembling carcinoma in situ observed in women. Retinoic acid-treated rafts formed epithelia composed of two to three cell layers of columnar-like cells resembling simple epithelium of the endocervix. Electron microscopy and cytokeratin expression patterns confirmed the histology of a differentiated endocervical phenotype after treatment with retinoic acid. Expression of HPV 16 E7 was modestly lower in treated epithelia, preferentially in basal cells.. Retinoic acid prevents the histology and cytokeratin differentiation markers of carcinoma in situ of HPV 16-immortalized endocervical cells. Because the epithelia closely mimic HPV 16-containing severe dysplasias and native endocervical epithelium in women, this immortalized endocervical cell-raft system may be useful as a model to assess the efficacy of agents such as retinoic acid for preventing progression of these lesions to malignant cervical carcinoma.

Topics: Carcinoma in Situ; Cell Line, Transformed; Cell Transformation, Neoplastic; Cells, Cultured; Cervix Uteri; Female; Humans; Keratins; Microscopy, Electron; Papillomaviridae; Tretinoin; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Multiple mammary epithelial cell (MEC) types are observed both in mammary ducts in vivo and in primary cultures in vitro; however, the oncogenic potential of different cell types remains unknown. Here, we used human papilloma virus 16 E6 and E7 oncogenes, which target p53 and Rb tumor suppressor proteins, respectively, to immortalize MECs present in early or late passages of human mammary tissue-derived cultures or in milk. One MEC subtype was exclusively immortalized by E6; such cells predominated in late-passage cultures but were rare at early passages and apparently absent in milk. Surprisingly, a second cell type, present only in early-passage tissue-derived cultures, was fully immortalized by E7 alone. A third cell type, observed in tissue-derived cultures and in milk, showed a substantial extension of life span with E7 but eventually senesced. Finally, both E6 and E7 were required to fully immortalize milk-derived MECs and a large proportion of MECs in early-passage tissue-derived cultures, suggesting the presence of another discrete subpopulation. Identification of MECs with distinct susceptibilities to p53- and Rb-targeting human papillomavirus oncogenes raises the possibility that these cells may serve as precursors for different forms of breast cancer.

Topics: Breast; Cell Cycle; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Female; G1 Phase; Gamma Rays; Humans; Keratins; Lactation; Mammaplasty; Mastectomy; Milk, Human; Oncogene Proteins, Viral; Oncogenes; Papillomaviridae; Papillomavirus E7 Proteins; Protein-Tyrosine Kinases; Repressor Proteins; Retinoblastoma Protein; RNA, Messenger; Tumor Suppressor Protein p53

Five out of 4,172 operated maxillary cysts (3 developed within the maxillary and 2 in the mandible) presented a malignant change of epithelium. In all these 5 cases histology demonstrated a transition from normal to cancerous epithelium. The overall percentage of malignant change was of 0.12%, with 0.077% for non-keratinized epithelium lining odontogenic cysts (3 cases) and 0.65%, that is 8 times higher, for keratinized ones (2 cases). Keratinization of cystic epithelium and chronic inflammatory lesions were the main risk factors.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epidermal Cyst; Epithelium; Female; Humans; Keratins; Male; Mandibular Diseases; Mandibular Neoplasms; Maxillary Diseases; Maxillary Neoplasms; Middle Aged; Odontogenic Cysts; Radicular Cyst

Recently, we discovered that stable introduction of a carboxyl-terminally truncated retinoic acid receptor gamma (tRAR gamma) into an epidermal keratinocyte line blocked the ability of these cells to differentiate, as judged by their failure to express late markers of squamous differentiation. We now demonstrate a correlation between the level of residual endogenous RAR activity of tRAR gamma-expressing keratinocyte lines and degree of terminal differentiation. Mutagenesis studies localize the effects to the A/B subdomain of the truncated receptor. Despite tRAR gamma's capacity to interfere with RAR-mediated transactivation of retinoic acid response elements (RAREs) in keratinocytes, the effects of the truncated receptor are independent of its ability to bind DNA and directly interact with endogenous RARs. tRAR alpha also inhibits RARE-mediated gene expression in keratinocytes, even though its full-length counterpart enhances RARE activity in these cells. Intriguingly, both tRAR gamma and RAR gamma suppress keratin promoter activity in epidermal cells, although for tRAR gamma, the effect is mediated through the A/B domain whereas for RAR gamma, the effects require DNA binding. Taken together, these findings suggest that the truncation allows for new and aberrant interactions with transcriptional proteins/cofactors that participate in governing RARE activity. This discovery may have relevance in tumorigenesis, where genetic lesions can result in mutant RARs or in loss of receptor expression.

Topics: Base Sequence; Cell Differentiation; Cell Transformation, Neoplastic; DNA; DNA Primers; Epidermal Cells; Humans; Keratinocytes; Keratins; Molecular Sequence Data; Mutagenesis, Insertional; Oligodeoxyribonucleotides; Polymerase Chain Reaction; Promoter Regions, Genetic; Receptors, Retinoic Acid; Recombinant Proteins; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Sequence Deletion

We have shown previously that the ras and myc oncogenes can induce poorly differentiated mouse prostate carcinomas in vivo with high frequency (greater than 90%) using inbred C57BL/6 mice in the mouse prostate reconstitution model system. To study the androgen sensitivity of these carcinomas, we have developed an in vitro model system which includes a cell line from normal urogenital sinus epithelium (CUGE) and cell lines from three ras + myc transformed mouse prostate carcinomas (RM-9, RM-1, and RM-2). CUGE cells, as well as all prostate carcinoma cell lines, were positive for cytokeratin 18 mRNA and immunoreactive to cytokeratin-specific antiserum. Two out of three of the early passage carcinoma cell lines were clonal with respect to Zipras/myc 9 retrovirus integration as determined by Southern blot analysis. Whereas significant mitogenic effects of testosterone (10 nM) were not seen in CUGE cells grown in serum-free medium, under similar conditions approx. 2-fold increases in cell number were seen in all low passage prostate carcinoma cell lines. Also, in the presence of growth inhibitory levels of suramin (50 micrograms/ml), testosterone was capable of significant growth stimulation in the carcinoma cell lines. With further propagation from low passage [20-25 population doublings (PD)] to high passage (75-100 PD), all carcinoma cell lines demonstrated increased and similar growth rate in the presence and absence of testosterone. These cell lines maintained stable androgen receptor numbers and binding kinetics during the transition from testosterone-responsive growth to reduced responsivity over multiple passages in culture (> 150 PD). Overall, our studies indicate that the capacity to bind testosterone is stably maintained through the transition of the androgen-sensitive to insensitive phenotype and raise the possibility that androgen sensitivity can persist throughout progression but is masked by the acquisition of autocrine pathways.

Topics: Androgens; Animals; Blotting, Southern; Cell Division; Cell Transformation, Neoplastic; Disease Models, Animal; DNA Probes; Drug Tolerance; Female; Genes, myc; Genes, ras; Keratins; Kinetics; Male; Metribolone; Mice; Mice, Inbred C57BL; Receptors, Androgen; RNA, Messenger; Testosterone; Tumor Cells, Cultured

The detection of cytokeratins in neoplastic tissues by immunohistochemical methods has numerous diagnostic and investigative applications, because cytokeratins are usually conserved in tumour cells during malignant transformation. Recently, however, it has been reported that progression to malignancy is associated with commencement of expression of low-molecular-weight cytokeratins. In the present study, 42 specimens from 35 cases of squamous cell carcinoma (SCC) of the skin were analysed by immunohistochemical techniques, using polyclonal anti-involucrin antibody and a panel of monoclonal antikeratin antibodies, in order to investigate the nature and differentiation of SCCs. The expression of cytokeratins and involucrin in well-differentiated SCCs was similar to that in normal epidermis. In contrast with well-differentiated SCCs, the expression of differentiation-specific cytokeratins and involucrin was diminished in the immature tumour cells in proportion to the malignancy of the SCCs. Some antibodies, however, stained all tumour cells, irrespective of the degree of malignancy. Furthermore, expression of simple epithelial and non-cornifying stratified squamous epithelial cytokeratins was observed in atypical tumour cells of poorly differentiated SCCs. It is of interest that similar expression was noted in many tumour cells in the lymph node metastases and in some tumour cells in the primary cutaneous lesions. Cytokeratin expression similar to that in normal epidermal keratinocytes was conserved in well-differentiated SCCs, but the expression of cytokeratins changed during progression to malignant transformation. The expression of simple epithelial or non-cornifying stratified squamous epithelial cytokeratins in cutaneous SCCs may be a marker for their capability of invasion and metastatic potential.

Topics: Antibody Specificity; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Keratins; Lymph Nodes; Lymphatic Metastasis; Protein Precursors; Skin Neoplasms

Thirteen cases of combined hepatocellular (HCC) and cholangiocellular carcinoma (CCC) were examined. In addition to routine pathology, immunoreactivities for carcinoembryonic antigen, alpha-fetoprotein (AFP), cytokeratin (Cam 5.2 and AE1), epithelial membrane antigen (EMA) and tumor-associated glycoprotein 72 (B72.3) were also examined. The average age of the 13 cases was 64.8 years, which lay between the average ages of pure HCC and CCC cases. They were categorized as separate type (2), collision type (6), and intermingled type (5). AE1 and EMA were the best markers to differentiate the CCC from the HCC area. B72.3 immunoreactivity was detected only in CCC (46%). There were no transitional features between HCC and CCC in two cases of the separate type and two cases of the collision type. However, focal transitional features from HCC to CCC were observed in all cases of the intermingled type and in four of six cases of the collision type. In one case of the intermingled type, many cancer cells contained both bile and mucus simultaneously, and revealed dual immunoreactivities. The conclusions are: 1) the combined type is generated from two sources; one is the intrahepatic double cancer (thoroughly separate type and a part of the collision type) and another is the stem cell origin with diverse phenotypes (intermingled type and a part of the collision tumor); and 2) AE1 was the most helpful marker to differentiate the CCC area from HCC, and other markers, e.g. AFP for HCC and EMA, CEA, and B72.3 for CCC, were also supportive but somewhat limited in the differential diagnosis.

Topics: Aged; alpha-Fetoproteins; Antigens, Neoplasm; Biomarkers, Tumor; Carcinoembryonic Antigen; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Cholangiocarcinoma; Female; Glycoproteins; Humans; Keratins; Liver; Liver Neoplasms; Male; Membrane Glycoproteins; Middle Aged; Mucin-1; Mucins; Neoplasms, Multiple Primary; Neoplastic Stem Cells

Intermediate filament and S100 expression was studied in five cases of choroid plexus papilloma. All the cases showed positivity for S100 and Nimentin. Cytokeratin and glial fibrillary protein (GFAP) intermediate filaments were present in 80 pc of the cases. None of the cases expressed epithelial membrane antigen. These results confirm the divergent differentiation of choroid plexus tumours.

Topics: Adolescent; Adult; Cell Transformation, Neoplastic; Child, Preschool; Choroid Plexus Neoplasms; Diagnosis, Differential; Female; Glial Fibrillary Acidic Protein; Glioma; Humans; Immunohistochemistry; Infant; Keratins; Male; S100 Proteins; Vimentin

As part of a study on the relationship of tumour phenotype and behaviour, we have characterized two head and neck squamous cell carcinoma cell lines, derived from human laryngeal carcinomas and designated HLaC'79 and HLaC'82. Cytogenetic analysis revealed that HLaC'79 and HLaC'82 shared 10 major chromosome rearrangements indicating that the cell lines had a common origin. In the extremely complex chromosomal patterns, abnormalities were found in chromosomes 1, 3 (surplus 3q) and 5 (i(5p) x 2). Both cell lines displayed constitutive expression of vimentin and were capable of anchorage-independent growth in agarose gels. However, in spite of their common origin specific differences were found. Cells of HLaC'79 were spindle shaped and formed tumours in athymic mice. In contrast, cells of HLaC'82 had a compact morphology, contained less vimentin, were more contact inhibited and were not tumorigenic. These results indicate that malignant transformation in HLaC'82 was partially reversed.

Topics: Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Humans; Intermediate Filament Proteins; Intermediate Filaments; Karyotyping; Keratins; Laryngeal Neoplasms; Tumor Cells, Cultured; Vimentin

Transgenic mice that expressed v-fos exclusively in the epidermis by means of a human keratin K1-based targeting vector (HK1.fos) developed preneoplastic epidermal hyperplasia and hyperkeratosis after long latency and an associated wound promotion stimulus. To assess the requirements for papilloma formation and malignant conversion and determine the sensitivity to a chemical promotion stimulus, HK1.fos mice were promoted with 12-O-tetradecanoylphorbol-13-acetate (TPA). HK1.fos mice were sensitive to TPA promotion but developed papillomas only after long latency (20-30 weeks of promotion) and in relatively few numbers per animal, suggesting the necessity of an additional genetic event prior to overt lesion formation. Consistent with this idea, at 60 weeks, on cessation of TPA promotion, these HK1.fos TPA-papillomas were found to be autonomous, TPA-independent tumors which persisted, grew larger, and converted to malignancy. Analysis of HK1.fos tumor RNA and DNA identified endogenous c-rasHa mutations at codons 12 and 61 in papillomas and carcinomas; however, no p53 tumor suppressor gene mutations were detected. These data indicate that epidermal expression of v-fos induces sensitivity to TPA promotion, but since additional genetic events, such as endogenous c-rasHa activation, appear to be required in tumorigenesis, v-fos may predominantly play a role in the mechanism of promotion to achieve papilloma autonomy and TPA independence. Furthermore, spontaneous malignant conversion in this model does not appear to involve mutations in the p53 tumor suppressor gene.

Topics: Animals; Base Sequence; Biomarkers; Carcinoma; Cell Differentiation; Cell Transformation, Neoplastic; DNA, Neoplasm; Epidermis; Gene Expression Regulation, Neoplastic; Genes, fos; Genes, p53; Genes, ras; Hyperplasia; Keratins; Keratosis; Mice; Mice, Transgenic; Molecular Sequence Data; Mutation; Oncogene Proteins v-fos; Papilloma; ras Proteins; RNA, Neoplasm; Tetradecanoylphorbol Acetate

The proto-oncogene c-fos is a major nuclear target for signal transduction pathways involved in the regulation of cell growth, differentiation, and transformation. Using the multistep skin carcinogenesis model, we have directly tested the ability of c-fos-deficient mice to develop cancer. Upon treatment with a tumor promoter, c-fos knockout mice carrying a v-H-ras transgene were able to develop benign tumors with similar kinetics and relative incidence as wild-type animals. However, c-fos-deficient papillomas quickly became very dry and hyperkeratinized, taking on an elongated, horny appearance. While wild-type papillomas eventually progressed into malignant tumors, c-fos-deficient tumors failed to undergo malignant conversion. Experiments in which v-H-ras-expressing keratinocytes were grafted onto nude mice suggest that c-fos-deficient cells have an intrinsic defect that hinders tumorigenesis. These results demonstrate that a member of the AP-1 family of transcription factors is required for the development of a malignant tumor.

Topics: Animals; Cell Differentiation; Cell Transformation, Neoplastic; Epidermal Cells; Gene Expression; Genes, fos; Genes, ras; Keratinocytes; Keratins; Mice; Mice, Nude; Mice, Transgenic; Mutation; Neoplasm Transplantation; Oncogene Protein p21(ras); Papilloma; Skin Neoplasms; Time Factors; Transcription Factor AP-1

We have analyzed differential gene expression in normal versus jun-transformed avian fibroblasts by using subtracted nucleic acid probes and differential nucleic acid hybridization techniques for the isolation of cDNA clones. One clone corresponded to a gene that was strongly expressed in a previously established quail (Coturnix japonica) embryo fibroblast line (VCD) transformed by a chimeric jun oncogene but whose expression was undetectable in normal quail embryo fibroblasts. Furthermore, the gene was expressed in quail or chicken fibroblast cultures that were freshly transformed by retroviral constructs carrying various viral or cellular jun alleles and in chicken fibroblasts transformed by the avian retrovirus ASV17 carrying the original viral v-jun allele. However, its expression was undetectable in a variety of established avian cell lines or freshly prepared avian fibroblast cultures transformed by other oncogenes or a chemical carcinogen. The nucleotide and deduced amino acid sequences of the cDNA clone were not identical to any sequence entries in the data bases but revealed significant similarities to avian beta-keratin genes; the highest degree of amino acid sequence identity was 63%. The gene, which we termed bkj, may represent a direct or indirect target for jun function.

Topics: Amino Acid Sequence; Animals; Base Sequence; Cell Line, Transformed; Cell Transformation, Neoplastic; Chickens; Cloning, Molecular; Coturnix; DNA, Complementary; Fibroblasts; Gene Expression Regulation, Neoplastic; Genes, jun; Keratins; Molecular Sequence Data; Multigene Family; Nucleic Acid Hybridization; Protein Conformation

The DNA of human papillomavirus (HPV) types found in cervical carcinomas can immortalize primary human keratinocytes. However, in analogy to tumor progression in vivo, HPV-immortalized keratinocytes require secondary events for malignant conversion. Here, we report on an HPV16-immortalized keratinocyte cell line (HPKIA) which after gamma-irradiation and long term culturing in vitro has acquired the ability to form squamous cell carcinomas in nude mice. The HPV16 integration locus and the viral transcript pattern of HPKIA cells at different passages have remained unaltered. A difference in cytokeratin expression was noted for HPKIA-induced cysts and HPKIA-induced carcinomas. In addition to the expression of suprabasal markers such as cytokeratin 10 and involucrin, carcinomas also express cytokeratin 8 and 18. The latter cytokeratin pair is often expressed in high-grade cervical neoplasia and cervical squamous cell carcinomas. Extensive cytogenetic analyses of nontumorigenic HPKIA cells and their tumorigenic segregants has revealed no single chromosomal abnormality which is confined to all tumorigenic cells. A consistent net loss of chromosomes 3, 5, 9, 12, and 22 was evident for all malignant cells. HPKIA cells represent all stages of transformation and are thus useful for defining secondary genetic events that potentially mark malignant progression in human cells in vivo.

Topics: Animals; Cell Line, Transformed; Cell Transformation, Neoplastic; Female; Humans; Karyotyping; Keratinocytes; Keratins; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Invasiveness; Papillomaviridae; RNA, Messenger

Granular cell tumor (GCT) is a morphologic designation for tumors of varied histogenesis. Most GCTs in human beings are derived from Schwann cells, and rat meningeal GCTs are believed to originate in the neural crest. Three equine pulmonary GCTs from aged horses were studied immunohistochemically with primary antibodies directed against vimentin, cytokeratins (AE1/AE3), S-100, Leu 7, desmin, and neuron-specific enolase (NSE) using a steptavidin-biotin procedure. All three tumors stained similarly with strong and diffuse staining of neoplastic cells for vimentin and S-100 and negative staining with all other antibodies. On the basis of the immunohistochemical results and the previously described histologic and ultrastructural characteristics, equine pulmonary GCT is designated as neural crest and possibly Schwann cell derived, similar to GCT in rats and human beings.

Topics: Animals; Cell Transformation, Neoplastic; Desmin; Female; Granular Cell Tumor; Horse Diseases; Horses; Immunohistochemistry; Keratins; Lung; Lung Neoplasms; Phosphopyruvate Hydratase; Schwann Cells; Vimentin

A five-month-old male baby presented with an abdominal mass which was found on computerised tomography (CT) to be involving the left kidney. Nephrectomy and histopathological study showed morphological featues of a malignant rhabdoid tumour. The tumour cells stained strongly for cytokeratin and epithelial membrane antigen and less intensely for vimentin. Electron microscopy revealed concentric whorled arrays of intermediate filaments within the tumour cell cytoplasm. The child was put on post-operative chemotherapy and radiotherapy but developed bilateral lung metastases and died three months after surgery.

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Fatal Outcome; Humans; Immunoenzyme Techniques; Infant; Keratins; Kidney Glomerulus; Kidney Neoplasms; Male; Mucin-1; Rhabdoid Tumor; Vimentin

Coexpression of the human Met receptor and its ligand, hepatocyte growth factor/scatter factor (HGF/SF), in NIH 3T3 fibroblasts causes the cells to become tumorigenic in nude mice. The resultant tumors display lumen-like morphology, contain carcinoma-like focal areas with intercellular junctions resembling desmosomes, and coexpress epithelial (cytokeratin) and mesenchymal (vimentin) cytoskeletal markers. The tumor cells also display enhanced expression of desmosomal and tight-junction proteins. The apparent mesenchymal to epithelial conversion of the tumor cells mimics the conversion that occurs during embryonic kidney development, suggesting that Met-HGF/SF signaling plays a role in this process as well as in tumors that express both epithelial and mesenchymal markers.

Topics: 3T3 Cells; Animals; Cell Transformation, Neoplastic; Desmosomes; Epithelial Cells; Hepatocyte Growth Factor; Keratins; Kidney; Mesoderm; Mice; Mice, Nude; Neoplasms, Experimental; Proto-Oncogene Mas; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-met; Proto-Oncogenes; Receptor Protein-Tyrosine Kinases; Signal Transduction; Transfection; Vimentin

Intratubular germ cell neoplasia (ITGCN) is now considered to be the preinvasive phase of testicular germ cell tumors with the exceptions of spermatocytic seminoma, pure yolk sac tumor, and mature teratoma. Pagetoid spread of ITGGN into rete testis is a common yet unpublished finding in these cases. We reviewed 100 cases of testicular germ cell tumors from the Surgical Pathology service of Parkland Memorial Hospital (Dallas, TX) to evaluate the frequency of this pattern of spread. Additional sections were obtained from selected cases and were stained with anti-placental alkaline phosphatase, anti-low molecular weight keratin (clone AE1), and various lectins to highlight the process. Pagetoid spread of ITGCN into rete testis was identified in 24 of 60 cases (40%) in which histologic sections contained both ITGCN and rete testis. The incidence of pagetoid ITGCN involvement of the rete testis was lower in pure seminoma (seven of 25 cases [28%]) than in testes containing nonseminomatous germ cell tumors (17 of 35 cases [49%]). AE1 stained the epithelial cells of the rete testis but not the cells of the ITGCN, whereas placental alkaline phosphatase stained the neoplastic cells but not the epithelial cells of the rete testis. These stains were useful in delineating two cases in which the pagetoid involvement was so extensive that they were misdiagnosed as invasive seminomas. Pagetoid spread of ITGCN is a relatively common finding in testicular germ cell tumors and rarely can be mistaken for invasive seminoma. Immunohistochemistry can be helpful in distinguishing florid pagetoid spread from invasive seminoma.

Topics: Adolescent; Adult; Alkaline Phosphatase; Carcinoma in Situ; Cell Transformation, Neoplastic; Diagnosis, Differential; Epithelium; Germinoma; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Orchiectomy; Paget Disease, Extramammary; Seminoma; Testicular Neoplasms

A panel of nine monoclonal antibodies was used to characterize human mesothelioma cell lines that we established from human malignant mesothelioma. The antigens detected were cytokeratin, vimentin, epithelial membrane antigen, carcinoembryonic antigen, Leu-M1 (CD15), desmin, factor VIII-related antigen (von Willebrand factor antigen), OV632, and ME1, a specific monoclonal antibody directed against human malignant mesothelioma. The technique used was the alkaline phosphatase anti-alkaline phosphatase method. All 30 cell lines, either epithelial, sarcomatous, or mixed, showed strong reactivity with cytokeratin and vimentin antibodies. None of the cell lines demonstrated any reactivity with carcinoembryonic antigen, Leu-M1, or factor VIII antibodies; moreover, all of 22 cell lines studied were positive for ME1 antibody and 10 of 12 cell lines studied were positive for OV632. Some interesting features were noted: only two of the 30 cell lines presented a weak positive staining with epithelial membrane antigen, and nine of 19 cell lines tested demonstrated a cytoplasmic staining pattern with desmin antibody. These results show that established human mesothelioma cell lines still possess the immunocytochemical characteristics that are basically consistent with the immunohistochemical features described in tumor tissues of malignant mesothelioma. These characteristics can be used to identify the mesothelioma cells grown from human malignant mesothelioma. Hence, the mesothelioma cell lines will provide a useful tool for the investigation of the cell biology of the tumor and the mechanisms of mesothelial cell transformation, as well as the in vitro evaluation of the effects of some drugs in order to develop new therapies for malignant mesothelioma.

Topics: Actins; Aged; Aged, 80 and over; Antibodies, Monoclonal; Antigens, Differentiation, Myelomonocytic; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; Desmin; Factor VIII; Female; Humans; Immunohistochemistry; Keratins; Male; Membrane Glycoproteins; Mesothelioma; Middle Aged; Mucin-1; Tumor Cells, Cultured; Vimentin; von Willebrand Factor

Several lines of evidence have indicated that rat liver epithelial (RLE) cell lines may be related to a dormant stem cell compartment in the liver in vivo. We have demonstrated that keratin 14 (K14) is expressed together with vimentin in undifferentiated RLE cells. However, upon spontaneous transformation and differentiation to hepatoblast-like progeny the expression of these intermediate filaments (IF) is abrogated, while expression of another set of genes, among others keratin 18 (K18) and alpha-fetoprotein (AFP), is induced (Bisgaard et al., 1994, J. Cell. Physiol., in press). To better understand the mechanisms underlying IF expression during transformation and differentiation of RLE cells we examined the expression and regulation of IFs in clonal cell lines of chemically, oncogene, and spontaneously transformed RLE cells and their resulting tumors. These clonal lines provided a wide variety of tumor phenotypes including trabecular, solid and tubular adenocarcinomas, undifferentiated carcinomas, and spindle cell carcinomas. Northern blot analysis of the cell lines confirmed the differential expression of IF mRNAs. While keratin 8 (K8) was expressed at similar steady-state levels in all cell lines, K14 and vimentin but not K18 were expressed in the majority of cell lines chemically transformed with aflatoxin B1 or by transduction of oncogenes. In contrast, cell lines transformed spontaneously by prolonged passage in vitro expressed K18, while K14 and vimentin were absent. The keratin expression pattern in vitro was retained in the majority of the resulting tumors. However, the keratins expressed in vitro did not accurately predict the tumor phenotype in vivo. In particular, in tumors typed morphologically as adenocarcinomas, the keratin pair typically expressed in chemically transformed tumor cells was K8/K14, whereas K8/K18 was expressed in the tumors derived from spontaneously transformed cell lines. Finally we showed by nuclear run-on and in vitro translation analyses that the expression of K14, K18, and vimentin in transformed RLE cell lines was regulated at the transcriptional level, whereas that of K8 appeared to be posttranslational. These findings suggest that events controlling the differential expression of IF genes are involved in the processes leading to transformation and differentiation of the RLE cell lines. We conclude that the transformed RLE cell lines provide a valuable model to further examine the regulatory mechanisms involved in h

Topics: alpha-Fetoproteins; Animals; Cell Line; Cell Nucleus; Cell Transformation, Neoplastic; Cells, Cultured; Epithelium; Female; Gene Expression; Genes, myc; Keratins; Liver; Mice; Mice, Nude; Oncogene Proteins v-raf; Oncogenes; Phenotype; Protein Biosynthesis; Rats; Rats, Inbred F344; Retroviridae Proteins, Oncogenic; Transcription, Genetic; Transfection; Transplantation, Heterologous; Tumor Cells, Cultured; Vimentin

The cytotoxicity and frequencies of transformation induced by 5 environmental polycyclic aromatic hydrocarbons (PAH) in hamster (M3E3/C3) and rat (WRB K3) lung cells were compared. Both cell strains investigated here retain major metabolic characteristics of the target cells in vivo and are thus able to effectively metabolize, i.e. activate, PAH. Cytotoxic effects of the carcinogen were determined in colony-forming assays and the PAH tested induced dose-dependent cytotoxic responses in the M3E3/C3 and WRB cells. They could then be classified into strong and weak cytotoxicity. Compared to the hamster cell system, the WRB cells were generally shown to be more sensitive. The transforming capacity of the compounds was determined by a soft agar colony formation assay detecting cells with anchorage independency (AI). All PAH investigated induced transformation to AI growth in both cell systems. The transforming activity of the PAH, relative to benzo[a]pyrene (B[a]P) as a reference substance, was determined to facilitate their ranking. This order of transforming potency appears to be similar to that observed in animal studies.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Clone Cells; Cricetinae; Epithelial Cells; Epithelium; Immunohistochemistry; Keratins; Kinetics; Lung; Mesocricetus; Polycyclic Compounds; Rats; Rats, Wistar

Two monoclonal antibodies (MAbs) were established in 20 clones of MAbs generated against cytokeratin fraction extracted from canine squamous cell carcinoma to investigate the expression of intermediate filament proteins during tumorigenesis. These MAbs were confirmed to react with purified cytokeratin by ELISA. One monoclonal antibody, MAb32 reacted all layers of epidermis except the cornified layer and mammary myoepithelial cells but not any epithelial cells. Another antibody named MAb24 exclusively reacted the basal monolayer of epidermis, the stratum germinativum. Any positive reactions with MAb24 were not detected in normal mammary gland. From the analysis by two-dimensional gel electrophoresis followed by immunoblotting, it was revealed that MAb24-recognizing cytokeratin subunit gave a molecular weight of 57 kilodalton and a isoelectric-pH value of pI5.1, indicating type I (acidic) cytokeratin. In intraductal papillomas developed in canine mammary glands, most tumor cells were positively stained with MAb32 in addition of myoepithelial cells while no positive reaction with MAb24 was seen. In ductal carcinomas, MAb24-positive cytokeratin was begun to express by tumor cells with positive reaction of MAb32 where these cells showed infiltrative growth into the stroma. We therefore proposed that 57 kilodalton-type I cytokeratin was a molecular marker for malignant transformation in canine mammary tumor and these antibodies could be useful tools to investigate the change of cytokeratin expression during tumorigenesis.

Topics: Adenofibroma; Animals; Antibodies, Monoclonal; Biomarkers, Tumor; Carcinoma, Ductal, Breast; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Dog Diseases; Dogs; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Epithelium; Female; Immunoblotting; Keratins; Mammary Glands, Animal; Mammary Neoplasms, Animal; Papilloma, Intraductal

Expression of cytoskeletal proteins has been shown to be dependent on the differentiation status of the tissue. In the present study, expression of two cytoskeletal proteins normally present in terminally differentiated keratinocytes, namely cytokeratin types 10/11 and involucrin, were studied in different stages of tumour progression in the oral mucosa. Results showed that cytokeratins 10/11 and involucrin strongly correlated with the differentiation status of cells. High expression was observed in non-dysplastic hyperplastic epithelium as compared to normal, dysplastic and neoplastic epithelium. In addition, various grades of dysplasia showed an inverse correlation with expression of these proteins. Statistical analysis of the results also showed a negative correlation between the differentiation of upper spinal cells and the stage of tumour progression. It therefore appears that the proteins studied may be useful as markers for epithelial carcinogenesis.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Humans; Immunoenzyme Techniques; Keratinocytes; Keratins; Leukoplakia, Oral; Mouth Mucosa; Mouth Neoplasms; Protein Precursors

The stability of cytokeratin during tumor transformation in hepatocellular carcinoma was studied. We applied biochemical methodology to look into the switching of cytokeratin molecules in tumor transformation. First, by centrifugation the cytokeratin molecules were extracted from both liver and hepatoma tissues. The extracts were then soaked with cyanogen bromide-activated Sepharose 4B beads previously coated by monoclonal anti-cytokeratin antibody. The bound molecules were then released from the resin with salt. Second, the isolated molecules of both were treated with lysosomal enzyme and analyzed on two-dimensional gels. The results demonstrated that there was a modulation in cytokeratin molecules, and the hepatoma cytokeratin was generated from the hepatocyte cytokeratin.

Topics: Blotting, Western; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Electrophoresis, Gel, Two-Dimensional; Electrophoresis, Polyacrylamide Gel; Humans; Keratins; Liver; Liver Neoplasms; Peptide Mapping; Precipitin Tests

We examined the expression of carcinoembryonic antigen (CEA) and cytokeratin (CK) as well as the sialo- and sulphomucin content of 40 hyperplastic polyps (HPs), 6 mixed hyperplastic-adenomatous polyps, 30 adenomas and 40 adenocarcinomas of the colorectum. HPs showed a positive CEA expression in 95% of cases and a decreased sialo- and sulphomucin content compared with normal mucosa. Similar changes were observed in adenomas with low-grade dysplasia. The increase in CEA expression from HPs and adenomas to carcinomas was accompanied by a reduction of sialo- and sulphomucins with about three fourths of carcinomas being sialo- and sulphomucin negative. Oncofetal antigen expression concomitant with mucin changes observed in HPs may indicate impaired cellular maturation at a functional level before dysplastic changes become visible. CEA and CK positive macrophages were found in carcinomas predominantly at sites of tumor disruption and necrosis as well as within veins and lymphatic vessels. Our findings suggest that macrophages may play a role in CEA and CK release into the circulation and thus may be determinants of tumor marker serum levels.

Topics: Adenocarcinoma; Adenocarcinoma, Mucinous; Adenomatous Polyps; Biomarkers, Tumor; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; Colon; Colonic Polyps; Colorectal Neoplasms; Humans; Hyperplasia; Keratins; Macrophages; Mucins; Necrosis; Neoplastic Cells, Circulating; Precancerous Conditions; Rectum

The presence of immunoreactive neurofilament proteins has previously been reported in Merkel cell carcinomas but not in normal human epidermal and dermal Merkel cells. We have studied the immunoreactivity of epidermal Merkel cells for neurofilament triplet proteins (68 KD, 70 KD, 160 KD, 200 KD), using epidermal sheets prepared from the plantar skin of human adults, which enabled us to survey large numbers of Merkel cells. Neurofilament protein 200 KD-positive cells were readily identified, while neurofilament protein 68 KD-, 70 KD- and 160 KD-positive cells were largely absent. 200 KD-positive cells in the epidermis were confirmed to represent Merkel cells by a sequential immunoenzyme labeling for the simple epithelial type cytokeratin (No. 8). 200 KD-positive cells were 5.9% of the total number of epidermal Merkel cells. Despite a heterogeneous expression of neurofilament protein subspecies between the normal and transformed Merkel cells, the presence of neurofilament proteins in epidermal Merkel cells may link them to Merkel cell carcinomas.

Topics: Adult; Carcinoma, Merkel Cell; Cell Transformation, Neoplastic; Dendrites; Epidermis; Epithelium; Gene Expression; Humans; Immunohistochemistry; Keratins; Mechanoreceptors; Melanoma; Nerve Tissue Proteins; Neurofilament Proteins; Nevus, Pigmented; Skin; Skin Neoplasms

Wilms tumors (WTs) are embryonic neoplasms of the kidney that are believed to arise from primitive metanephrogenic blastema. Our previous reports and those of others indicate that WTs show an increased expression of insulin-like growth factor II (IGF-II) mRNA. However, the precise role of IGF-II on Wilms tumorigenesis is not known. A central question is to determine whether the increased IGF-II expression in WTs simply reflects the fetal nature of WTs (effect), or is induced by specific changes in gene expression (cause).. This study included 31 sporadic WTs, 7 fetal and 3 adult kidneys and 1 yolk sac tumor. Clinical and histologic summaries of WT cases are shown in Table 1. In WTs, the relative area of blastemal, epithelial, poorly differentiated spindle cell and heterologous cell components were assessed. Dot and Northern blot hybridization, using cDNA probes, were done to assess the level of IGF-II mRNA expression. In situ RNA hybridization was employed to localize IGF-II transcripts. Immunohistochemistry was applied to frozen sections to demonstrate cytokeratin and type-IV collagen. These results were then correlated with the histology of WTs and their precursor lesions, i.e., nephrogenic rests (NRs).. Dot blot hybridization indicated that IGF-II transcripts were 32- to 64-fold more abundant in WTs than in the adjacent uninvolved kidneys. In situ hybridization showed that WTs, NRs, and fetal kidney shared a common feature in which IGF-II transcripts were predominantly associated with blastema. However, WTs and NRs differed from fetal kidney in that occasional epithelial structures and dense blastema showed aberrant, sustained IGF-II expression.. The data indicate two points. 1) There is an inverse correlation between nephroblastic differentiation and IGF-II expression in developing fetal kidney. 2) The IGF-II expression in WTs and NRs does not simply reflect the embryonal nature of the tumor but is rather significantly altered, suggesting a role as a transforming growth factor in Wilms tumorigenesis.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Blotting, Northern; Cell Transformation, Neoplastic; Child; Collagen; Endodermal Sinus Tumor; Epithelium; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization; Insulin-Like Growth Factor II; Keratins; Kidney; Kidney Neoplasms; Male; Middle Aged; Precancerous Conditions; RNA, Messenger; Statistics as Topic; Transcription, Genetic; Transforming Growth Factors; Wilms Tumor

The profile of keratin expression in benign warts from various cutaneous and mucosal sites along with dysplastic warts and squamous cell carcinomas has been examined using a panel of monospecific antibodies to epithelial keratins. Viral warts and verrucous keratoses from immunosuppressed renal transplant recipients show a spectrum of squamous atypia from benign lesions, from minimal changes to full thickness dysplasia. Changes associated with malignancy include loss of differentiation-specific keratins 1 and 10 together with expansion of basal cell epitopes and inappropriate expression of simple epithelial keratins 8, 18, and 19 in advanced squamous cell carcinoma. This late expression of keratins 8 and 18 contrasts with early expression of keratin 17 in all dysplastic lesions examined. Keratin 17 is found suprabasally in hyperproliferative lesions, including benign warts, but marked basal plus suprabasal expression is seen increasingly in malignantly transformed epidermis. These findings were not specific to immunosuppression, as shown by identical findings in control squamous cell carcinoma from nonimmunosuppressed individuals. Keratin 17 expression may prove prognostically helpful when assessing dysplasia in epidermal tumors.

Topics: Biomarkers; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cell Transformation, Viral; Condylomata Acuminata; Epithelium; Female; Humans; Immunohistochemistry; Keratins; Skin; Skin Diseases; Skin Neoplasms; Warts

In this study, we attempted to find a gene or genes which were differentially expressed between a non-tumorigenic rat bladder cell line and a highly tumorigenic/metastatic bladder carcinoma cell line that was derived from the former after treatment in vitro with N-methyl-N-nitrosourea. We cloned a rat keratin 5 cDNA by a differential hybridization technique and found that all of the non-tumorigenic cells (7/7) and normal bladder tissue expressed keratin 5, but most of the tumorigenic cells (8/10) did not express keratin 5. Furthermore, in a spontaneously transformed cell line, keratin 5 expression was lost during the transformation process. These results suggest that loss of keratin 5 expression is closely associated with acquisition of a tumorigenic phenotype by rat bladder non-tumorigenic cells.

Topics: Animals; Base Sequence; Biomarkers, Tumor; Cell Transformation, Neoplastic; Cloning, Molecular; DNA Primers; Epithelial Cells; Epithelium; Gene Expression Regulation, Neoplastic; Gene Library; Keratins; Male; Mice; Mice, Nude; Molecular Probe Techniques; Molecular Sequence Data; Rats; Tumor Cells, Cultured; Urinary Bladder Neoplasms

Topics: Adult; Animals; Breast; Breast Neoplasms; Cell Transformation, Neoplastic; Disease Models, Animal; Epithelium; Female; Humans; Keratins; Mice; Mice, Inbred BALB C; Tumor Cells, Cultured

We have studied in this report the expression of keratins in mouse epidermal keratinocytes transformed in culture by a chemical carcinogen (PDV) and in cell lines (PDVCM1, PDVCM2, PDVCV1, and PDVC57) derived by tumor transplantation of PDV cells in syngeneic C57Bl/6 mice. PDV, PDVCM1, PDVCM2, and PDVCV1 cell lines are weakly to moderately tumorigenic, giving rise to squamous cell carcinomas, although not at all injection points, whereas PDVC57 cells are more malignant, inducing highly anaplastic carcinomas at 100% of injection sites. All the cell lines synthesize anomalously simple epithelial keratins, substantial amounts of K8, and minor quantities of K18 and K19, but the level of expression is increased in PDVC57. We have found that in PDVC57 cells upregulation of K8 is linked to down-regulation of the normal keratins produced by epidermal keratinocytes in culture (i.e., K5, K6, K14, and K17). On the other hand, K8 does not generally colocalize with K13, a keratin also aberrantly expressed by epidermal cell cultures when induced to differentiate by high Ca2+ medium. K13, normally synthesized in internal stratified epithelia, is anomalously induced in mouse epidermal tumors and has been used as an early marker of carcinoma progression. In tumors induced by the cell lines upon injection in mice, K8 is found in the less differentiated regions as opposite to K13, restricted to the differentiating areas of the tumors. In PDV, PDVCM1, PDVCM2, and PDVCV1 carcinomas the overall expression of K13 is higher than that of K8. However, this relation is inverted in tumors induced by PDVC57 cells, in a good correlation with the tumoral phenotypes produced by the cell lines. Our results suggest that upregulation of the simple epithelial keratin K8, as found in transformed epidermal cell lines and tumors, is a late marker of malignant progression and is associated with the loss of the differentiated phenotype.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinoma; Carcinoma, Squamous Cell; Cell Line, Transformed; Cell Transformation, Neoplastic; Genes, ras; Keratinocytes; Keratins; Mice; Mice, Inbred C57BL; Neoplasm Transplantation; Skin Neoplasms; Up-Regulation

The marker profile of 18 samples of normal human ovarian tissues and 138 samples of their derived tumors was established using 51 monoclonal antibodies directed against intermediate filaments, ovarian carcinoma-specific antigens, general tumor-associated antigens and MHC-I/II antigens. Our data show that vimentin and keratins 7, 8, 18, and 19 were found in both epithelial and some nonepithelial ovarian tumors. Several tumor samples contained additional keratins 4, 10, 13, and 14, as well as desmin. BW 495/36 and to a lesser extent HMFG-2 were usually found in all ovarian tumors that contained simple epithelial keratins, except the absence of HMFG-2 in gonadal tumors as well as in dysgerminomas. In contrast to the keratin antibodies, these two panepithelial antibodies were negative in normal mesothelial cells and granulosa cells of the ovarian follicles. In general, the marker TAG-72 appeared useful for its discrimination between positively stained mucinous adenomas, the ovarian carcinomas as well as germ cell tumors, and the negatively stained gonadal tumors, serous adenomas, and cystomas. OV632 appeared useful in the distinction between negatively stained serous adenomas and positively stained serous carcinomas. In contrast, the monoclonal antibodies OC 125, OV-TL 3, OV-TL 16, and MOv 18 can be considered as pan-ovarian carcinoma markers, however without the discriminative capability as seen for OV632. These ovarian carcinoma-associated antigens were hardly found expressed in gonadal and germ cell tumors, except in the group of endodermal sinus tumors. HLA-I was found to be expressed in almost all nucleated cells, although loss of HLA-I expression was seen in areas of tumor cells. HLA-DR was negative in normal ovarian tissue, but heterogeneous expression was noticed in most of the epithelial tumors.

Topics: Adenoma; Antibodies, Monoclonal; Antigens, Differentiation; Biomarkers, Tumor; Cell Transformation, Neoplastic; Diagnosis, Differential; Epithelium; Female; Humans; Keratins; Neoplasms, Germ Cell and Embryonal; Ovarian Neoplasms; Ovary

The regulatory elements of the human keratin K1 gene have been used to target expression of the v-Ha-ras oncogene exclusively in the epidermis of transgenic mice. We developed 12 transgenic mouse lines that express the HK1.ras transgene, producing epidermal hyperplasia in neonates and hyperkeratosis in juveniles. Eventually this skin phenotype diminished but with time adult animals developed papillomas that could persist or regress. The rate and frequency of tumorigenesis appeared to be limited, which suggests that v-Ha-ras requires a second or even third event to elicit and maintain a benign phenotype in transgenic mice. Since in certain transgenic lines papillomas appeared at wound sites, it appears that the promotion stimulus from wounding may be a second event. We envision that such transgenic mice that express v-Ha-ras in the epidermis will become a powerful model for assessing how environmental and molecular factors affect the process of multistage skin carcinogenesis in vivo, as well as a model for evaluating novel therapeutic protocols.

Topics: Aging; Animals; Bacteriophage lambda; Base Sequence; Cell Transformation, Neoplastic; Disease Models, Animal; Female; Fluorescent Antibody Technique; Gene Expression; Gene Expression Regulation, Neoplastic; Genes, ras; Genetic Techniques; Hyperplasia; Keratins; Keratosis; Male; Mice; Mice, Transgenic; Molecular Sequence Data; Papilloma; Polymerase Chain Reaction; Regulatory Sequences, Nucleic Acid; RNA, Neoplasm; Skin Neoplasms

Keratinocyte growth factor (KGF) is a member of the fibroblast growth factor (FGF) family. Synthesized by cells of the dermal component of skin, KGF's potent mitogenic activity is on the epidermal component, which harbors the receptors for this factor. To explore the possible role of KGF in mesenchymal-epithelial interactions in skin, we used a human keratin 14 promoter to target expression of human KGF cDNA to the stratified squamous epithelia of transgenic mice. Mice expressing KGF in their epidermis typically appeared frail and weak, and often had grossly wrinkled skin. These mice exhibited a gross increase in epidermal thickness accompanied by alterations in epidermal growth and differentiation. Most remarkably, animals displayed several striking and unexpected changes, including a marked suppression of hair follicle morphogenesis and suppression of adipogenesis. With age, some animals developed gross transformations in the tongue epithelium and in epidermis. In addition, they exhibited elevated salivation and their salivary glands showed signs of altered differentiation. Collectively, our findings provide new and important insights into the roles of KGF, implicating this potent growth factor in eliciting global effects not only on growth, but also on development and differentiation, of skin and other tissues. In particular, KGF seems to interfere with signalling of some mesenchymal-epithelial interactions.

Topics: Animals; Animals, Newborn; Base Sequence; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; DNA, Single-Stranded; Epithelial Cells; Epithelium; Fibroblast Growth Factor 10; Fibroblast Growth Factor 7; Fibroblast Growth Factors; Growth Substances; Hair; Humans; Keratinocytes; Keratins; Mesoderm; Mice; Mice, Transgenic; Microscopy, Electron; Molecular Sequence Data; Morphogenesis; Phenotype; Polymerase Chain Reaction; Promoter Regions, Genetic; Skin; Skin Physiological Phenomena

Topics: Biomarkers, Tumor; Biopsy; Cell Transformation, Neoplastic; Gastric Mucosa; Gastritis; Gastroscopy; Humans; Keratins; Metaplasia; Polyps; Precancerous Conditions; Stomach Neoplasms; Stomach Ulcer

Intermediate filament proteins have been used to diagnose the origin of specific cells. Classically, vimentin is found in mesenchymal cells, and keratins are present in epithelial cells. However, recent evidence suggests that the coexpression of these phenotype-specific proteins augments tumor cell motility, and hence, metastasis. In the present study, we used the mouse L-cell model to determine if a direct correlation exists between the expression of additional keratins in these cells, which normally express only vimentin, and their migratory ability. Mouse L cells were transfected with human keratins 8, 18, and both 8 and 18. The results indicate that the cells expressing complete keratin filaments have a higher migratory and invasive ability (through extracellular matrix-coated filters) compared with the parental and control-transfected clones. Furthermore, there is an enrichment of keratin-positive cells from a heterogeneous population of L clones selected over serial migrations. This migratory activity was directly correlated with the spreading ability of the cells on Matrigel matrix, in which the keratin-positive transfectants maintain a round morphology for a longer duration, compared with the other L-cell populations. Collectively, these data suggest that keratins may play an important role(s) in migration, through a special interaction with the extracellular environment, thereby influencing cell shape.

Topics: Animals; Cell Adhesion; Cell Movement; Cell Transformation, Neoplastic; DNA; Fluorescent Antibody Technique; Intermediate Filaments; Keratins; Kinetics; L Cells; Mice; Neoplasm Invasiveness; Transfection

Mouse skin carcinomas arise from a small subpopulation of benign papillomas with an increased risk of malignant conversion. These papillomas arise with limited stimulation by tumor promoters, appear rapidly, and do not regress, suggesting that they differ in growth properties from the majority of benign tumors. The transforming growth factor beta (TGF-beta) proteins are expressed in the epidermis and are growth inhibitors for mouse keratinocytes in vitro; altered TGF-beta expression could influence the growth properties of high-risk papillomas. Normal epidermis, tumor promoter-treated epidermis, and skin papillomas at low risk for malignant conversion express TGF-beta 1 in the basal cell compartment and TGF-beta 2 in the suprabasal strata. In low-risk tumors, 90% of the proliferating cells are confined to the basal compartment. In contrast, the majority of high-risk papillomas are devoid of both TGF-beta 1 and TGF-beta 2 as soon as they arise; these tumors have up to 40% of the proliferating cells in the suprabasal layers. Squamous cell carcinomas are also devoid of TGF-beta, suggesting that they arise from the TGF-beta-deficient high-risk papillomas. In some high-risk papillomas, TGF-beta 1 loss can occur first and correlates with basal cell hyperproliferation, while TGF-beta 2 loss correlates with suprabasal hyperproliferation. Similarly, TGF-beta 1-null transgenic mice, which express wild-type levels of TGF-beta 2 in epidermis but no TGF-beta 1 in the basal layer, have a hyperproliferative basal cell layer without suprabasal proliferation. In tumors, loss of TGF-beta is controlled at the posttranscriptional level and is associated with expression of keratin 13, a documented marker of malignant progression. These results show that TGF-beta expression and function are compartmentalized in epidermis and epidermal tumors and that loss of TGF-beta is an early, biologically relevant risk factor for malignant progression.

Topics: Animals; Base Sequence; Carcinoma, Squamous Cell; Cell Division; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Female; Keratins; Mice; Mice, Transgenic; Molecular Sequence Data; Papilloma; Risk; Skin Neoplasms; Transforming Growth Factor beta

Altered cellular responsiveness to growth factors is one of the factors involved in carcinogenesis. In order to study the role of growth factors in transitional-cell carcinogenesis, we established 3 urothelial cell lines from normal mouse urothelium, designated g/G, NUC-5 and NUC-I. These cell lines were studied by light and transmission electron microscopy, karyotyping, grafting in syngeneic mice, growth-factor response in vitro under serum-free conditions, and EGF receptor expression. In the presence of insulin or insulin-like growth factors I and II, proliferation of the non-tumorigenic DNA-tetraploid g/G and DNA aneuploid NUC-5 cells is stimulated by EGF, TGF alpha, bFGF and aFGF. This stimulation can be abolished in g/G but not in NUC-5 cells by simultaneous addition of TGF beta. Proliferation of g/G and NUC-5 cells can also be stimulated by PDGF-AA. The spindle-cell-like NUC-I cells are DNA aneuploid and tumorigenic in syngeneic mice; they express low levels of EGF receptors and their autonomous proliferation is only affected by insulin or insulin-like growth factors. Each of these cell lines seems to reflect a different phase in transitional-cell carcinogenesis: g/G cells have gained immortality, have become tetraploid, but are non-tumorigenic and growth-factor-dependent. NUC-5 cells have become aneuploid, have a growth-factor responsiveness different from that of normal epithelial cells, but are still non-tumorigenic. NUC-I cells are aneuploid, tumorigenic, and growth-factor-independent. These urothelial cell lines provide a suitable tool for further studies in transitional-cell carcinogenesis.

Topics: Animals; Carcinoma, Transitional Cell; Cell Division; Cell Line; Cell Transformation, Neoplastic; Epithelial Cells; ErbB Receptors; Female; Growth Substances; Keratins; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Nude; Ploidies; Urinary Tract; Urologic Neoplasms

Primary diethylstilbestrol-induced kidney tumors from Syrian hamsters were grown in vitro and maintained in culture for 6 mo. Combined immunohistochemical studies using antibodies to intermediate filaments and ultrastructural studies of tumor cells in culture exhibited characteristics similar to tumor cells in vivo. Furthermore, the cells manifested transformed properties in culture; they grew both as multilayered colonies attached to the tissue culture substrate and as floating multicellular colonies (spheroids). When cultured cells were injected into diethylstilbestrol-treated recipient hamsters, tumors developed at the injection sites. In contrast, renal tubules or whole kidney cortex from control hamsters cultured in the same medium underwent only short-term growth, with senescence developing after approximately 1 mo. However, cell cultures of kidney cortex from animals treated in vivo for 5 mo. with diethylstilbestrol formed a cell line. This diethylstilbestrol-induced cell line has been maintained in culture for 1.5 yr and has the following characteristics: a) it is anchorage-dependent, b) it is negative in in vivo tumorigenicity tests, and c) cultured cells are histochemically and ultrastructurally similar to cultured tumor cells. This culture system should prove to be of use in studying hormonal carcinogenesis in vitro.

Topics: Animals; Cell Transformation, Neoplastic; Cricetinae; Desmin; Diethylstilbestrol; Estrogens; Immunohistochemistry; Intermediate Filaments; Keratins; Kidney Cortex; Kidney Neoplasms; Mesocricetus; Microscopy, Electron; Tumor Cells, Cultured; Vimentin

Squamous carcinomas of both human and rodent origin can undergo a transition to a more invasive, metastatic phenotype involving reorganization of the cytoskeleton, loss of cell adhesion molecules such as E-cadherin and acquisition of a fibroblastoid or spindle cell morphology. We have developed a series of cell lines from mouse skin tumors which represent different stages of carcinogenesis, including benign papillomas, and clonally related squamous and spindle carcinomas derived from the same primary tumor. Some spindle cells continue to express keratins, but with a poorly organized keratin filament network, whereas in others no keratin expression is detectable. All of the spindle cells lack expression of the cell adhesion molecule E-cadherin and the desmosomal component desmoplakin. Loss of these cell surface proteins therefore appears to precede the destabilization of the keratin network. At the genetic level, it is not known whether such changes involve activation of dominantly acting oncogenes or loss of a suppressor function which controls epithelial differentiation. To examine this question, we have carried out a series of fusion experiments between a highly malignant mouse skin spindle cell carcinoma and cell lines derived from premalignant or malignant mouse skin tumors, including both squamous and spindle carcinoma variants. The results show that the spindle cell phenotype as determined by cell morphology and lack of expression of keratin, E-cadherin, and desmoplakin proteins, is recessive in all hybrids with squamous cells. The hybrids expressed all of these differentiation markers, and showed suppression of tumorigenicity to a variable level dependent upon the tumorigenic properties of the less malignant fusion partner. Our results suggest that acquisition of the spindle cell phenotype involves functional loss of a gene(s) which controls epithelial differentiation.

Topics: Animals; Antigens, Differentiation; Cadherins; Carcinoma; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Cytoskeleton; Desmoplakins; Epidermis; Fluorescent Antibody Technique; Gene Expression Regulation; Genes, Recessive; Genes, Suppressor; Hybrid Cells; Immunohistochemistry; Keratins; Mice; Phenotype; Skin Neoplasms; Tumor Cells, Cultured

We report a schwannoma with well differentiated ducts resembling cutaneous sweat ducts. The tumor presented as a painless mass near the left knee of a 41-year-old female. The mass had been present for many years. Some increase in size had been noticed over the previous 2 to 3 yr. The bulk of the tumor was composed of spindle cells with the appearance of Antoni A and Antoni B tissue. Rare mitotic figures were noted. In several areas of the tumor, numerous well-differentiated ducts were present. Most resembled normal cutaneous sweat ducts. In some areas, cystic dilatation of the ducts was present. Focal areas demonstrated poorly formed ducts and single cells with prominent nuclei and ample cytoplasm. Well formed ducts, poorly formed ducts, and single cells marked with AE 1/3 (keratin) and epithelial membrane antigen. The spindle cell proliferation marked for S 100 protein and vimentin. Sweat duct differentiation has not been reported previously in either benign or malignant schwannoma. Light and electron microscopic features of this tumor are presented.

Topics: Adult; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Microscopy, Electron; Neurilemmoma; S100 Proteins; Skin Neoplasms; Sweat Glands; Vimentin

Localization of cells with proliferative capacity in human major salivary glands lacks extensive study. Minced fragments of human parotid (n = 3) and submandibular (n = 3) glands embedded in a floating collagen gel matrix and cultured for up to 28 days allowed maintenance of the three-dimensional relationship of the various cell types in these glands. Immunocytochemistry and electron microscopy of a time-dependent series of cultured gland fragments showed gradual cytologic modification of acinar cells so that acini became duct-like but also established that even after 28 days of culture certain cellular features allowed continued identification of acinar cells. Serial section immunostaining for amylase, cytokeratins, and proliferating cell nuclear antigen (a specific marker for cycling cells) revealed that acinar, intercalated duct, and excretory duct (both basal and luminal) cells are all capable of entering the cell cycle. At day 5 of culture, the number of cycling cells increased 16-fold in the parotid gland and 9-fold in the submandibular gland over that in the respective in situ gland. In this in vitro system, which perhaps simulates regenerative processes in human salivary glands, none of the samples showed cycling cells localized only to segments of intercalated duct or the basal cells of excretory duct as suggested by current histogenetic concepts.

Topics: Cell Adhesion; Cell Count; Cell Cycle; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Collagen; Gels; Histocytochemistry; Humans; Immunoenzyme Techniques; Isoproterenol; Keratins; Nuclear Proteins; Parotid Gland; Proliferating Cell Nuclear Antigen; Submandibular Gland; Tissue Embedding

The finding of cytokeratin positivity in non-epithelial cells in 9/15 cases of performing or penetrating gastric ulcer is reported. Two different monoclonal anti-cytokeratin antibodies were used and both produced a strong positivity in spindle and polyglonal cells in the gastric wall. These cells were often distributed near the peritoneal surface, but they were also found in central parts of the gastric wall. The cytokeratin-positive cells had no connection with the gastric mucosa, lacked epithelial features in routinely stained sections and were not positively stained by antibodies to other epithelial markers (EMA and Ber-Ep 4). Many of the cytokeratin-positive cells were also positive for vimentin. There was no evidence of malignancy in any of the cases, but cytokeratin-positive cells like those in the present study may be erroneously interpreted as infiltrating carcinoma. The true nature of the cytokeratin-positive cells was not revealed in the present study. It is concluded that cytokeratin positivity must be evaluated with care and that it is valuable to add antibodies other than anti-cytokeratins for the recognition of epithelial cell differentiation.

Topics: Antibodies, Monoclonal; Cell Transformation, Neoplastic; Gastric Mucosa; Humans; Immunohistochemistry; Keratins; Stomach; Stomach Neoplasms; Stomach Ulcer; Vimentin

Transgenic mice have been previously established that express v-rasHa or v-fos exclusively in the epidermis by means of a targeting vector based on the human keratin 1 gene (HK1). Epidermal expression of v-rasHa (HK1.ras) or v-fos (HK1.fos) resulted in hyperplasia, hyperkeratosis, and later, in benign tumors. To assess the potential for oncogene cooperation in vivo mating experiments were performed. Resultant HK1.fos/ras mice exhibited an obvious increase in the severity of neonatal and juvenile preneoplastic phenotypes, together with the immediate onset of tumorigenesis as compared to single oncogene sibling controls. The HK1.fos/ras tumors grew aggressively and often compromised the animals by 10-12 weeks. However, tumors remained benign as determined by histotype and specific keratin markers. These data indicate that v-fos can cooperate with an initiating v-rasHa phenotype to generate autonomous papillomas, but additional events are required for malignant conversion.

Topics: Animals; Animals, Newborn; Base Sequence; Cell Transformation, Neoplastic; DNA Primers; Fluorescent Antibody Technique; Genes, fos; Genes, ras; Genetic Vectors; Humans; Hyperplasia; Introns; Keratins; Mice; Mice, Transgenic; Molecular Sequence Data; Papilloma; Polymerase Chain Reaction; Skin; Skin Neoplasms; TATA Box

Thirty-one dermal appendage tumors of sweat gland differentiation including 7 spiradenomas (SPA), 8 cylindromas (CYL), 8 acrospiromas (ACS), and 8 chondroid syringomas (CS) were analyzed using antibodies to epithelial membrane antigen (EMA), cytokeratin (AE1, AE3, CAM 5.2, 34BE12), S-100 protein, actin (ACT), and desmin (DES) to characterize the immunocytochemical profile of benign sweat gland tumors. Cytokeratin expression was variable; AE1, 34BE12, AE3, and CAM 5.2 were present in 31, 24, 23, and 22 tumors respectively; 29 tumors contained EMA. Seventeen tumors, (6 SPA, 8 CYL, 2 ACS, 1 CS) stained with antibody to alpha smooth muscle actin, and 26 (7 SPA, 7 CYL, 4 ACS, 8 CS) expressed S-100 protein. Although some prior studies had reported actin filaments on electron microscopy in both spiradenoma and cylindroma, these tumors have previously been considered to be negative for myoepithelial differentiation. All spiradenomas and cylindromas we studied demonstrated actin and/or S-100 protein positivity in basal epithelial cells, consistent with myoepithelial differentiation. The organization of actin and S-100 protein positivity displayed by the spiradenomas and cylindromas we studied suggests that the tumors are differentiated towards the secretory portion of the eccrine sweat gland.

Topics: Acrospiroma; Actins; Adenoma, Sweat Gland; Cell Transformation, Neoplastic; Desmin; Epithelium; Humans; Immune Sera; Immunohistochemistry; Keratins; Membrane Glycoproteins; Mucin-1; S100 Proteins; Sweat Gland Neoplasms; Syringoma

To gain more insight into the differentiation characteristics of the epithelial cells of the adamantinoma of the long bones, we studied the specific keratin immunoreactivity pattern using monoclonal antibodies on 34 primary, recurrent, or metastatic specimens of 22 patients. The results revealed the widespread presence of keratins 14 and 19 in all specimens studied; 74% showed immunoreactivity of keratin 5, and focal staining of keratin 17 was detected in 50%. Keratins 7 and 13 were found in three adamantinoma specimens. This keratin immunoreactivity pattern was independent of histologic subtype, despite marked variety in differentiation pattern, suggesting a common histogenesis for all subtypes of adamantinoma. Furthermore, the pattern was conserved both in local recurrences and in metastasis. The major pattern found in our study differs significantly from other bone and soft tissue tumors with known epithelial characteristics, e.g., synovial sarcomas, chordomas, and epithelioid sarcomas, in that it lacks immunoreactivity of keratins 8 and 18. Our results suggest a basal epithelial cell-like differentiation of adamantinomas.

Topics: Adolescent; Adult; Aged; Ameloblastoma; Antibodies, Monoclonal; Bone Neoplasms; Cell Transformation, Neoplastic; Child; Child, Preschool; Female; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Neoplasm Recurrence, Local

An expression of desmosomal glycoprotein 1 (DG 1) was immunohistochemically examined in 77 biopsies and 21 metastatic cervical lymph nodes of oral squamous cell carcinomas (SCC). In the primary tumors the DG 1 expression was significantly reduced at the invasive site of poorly differentiated and highly invasive tumors. In cases of metastases in cervical lymph nodes, the DG 1 staining at the invasive site of the primary tumor was significantly less than that of nonmetastatic cases. The DG 1 expression in the metastatic lymph nodes was as weak as that in the primary tumor. Thus, we suggest that immunohistochemical investigation of DG 1 expression in oral SCC is valuable in predicting tumor behavior.

Topics: Carcinoma, Squamous Cell; Cell Adhesion Molecules; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Desmoglein 1; Desmoplakins; Desmosomes; Humans; Immunoenzyme Techniques; Immunohistochemistry; Keratins; Lymphatic Metastasis; Mouth Neoplasms; Neoplasm Invasiveness; Neoplasm Staging; Staining and Labeling

A retrospective study of 878 biopsy specimens from 692 patients with laryngeal hyperplastic aberrations was performed according to the Kambic-Lenart classification. Special attention was focused on 88 patients with persistent or recurring disease. In these carcinoma developed in 17 (2.4%) patients, 12 (1.7%) of whom had had atypical hyperplasia. We therefore propose that the term precancerosis, which so definitely implies cancer, should be replaced with the expression risky epithelium where nothing is determined in advance, but a careful follow-up of the patients is imperative. In particular cases of laryngeal hyperplastic lesions, mainly in abnormal and in atypical hyperplasias when the tissue specimens are cut tangentially, the exact identification and position of individual epithelial cells is essential. In such cases histochemical and immunohistochemical methods yield more precise evaluation. Lectins and cytokeratins provide good markers of epithelial maturation. These results contribute to a more useful evaluation of laryngeal hyperplastic lesions, crucial for the choice of adequate therapy.

Topics: Adult; Aged; Aged, 80 and over; Cell Transformation, Neoplastic; Female; Follow-Up Studies; Histocytochemistry; Humans; Hyperplasia; Keratins; Laryngeal Mucosa; Lectins; Male; Middle Aged; Retrospective Studies; Vocal Cords

Activation of a Harvey ras (H-ras) protooncogene is a frequent event associated with mouse epidermal carcinogenesis. We report that the transfection of a human H-ras oncogene into an immortalized mouse epidermal cell line (MCA3D) induces the anomalous expression of cytokeratins (CKs) 8 and 18 characteristic of simple epithelia. The comparison of various transfectant cell clones indicated a direct correlation between the levels of CK8 expression and the mutated H-ras p21s. The expression of simple epithelial CKs is also described in cell lines derived from mouse skin carcinomas (HaCa4, CarC) and in keratinocytes transformed in vitro by a chemical carcinogen (PDV, PDVC57), all of which contain altered H-ras genes. The induction of CK8 and CK18 occurs at the mRNA level and, although both CK8 and CK18 mRNAs are expressed, CK18 protein does not accumulate whereas CK8 is incorporated into intermediate filaments. Immunofluorescence studies show that the pattern of CK8 protein expression is heterogeneous; some cells express very low amounts of CK8, whereas others synthesize relatively high levels of this protein. However, selection of strongly CK8-positive cells was found in one case where a more malignant population of cells (PDVC57) was derived by tumor transplantation of PDV. Our results suggest that activation of a H-ras gene can alter the normal differentiation program of epidermal cells and that the ability to synthesize CK8 and CK18 could be related to tumor progression.

Topics: Animals; Blotting, Northern; Cell Line; Cell Transformation, Neoplastic; Codon; Epidermis; Fluorescent Antibody Technique; Gene Expression; Genes, ras; Harvey murine sarcoma virus; Humans; Intermediate Filament Proteins; Keratinocytes; Keratins; Mice; Mice, Nude; Mutagenesis, Site-Directed; RNA; Transfection

An immunohistochemical study of squamous metaplasia (n = 10), dysplasia (n = 18), squamous cell carcinoma (n = 48) and 3 cases of adenosquamous carcinoma of the uterine cervix with anti-56KD keratin and 68KD keratin antibodies was performed. In the cases of squamous metaplasia, there were two types of staining of which one type had 56KD positive and 68KD negative and another type had both positive. In the cases of dysplasia, there were two types of staining the same as in squamous metaplasia. But in the cases of carcinoma in situ (CIS) (n = 25), there were three types of staining of which the first type had both 56KD and 68KD negative (n = 7), the second type had 56KD positive and 68KD negative (n = 15), and the third type had both 56KD and 68KD positive (n = 3). In invasive carcinoma (n = 23), there were two types of staining the same as in dysplasia of which one type had 56KD positive and 68KD negative (n = 17) and another type had both positive (n = 6). The keratin negative cases in CIS showed morphologically atypical reserve cell hyperplasia composed of atypical small cells with round nuclei and had a small lesion compared with other types. This result suggested that keratin negative CIS was an early form of CIS which was keratin positive. The results indicating that all dysplasia had 56KD keratin positive and CIS had not always 56KD keratin positive suggested that dysplasia was not always a precursor lesion of CIS.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenocarcinoma; Antibodies, Monoclonal; Carcinoma in Situ; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cervix Uteri; Female; Humans; Hyperplasia; Immunoenzyme Techniques; Keratins; Metaplasia; Molecular Weight; Neoplasms, Multiple Primary; Uterine Cervical Neoplasms

Recent ultrastructural, cytogenetic, and ploidy analyses indicate that seminoma acts as a precursor from which other forms of testicular germ cell tumor may originate. Ten cases of primary or metastatic testicular germ cell tumors were investigated that showed possible transformation of seminoma to yolk sac tumor. Such transformation was identified in six cases in which foci of abrupt change from seminoma to various patterns of yolk sac tumor occurred, often at the periphery of otherwise pure lobules of seminoma. Immunostains for cytokeratins, placental-like alkaline phosphatase, and alpha-fetoprotein demonstrated the expected changes in reactivity at the foci of such transformation. Four additional cases were regarded as either seminomas with artifactual microcystic change or the close association of seminoma and yolk sac tumor but lacking evidence for transformation. These data support the theory that seminoma is not an "endpoint" neoplasm but may serve a precursor role in the progression to nonseminomatous germ cell tumors.

Topics: Adolescent; Adult; Alkaline Phosphatase; alpha-Fetoproteins; Biomarkers, Tumor; Cell Transformation, Neoplastic; Dysgerminoma; Humans; Immunohistochemistry; Isoenzymes; Keratins; Male; Mesonephroma; Middle Aged; Testicular Neoplasms

Thirty cases of poorly differentiated carcinomas of the skin were examined for the expression of vimentin. All cases expressed cytokeratins; in addition, 12 cases were positive for vimentin. These were all non-reactive with antibodies to S100 protein, HMB45 and desmin. The finding of vimentin in poorly differentiated squamous cell carcinomas underscores the need for caution in the use of immunohistochemical stains for tumor typing. Cutaneous squamous cell carcinomas are an addition to the list of epithelial tumors which are known to coexpress vimentin intermediate filaments. Other carcinomas which consistently express vimentin include those of renal, endometrial, thyroid, pulmonary, ovarian, salivary gland, adrenal and more recently, those of breast and prostatic origin.

Topics: Actin Cytoskeleton; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Keratins; Skin Neoplasms; Vimentin

The distribution of cytokeratin (CK) polypeptides expressed in syringomas (12 cases) was compared with that in normal eccrine sweat ducts using immunohistochemical techniques on paraffin-embedded tissue. Intradermal and intraepidermal segments of the eccrine duct showed reactivity with an antibody to CK1/5/10/11 in all cell layers, whereas CK19 expression was restricted to the luminal cell layer. CK14 was expressed in all cells of the eccrine duct except for the peripheral cells of the intraepidermal duct. Expression of CK5/6 was seen in the basal cells of the dermal duct and of the lower intraepidermal duct (sweat duct ridge) exclusively. Reactivity with an antibody to CK1 was found in the intermediate cells of the uppermost part of the eccrine dermal duct. In addition, this antibody gave a strong staining of the peripheral cells of the intraepidermal duct, leaving basal cells of the sweat duct ridge and luminal cells unstained. In syringoma, CK distribution was essentially comparable with that found in the uppermost part of the dermal duct and in the sweat duct ridge. Namely, ductal luminal cells expressed CK1/5/10/11, CK19, and variably CK14. Intermediate cells of ductal structures and solid nests were homogeneously stained by antibodies to CK1 and CK1/5/10/11, whereas CK14 was expressed heterogeneously. The basal or outermost layer of ductal structures and solid nests was reactive with antibodies to CK1/5/10/11, CK5/6, and CK14. With regard to CK expression, the results indicate that syringoma represents a tumor differentiating toward both the uppermost part of the dermal duct and the lower intraepidermal duct (sweat duct ridge) of the eccrine sweat gland.

Topics: Adenoma; Cell Differentiation; Cell Transformation, Neoplastic; Eccrine Glands; Epidermis; Humans; Keratins; Peptides; Skin; Sweat Gland Neoplasms

Short-term maintenance of fetal rat colonic tissue in vitro has been demonstrated using a collagen matrix organ culture system. The introduction of single (v-myc, v-rasH, v-src) oncogenes or combinations of oncogenes (v-myc/rasH, v-myc/src) into normal colon mucosal elements was established using retroviral vectors, resulting in enhanced proliferation and migration of epithelial cells from the lumen of tissue implants. Expression of a single oncogene in normal colon epithelium did not result in the establishment of cell lines. In contrast, expression of cooperating oncogenic elements resulted in cell lines in greater than 80% of experiments, revealing different morphological characteristics dependent upon the oncogene combination used. Confirmation of the expression of viral transcripts was determined using Northern blot analysis and viral oncoprotein expression using Western blot analysis (p21) and an immunoprecipitation kinase assay (src). Expression of keratin filaments was lost following passaging of cell lines but could be induced by the myc/ras transformants by growth on Rat-1 feeder layers. This induction phenomenon was not observed with myc/src lines, and although these expressed high levels of sucrase isomaltase the epithelial origin of these cells is unclear. Karyotypic analysis performed on three myc/ras-transformed cell lines revealed a normal chromosome complement associated with transformation. In this report we describe a novel in vitro transformation system for normal rat colonic epithelium mediated by the introduction of oncogene elements using different retroviral vector constructs. The potential to generate cell lines representing different stages of neoplastic progression using relevant genetic components presents significant advantages for the study of cellular and molecular interactions underlying colon neoplastic progression.

Topics: Animals; Blotting, Northern; Blotting, Western; Carrier Proteins; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; CSK Tyrosine-Protein Kinase; Fluorescent Antibody Technique; Gene Expression; Genes, myc; Genes, ras; Genes, src; Karyotyping; Keratins; Membrane Glycoproteins; Microfilament Proteins; Oncogenes; Organ Culture Techniques; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-myc; Proto-Oncogene Proteins p21(ras); Proto-Oncogene Proteins pp60(c-src); Rats; Rats, Inbred WF; RNA; src-Family Kinases; Sucrase-Isomaltase Complex

Interrelationships between neoplastic progression and the expression of intermediate filaments were examined in primary cultures, immortal lines, and Kirsten murine sarcoma virus (KiMSV) transformed lines of rat ovarian surface epithelial (ROSE) cells. Immunofluorescence microscopy revealed abundant keratin filaments in all cells of primary cultures. In immortal, nontumorigenic lines, keratin filaments were detected in fewer cells, in smaller numbers, and in microscopically altered forms. The percentage of keratin-positive cells ranged from 4 to 54%. Its expression was inversely proportional to cell density. Keratin expression was similar in the two immortal lines, although one had retained a monolayered epithelial growth pattern resembling primary cultures, while in the other the growth pattern of the cells was more atypical. The two KiMSV-transformed lines were previously shown to produce tumors in vivo that resemble human ovarian endometrioid stromal sarcomas. In spite of this histologic appearance, the proportion of keratin-positive cells in these cells was increased over the immortal lines. Keratin expression was unrelated to cell density, and keratin in most virally transformed cells was limited to few, fine filaments. In thymidine-labelled immortal and virus-transformed cultures stained for keratin, no correlation was found between keratin expression and proliferative activity. The keratin profiles of primary and immortal cultures were identical on Western blots, with subtypes ranging from 52 to 66 kDa. The two virally transformed lines lacked some of the subtypes. Vimentin networks were faint or absent in primary cultures. In the immortal and the virus-transformed lines, neoplastic progression was associated with increasing vimentin expression but with no changes in filament morphology and distribution. The results show that the abnormalities in intermediate filament expression that accompany immortalization do not preclude the retention of a normal epithelial morphology and growth pattern in this cell type. Furthermore, the number of intermediate filaments and their intracellular distribution appear to be altered at an earlier stage in neoplastic progression than those mechanisms that select for specific keratin subtypes, or those that respond to regulation by cell density. Finally, the presence of keratin in the KiMSV-transformed lines examined in this study supports the hypothesis that human ovarian stromal sarcomas can arise in the OSE.

Topics: Animals; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Epithelium; Female; Immunohistochemistry; Intermediate Filaments; Keratins; Ovarian Neoplasms; Ovary; Rats; Vimentin

Initiation and promotion in mouse skin carcinogenesis produce multiple benign tumors, squamous papillomas, but only a few squamous cell carcinomas. The spontaneous conversion from the benign to the malignant phenotype occurs over many months and in stages, but induced malignant conversion can be accomplished more rapidly by exposure of papilloma-bearing mice to mutagens or by transfection of papilloma cell lines with specific oncogenes. The analysis of genetic targets responsible for carcinogen-induced neoplastic progression would be facilitated by the development of in vitro models where the process is rapid, focal, and quantitative. To this end, primary newborn mouse keratinocytes were initiated in vitro by the introduction of the v-rasHa oncogene via a defective retrovirus. Recipient cells produce squamous papillomas and have a high proliferation rate in culture medium with 0.05 mM Ca2+, but fail to grow in medium with 0.5 mM Ca2+ which is permissive for growth of malignant keratinocytes. When v-rasHa-keratinocytes were exposed to mutagens in vitro, proliferative foci emerged after culture in 0.5 mM Ca2+ for 4 weeks. These foci stained intensely red with rhodamine stain, could be easily quantitated, and readily incorporated bromodeoxyuridine. Dose-response studies with several mutagens indicated that the number of foci increased with concentration to the point where excessive cytotoxicity developed. Mutagens varied in potency for producing foci in the following order: cis-diamminedichloroplatinum greater than or equal to benzo(a)pyrene diolexpoxide I greater than N-methyl-N'-nitro-N-nitrosoguanidine greater than or equal to 4-nitroquinoline-N-oxide greater than N-acetoxy-acetyl- aminofluorene. The tumor promoter 12-O-tetradecanoylphorbol-13-acetate was inactive in the assay. A subset of cell lines derived from foci produced malignant tumors in vivo, while others were not tumorigenic. Analysis of DNA from cell lines and tumors revealed that most tumorigenic cell lines maintained the v-rasHa genome, whereas the viral sequences were deleted in nontumorigenic cell lines. Immunohistochemical analysis indicated that proliferative foci and quiescent v-rasHa keratinocytes expressed keratin 8, a marker of v-rasHa expression in cultured keratinocytes. Cells in foci, but not v-rasHa control cells, expressed keratin 13, a marker which is strongly associated with the malignant progression of skin tumors in vivo. This in vitro assay provides a quantitative model to

Topics: Animals; Animals, Newborn; Base Sequence; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Genes, ras; Keratinocytes; Keratins; Methylnitronitrosoguanidine; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Oligodeoxyribonucleotides; Papilloma; Polymerase Chain Reaction; Skin Neoplasms; Tetradecanoylphorbol Acetate; Transfection

The importance of cervical squamous metaplasia and human papillomavirus 16 (HPV 16) infection for cervical carcinoma has been well established. Nearly 87% of the intraepithelial neoplasia of the cervix occur in the transformation zone, which is composed of squamous metaplastic cells with unclear origin. HPV DNA, mostly HPV 16, has been found in 90% of cervical carcinomas, but only limited experimental data are available to discern the role of HPV 16 in this tissue specific oncogenesis. We have initiated in vivo studies of cultured endocervical cells as an experimental model system for development of cervical neoplasia. Using a modified in vivo implantation system, cultured normal endocervical epithelial cells formed epithelium resembling squamous metaplasia, whereas those immortalized by HPV 16 developed into lesions resembling carcinoma in situ. In contrast, their ectocervical counterparts formed well differentiated stratified squamous epithelium and a lesion with mild dysplastic change, respectively. The HPV 16-immortalized cells showed in vivo cytokeratin expression patterns similar to their respective normal counterparts, confirming their different origins. Thus, this study provides direct experimental evidence for the transformation of simple epithelial cells of endocervical origin into stratified squamous metaplasia and indicates the differential susceptibility of endo- and ectocervical epithelial cells for conversion to cancer by HPV 16.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Transformed; Cell Transformation, Neoplastic; Cells, Cultured; Cervix Uteri; Epithelium; Female; Humans; Immunohistochemistry; Keratins; Metaplasia; Mice; Mice, Nude; Microscopy, Electron; Mucins; Papillomaviridae; Transplantation, Heterologous; Uterine Cervical Neoplasms

In embryogenesis, ovarian surface epithelial cells and ovarian granulosa cells arise through divergent differentiation from a common mesenchymal precursor, the urogenital ridge. In the adult rat, ovarian surface epithelial cells are nonsteroidogenic and keratin positive, while ovarian granulosa cells are steroidogenic and keratin negative. In culture, Kirsten murine sarcoma virus-transformed, tumorigenic ovarian surface epithelial cells continued to express keratin but also became steroidogenic. Transformed ovarian granulosa cells remained steroidogenic but also acquired keratins. Mesodermally derived cells from other sources did not show these differentiation-related changes in response to transformation. The results suggest that v-ras oncogenes may cause the reversion of adult, developmentally related cells to the phenotype of a common, multipotential precursor. They also demonstrate the capacity of v-ras to either induce or reduce the same differentiated characteristic, depending on the developmental history of the target cells.

Topics: 3-Hydroxysteroid Dehydrogenases; Animals; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Female; Fluorescent Antibody Technique; Genes, ras; Granulosa Cells; Keratins; Kirsten murine sarcoma virus; Methionine; Progesterone; Rats; Rats, Inbred F344; Transfection

Primary intraosseous carcinoma (P1OC) of the jaws is rare. They either arise de novo or as a consequence of malignant transformation of a benign cyst or tumor. A 56-year-old patient with a P1OC of the mandible arising from an odontogenic keratocyst is described.

Topics: Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Jaw Cysts; Keratins; Male; Mandibular Diseases; Mandibular Neoplasms; Middle Aged; Odontogenic Tumors

Lectins bind tightly to carbohydrate moieties on cell surfaces. Alterations in lectin binding have been reported to accompany epidermal cell differentiation, marking alterations in membrane sugars during this process. The presence of UEA I (Ulex europaeus agglutinin I) L-fucose-specific lectin-binding sites has been used as a marker for terminally differentiated (committed) keratinocytes. In this article, we report the presence of UEA-I-binding sites on squamous keratinocytes of well-differentiated squamous cell carcinomas, with patchy loss of UEA I positivity on poorly differentiated cells of squamous cell carcinomas, suggesting a possible use for this technique in the rapid assessment of less differentiated areas within the squamous cell tumor. The absence of UEA-I-binding sites on basal cell carcinomas may be related to an inability of cells comprising this tumor to convert the L-D-pyranosyl moiety on basal cells to the L-fucose moiety, resulting in an inability of basal cell carcinoma cell to undergo terminal differentiation into a committed keratinocyte.

Topics: Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Endothelium, Vascular; Epidermis; Epithelium; Fucose; Humans; Keratinocytes; Keratins; Lectins; Plant Lectins; Receptors, Cell Surface; Receptors, Mitogen; Skin Neoplasms

Cell cultures were established from 48 solid basal cell carcinomas (BCC) and from the normal epidermis of the same patients. The growth characteristics and differentiation of BCC cells in vitro were compared with normal keratinocytes (nKC) by using immunohistochemistry, two-dimensional gel electrophoresis including immunoblots, transmission electron microscopy, and soft agar suspension culture. After isolation of the tumor tissue under a stereodissection microscope, explants were cultured on feeder layers of mitomycin-treated 3T3 cells. After 3-5 d, 73% of all explants of BCC could be successfully cultured showing spindle-shaped outgrowing cells. Compared to nKC, cultured BCC cells had a lower growth rate and showed a wider intercellular polymorphism regarding size and shape. Their labeling pattern with a wide panel of monoclonal antibodies showed significant differences from that of nKC. In particular, only weak reactions for various cytokeratins, filaggrin and vimentin depending on the BCC cell type (small, middle, large) were found. Two-dimensional gel electrophoresis revealed expression of keratins 5, 6, 14, 16, and 17 in BCC cells and of K 5, 6, 13, 14, 16, 17, and 19 in nKC. These findings were confirmed by immunoblot. On the ultrastructural level, only a few desmosomes and a lower degree of keratinization markers were detected in BCC cells; finally, when cultured in soft agar BCC cells formed colonies whereas nKC did not. Our findings indicate that cultured BCC cells may preserve in vitro some in vivo characteristics and maintain a growth and differentiation pattern that differs from cultured nKC. The culture model presented here provides further insights into the cytogenetic and histogenetic characteristics of BCC.

Topics: Aged; Aged, 80 and over; Carcinoma, Basal Cell; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Gel, Two-Dimensional; Female; Filaggrin Proteins; Humans; Immunohistochemistry; Keratinocytes; Keratins; Male; Microscopy, Electron; Middle Aged

Keratin 19 (K-19) expression has been strongly correlated with dysplasia in oral epithelium. Expression of K-19 was evaluated by immunoperoxidase staining in formalin-fixed normal ectocervical tissue, normal endocervical tissue, cervical dysplasia, squamous metaplasia, atrophic epithelium, cervical condylomas, and invasive carcinoma to determine if a correlation of K-19 expression with dysplasia was present in the cervical epithelium. Uniform expression of K-19 was seen in endocervical epithelium and in the basal layer of normal ectocervical epithelium in all areas where these epithelia were present. Cervical dysplasia without associated condylomatous changes showed increased expression of K-19 in suprabasal epithelium, corresponding to the level of immature cells. Squamous metaplasia was characterized by scattered cells with increased staining (patch-quilt pattern). There was considerable overlap in the patterns of K-19 expression in dysplastic and metaplastic epithelium. Thus K-19 staining pattern could not be used as a distinctive marker for dysplasia in the cervical epithelium. Atrophic epithelium showed a characteristic uniform but low-level expression of K-19 in suprabasal areas. This pattern may be of diagnostic use in differentiating atrophic lesions from dysplasia. Condylomas showed focal loss of K-19 in the basal layer, suggesting induction of premature differentiation in the basal layer by human papillomavirus infection. Invasive carcinomas showed variable patterns. K-19 is a marker of immature cervical squamous epithelium, with generally distinctive but sometimes overlapping patterns of expression in various diagnostic categories.

Topics: Atrophy; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cervix Uteri; Condylomata Acuminata; Epithelium; Female; Humans; Immunohistochemistry; Keratins; Uterine Cervical Diseases; Uterine Cervical Neoplasms

We report an unusual large, multicystic, posterior fossa neuroepithelial neoplasm involving the cerebellum, brain-stem, and quadrigeminal cistern of a 9-month-old girl. The neoplasm consisted of variably sized, sharply demarcated nests of small cells with a high nuclear-cytoplasmic ratio and moderately basophilic nuclei, embedded in a desmoplastic, immature-appearing, mesenchymal stroma. The nests contained mitoses but none were seen in the stroma. Glial fibrillary acidic protein (GFAP), neurofilament protein, synaptophysin, and cytokeratin (AE-1) were expressed in the nests. Mesenchymal cells were negative for neural markers but positive for vimentin and desmin. The neoplasm was interpreted as a mixed mesenchymal and primitive neuroectodermal tumor (PNET) with histologic features reminiscent of a recently described intraabdominal desmoplastic small cell tumor. The tumor responded poorly to chemotherapy and a second operation was performed 1 year later. The second specimen bore no resemblance to the original and consisted of epithelial-like nests and clusters of neoplastic cells frequently interrupted by sinusoidal vessels. Tumor cells had medium-sized vesicular nuclei with small nucleoli, and a granular cytoplasm. Occasional less cellular islands of neuropil-like tissue contained larger cells having eccentric, vesicular nuclei with prominent nucleoli and abundant pink cytoplasm. Mitoses were not conspicuous. Many cells expressed synaptophysin, neurofilament protein, and GFAP. Neurofilament protein was strongly positive in the larger, neuron-like cells and synaptophysin stained the neuropil-like areas strongly but was less prominent in the neuronal perikarya. Unexpectedly, the neuropil-like areas expressed epithelial membrane antigen, whereas the neuronal cells were negative for chromogranin A. The peculiar histologic picture, combination of phenotypic markers, and remarkable biologic behavior of this unusual tumor defies classification according to existing nomenclature and exemplifies the broad range of phenotypes expressed by primitive neuro-epithelial neoplasms.

Topics: Brain Neoplasms; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Desmin; Female; Glial Fibrillary Acidic Protein; Humans; Immunohistochemistry; Infant; Karyotyping; Keratins; Medulloblastoma; Neoplasms, Germ Cell and Embryonal; Phenotype; Synaptophysin; Terminology as Topic; Vimentin

Undifferentiated (embryonal) sarcoma of the liver is a primitive mesenchymal neoplasm with predilection for individuals in the first 2 decades of life. In this study (10 boys, 6 girls), children in the age range of 6-10 years were most commonly affected (63%). Clinical features most frequently noted on presentation were abdominal pain or a palpable mass. In two cases there was cardiac involvement caused by invasion of the inferior vena cava with extension into the right atrium and ventricle; both children died of progressive dyspnea from tumor embolization to the lungs. One patient was a member of a kindred with the cancer family syndrome (Li-Fraumeni syndrome). There were 13 tumor-related deaths (86% mortality); on child was alive with recurrent tumor in the upper abdomen. Complete surgical resection was attempted in 10 of 15 children who underwent exploratory laparotomy; 2 were alive and well 1 and 5 years later, whereas 1 patient had a recurrence in the upper abdomen 3 years after diagnosis. Ultrastructural study (five cases) and immunohistochemistry (11 cases) supported a mesenchymal origin for the tumor, but failed to identify any diagnostic immunophenotype or specific line of differentiation. Coexpression of vimentin and cytokeratin was seen in three cases. Prompt detection of this aggressive tumor with complete surgical resection is the key to a successful outcome, but this is very difficult to achieve. Recent experience suggests that aggressive adjuvant chemotherapy may improve survival in some cases.

Topics: Adolescent; Adult; alpha 1-Antitrypsin; Antibodies, Monoclonal; Cell Transformation, Neoplastic; Child; Child, Preschool; Factor VIII; Female; Follow-Up Studies; Humans; Immunohistochemistry; Keratins; Liver Neoplasms; Male; Mesenchymoma; Microscopy, Electron

Keratin expression was studied immunohistochemically in 27 ameloblastomas using polyclonal antibody against wide-spectrum keratins (TTL) and monoclonal antibodies against lower- and higher-molecular-weight keratins (PKK1 and KL1), respectively, to clarify the tumor differentiation. Reactions with TTL and KL1 antibodies were generally positive in the stellate cells of the follicular or acanthomatous ameloblastomas. Cell nests of the basal cell type were positive for PKK1. On the other hand, the reactions with TTL or KL1 in the plexiform type were generally weak or absent. From these facts, it was concluded that the follicular, as well as acanthomatous, ameloblastoma is liable to undergo squamous differentiation, whereas the plexiform ameloblastoma remains in primitive stage of tumor differentiation.

Topics: Ameloblastoma; Antibodies; Antibodies, Monoclonal; Cell Nucleus; Cell Transformation, Neoplastic; Cytoplasm; Epithelium; Gingiva; Humans; Immunoenzyme Techniques; Keratins

Forty-seven cases of malignant mixed müllerian tumors were reviewed histologically and studied immunohistochemically with three major objectives: to analyze the histogenetic relationship between the carcinomatous and sarcomatous components of these neoplasms, to ascertain the practical role of immunohistochemical studies in diagnosis and classification, and to determine the prognostic significance of immunohistochemically verified rhabdomyoblastic and neuroendocrine differentiation. Epithelial differentiation (cytokeratin and/or epithelial membrane antigen expression) was confirmed in all carcinomatous components; within the sarcomatous areas, it was identified among individual cells (60% of cases) and within poorly formed clusters of cells (57% of cases). There was a statistically significant tendency for concordant expression of alpha-1-antichymotrypsin, Leu-M1, S-100, Leu-7, and neuron-specific enolase between the carcinomatous and sarcomatous components of individual cases. These two findings provide evidence of common origin for the sarcomatous and carcinomatous components of these neoplasms. Histologic review of metastases in 21 cases revealed a biphasic composition in the majority of metastatic lesions (62%), another feature that further supports a common origin for the two components. From a practical standpoint, immunohistochemistry may be helpful in accentuating the biphasic pattern of these neoplasms and in verifying the presence of rhabdomyoblastic differentiation. In most cases, however, careful morphologic examination and thorough sampling will suffice for correct diagnosis and subclassification. The presence of heterologous, rhabdomyoblastic, or neuroendocrine differentiation did not have a statistically significant influence on survival; the last of these was associated with a tendency for a more rapidly fatal course.

Topics: Actins; alpha 1-Antichymotrypsin; Antigens, Differentiation, Myelomonocytic; Cell Transformation, Neoplastic; Chromogranin A; Chromogranins; Desmin; Female; Genital Neoplasms, Female; Humans; Immunohistochemistry; Keratins; Membrane Glycoproteins; Mucin-1; Myoglobin; Neoplasms, Germ Cell and Embryonal; Prognosis; S100 Proteins; Substance P; Vimentin

Two cases are reported of intra-abdominal small cell tumours expressing concomitant neural and epithelial differentiation. These features were discernible on conventional microscopy and supported immunocytochemically. Immunoreactive vimentin was also revealed in both tumours, and, in addition, one showed focal desmin positivity. Epithelial differentiation in both tumours was confirmed ultrastructurally. The tumours were interpreted to represent a variant of peripheral primitive neuroectodermal tumour, and the report serves to emphasize a potential among such tumours for complex differentiation. The neoplasms are compared with other similar tumours reported recently in children.

Topics: Abdominal Neoplasms; Adolescent; Cell Transformation, Neoplastic; Epithelium; Humans; Immunohistochemistry; Keratins; Male; Membrane Glycoproteins; Microscopy, Electron; Mucin-1; Neoplasms, Germ Cell and Embryonal; Phosphopyruvate Hydratase

Cytogenetic analysis of two pulmonary chondroid hamartomas and nine breast adenofibromas revealed clonal chromosome aberrations in both hamartomas and in four breast tumors. To determine lineage of the cells with chromosome aberrations, a combined immunohistochemical/cytogenetic approach was developed that enabled simultaneous ascertainment of cytogenetic aberrations and immunohistochemical features in individual cells. Immunohistochemical/cytogenetic evaluation of one hamartoma and two adenofibromas demonstrated that neoplastic proliferation, in each case, was confined to the mesenchymal (stromal) component, whereas epithelial cells appeared to be reactive. Cytogenetically abnormal short-term cultures of the remaining hamartoma and another of the breast adenofibromas were composed entirely of mesenchymal elements, indicating mesenchymal clonality in those tumors as well. Our findings support redesignation of pulmonary chondroid hamartomas as 'pulmonary chondromas' and suggest that carcinomas developing within fibroadenomas arise from reactive epithelial proliferation. Combined immunohistochemical/cytogenetic analysis might be useful in the development of novel therapeutic approaches that selectively target neoplastic populations within solid tumors.

Topics: Adenocarcinoma; Breast Neoplasms; Cell Transformation, Neoplastic; Chromosome Aberrations; Hamartoma; Humans; Immunohistochemistry; Karyotyping; Keratins; Lung Neoplasms; Mesoderm; Trisomy; Tumor Cells, Cultured; Vimentin

A system of histological grading based on retrospective analysis of 239 patients with squamous cell carcinoma of the penis is presented. A new scoring system with 4 histological grades was used. The results of this study confirm previous reports that penile cancer is usually a highly (50%) or moderately (29%) differentiated squamous cell carcinoma. Poorly differentiated carcinomas and cancers of other types are very rare. The new grading system was found to be practical and a correlation between histological grade, clinical findings and prognosis was established. Patients with grade 1 tumours had an exceptionally favourable prognosis, with more than 80% being long-term survivors; for these patients, treatment with delayed side effects should be avoided and new forms of treatment should be explored.

Topics: Adult; Age Factors; Aged; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Inflammation; Keratins; Male; Methods; Middle Aged; Penile Neoplasms; Prognosis; Retrospective Studies

Six cases of primary hepatic carcinomas with a significant amount of sarcomatoid elements were examined by using immunohistochemical stainings. Four of the six cases were associated with ordinary hepatocellular carcinoma (HCC), one with cholangiocellular carcinoma (CCC), and one with mixed HCC and CCC. Alpha-fetoprotein and alpha-1-antitrypsin were negative in sarcomatoid cells of all cases; vimentin stained positively in sarcomatoid tumor cells in two of the six cases; and cytokeratin (CK8) was detected in five cases. The CK8 was not detected in tumor cells of two cases of hepatic angiosarcoma, two of metastatic leiomyosarcomas, and one of metastatic fibrosarcoma, although vimentin stained positively in all these true sarcomas. It was concluded that sarcomatoid dedifferentiation of liver carcinomas might derive from both HCC and CCC. In addition CK8 might be an excellent marker to make a differential diagnosis of sarcomatoid cancers from true metastatic or primary sarcomas of the liver.

Topics: Aged; alpha-Fetoproteins; Carcinoembryonic Antigen; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Diagnosis, Differential; Female; Fibrosarcoma; Hemangiosarcoma; Humans; Immunohistochemistry; Keratins; Leiomyosarcoma; Liver Neoplasms; Male; Middle Aged; Sarcoma; Vimentin

Fetal sinusoidal liver cells were isolated from human liver explant cultures and transfected with pCMVLT, a plasmid containing the immediate early promotor of cytomegalovirus (CMV) and the large tumor antigen (LT) coding part of the polyoma virus (py) genome. Whereas nontransfected cells stopped proliferating after 4 weeks, the transfected sinusoidal cells were stimulated to divide more quickly without changes in their morphology. Up to now, cells have been permanently cultured for more than 18 months and passaged over 130 times, corresponding to around 400 generations. This allows them to be regarded as "immortalized" cells. The presence of LT protein in the cells has been documented by means of immunoprecipitation and immunofluorescence. Expression of the v. Willebrandt factor VIII was the main criterion for classifying the cell population as endothelial cells. The presence of cytokeratins 7, 8, and 18 in these cells underlines their close ontogenic and functional relationship to mesothelial cells. Sinusoidal endothelial cells (SECs) synthesize vimentin and the typical extracellular matrix components collagen IV and fibronectin, but are negative for laminin and entactin. We used immortalized SEC's in co-culture experiments with fresh fetal human hepatocytes and adult mouse hepatocytes. They promoted survival of both types of hepatocytes over a period of 8-10 weeks. Control human fetal liver explant culture cells survived for only 3-4 weeks, whereas control adult mouse liver cells retained vitality for 8-10 days only.

Topics: Antigens, Polyomavirus Transforming; Cell Differentiation; Cell Division; Cell Line; Cell Transformation, Neoplastic; Collagen; Endothelium; Humans; In Vitro Techniques; Keratins; Liver; Vimentin; von Willebrand Factor

Topics: Antibodies, Monoclonal; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Breast; Breast Diseases; Breast Neoplasms; Cell Transformation, Neoplastic; Epitopes; Female; Humans; Immunoenzyme Techniques; Keratins; Lactoferrin; Membrane Glycoproteins; Mucin-1; Neoplasm Invasiveness; Neoplasm Metastasis; Precancerous Conditions

Two cases of well-differentiated hepatocellular carcinoma (HCC) with focal biliary differentiation are presented. The distinct histological features of these neoplasms and the unusually protracted clinical course of 8 and 10 years distinguish them from previously described pathological categories of primary hepatic tumors. Electron microscopic and immunohistochemical findings support a dual hepatic and bile duct differentiation of the tumor cells. If additional examples of this tumor are found to be associated with a similarly prolonged symptom-free survival, the distinction of this entity from traditional, rapidly fatal HCC becomes important. Less aggressive therapeutic options may be entertained.

Topics: Adult; alpha 1-Antichymotrypsin; alpha 1-Antitrypsin; Bile Ducts; Carcinoembryonic Antigen; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Chorionic Gonadotropin; Chromogranins; Female; Hepatic Duct, Common; Humans; Immunohistochemistry; Keratins; Liver Neoplasms; Male; Membrane Glycoproteins; Microscopy, Electron; Middle Aged; Mucin-1; Phosphopyruvate Hydratase; Prognosis; Time Factors; Vimentin

Pathologic features of eight cases of undifferentiated (embryonal) sarcoma of the liver (USL) in childhood were studied. Light microscopic examination showed a diffuse growth of spindle cells with occasional polygonal cells and multinucleated giant cells and also revealed focal areas of storiform pattern in four tumors, cambium layer formation in one tumor, and alveolar arrangement in one tumor. Immunohistochemical study showed positive staining of proliferating cells for suggestive histiocytic markers (A1AT in 6/6, A1ACT in 5/6, lysozyme in 4/6, and KP1 in 4/6) and for muscle markers (desmin in 4/6 and HHF35 in 3/6). Ultrastructural examination demonstrated that the individual tumors were composed of a mixture of cells having fibroblastic, histiocytoid, fibrohistiocytoid, myofibroblastic, and undifferentiated (primitive mesenchymal) morphologies. Also identified were cells with definite myoblastic morphology in three tumors: leiomyoblastic in one and rhabdomyoblastic in two. In conclusion, the tumor cells in USL show phenotypical diversity comparable to those of malignant fibrous histiocytoma with or without additional rhabdomyosarcomatous or leiomyosarcomatous differentiation.

Topics: Actins; alpha 1-Antichymotrypsin; alpha 1-Antitrypsin; alpha-Fetoproteins; Cell Transformation, Neoplastic; Desmin; Female; Humans; Immunohistochemistry; Keratins; Liver Neoplasms; Male; Membrane Glycoproteins; Mesenchymoma; Microscopy, Electron; Mucin-1; Muramidase; S100 Proteins; Vimentin

We have previously described the neoplastic transformation of immortalized human bronchial epithelial cells (BEAS-2B) by the combination of the c-raf-1 and c-myc protooncogenes and the concomitant induction of neuron-specific enolase mRNA expression (A. Pfeifer et al., Proc. Natl. Acad. Sci. USA, 86: 10075-10079, 1989). In this paper we describe the morphological, biochemical, and immunohistochemical characteristics of the primary c-raf-1/c-myc tumors, xenografts of these tumors, and tumors that originated from cell lines of the primary neoplasm. The tumors were morphologically characterized by the appearance of desmosomes and tonofilaments, microvilli, and dense core granules representing markers of squamous, glandular, and neuroendocrine differentiation, respectively. A total of 11 of 13 tumors were positive by immunohistochemical techniques for neuron-specific enolase, serotonin (nine of 13), and calcitonin (six of 13). Keratins were expressed in 11 of 13 tumors, and while specific keratins (K5, K7, K16/K17) decreased, there was an increase of vimentin in the tumor cells. Gastrin-releasing peptide immunoreactivity was detectable in a small number of tumors (five of 13). BEAS-2B cells transfected with the c-raf-1 and c-myc protooncogenes and cell lines established from the primary tumors expressed major histocompatibility Class II antigen which has been found on small cell lung carcinoma cells. The tumors induced by the c-raf-1 and c-myc protooncogenes resemble the multidifferentiated phenotype of small cell lung cancer frequently detected in vivo and present a defined model to study the relation between molecular markers, phenotypical appearance, and response to chemotherapeutic agents and radiation.

Topics: Adenocarcinoma; Animals; Antigens, Neoplasm; Bronchi; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Epithelium; Genes, myc; Histocompatibility Antigens Class II; Humans; Immunohistochemistry; Keratins; Lung Neoplasms; Mice; Neoplasm Transplantation; Phosphopyruvate Hydratase; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-raf; Proto-Oncogenes; Transplantation, Heterologous

Bronchial epithelial cells were cultured from an individual with no evidence of malignant disease. These cells, designated HB56B, had a greatly extended in vitro life-span, being able to undergo 50 passages and 200 population doublings in contrast to the usual 3 to 4 passages and 20 to 30 population doublings characteristic of normal human bronchial epithelial cells. HB56B cells had karyotypic evidence of an amplified region on the short arm of chromosome II. Unlike normal bronchial epithelial cells, which undergo terminal squamous differentiation in vitro in response to fetal bovine serum, HB56B cells were only minimally affected by serum. These cells were readily established as an immortalized cell line, HB56B/5T, following transfection with a plasmid containing SV40 early region DNA. HB56B cells were non-tumorigenic in athymic nude mice, but HB56B/5T cells within a few passages of transfection with the SV40 plasmid formed tumors of which 28/37 regressed. HB56B cells may offer an experimental system for the study of proliferation, differentiation, and senescence control in human bronchial epithelial cells.

Topics: Adult; Animals; Antigens, Polyomavirus Transforming; Bronchi; Cell Division; Cell Line; Cell Transformation, Neoplastic; Chromosome Aberrations; Chromosome Disorders; Chromosomes, Human, Pair 11; Culture Techniques; DNA, Neoplasm; Epithelial Cells; Female; Humans; Isoenzymes; Karyotyping; Keratins; Mice; Mice, Nude; Neoplasm Transplantation; Oncogenes; Simian virus 40; Transfection; Transplantation, Heterologous

Several tumorigenic (benign and malignant) clones have been raised from the human epidermal cell line HaCaT after transfection with the c-Ha-ras oncogene (val 12) (P. Boukamp et al., Cancer Res., 50: 2840-2847, 1990). In culture, these HaCaT-ras clones expressed epidermal differentiation markers, such as keratins K1 and 10, at high density or upon depletion of retinoic acid. Accordingly, as HaCaT cells, the clones formed well-differentiated stratified epithelia synthesizing K1 and 10 in surface transplants, while simple and internal epithelial keratins seen in culture were suppressed (as upon retinoic acid depletion in vitro). In transplants of HaCaT cells, in contrast to those of normal keratinocytes, K1 appeared prematurely already in basal cells, while K10 localized rather normally in the suprabasal position. Keratins 1 and 10 were also synthesized in transplants of HaCaT-ras clones (again K1 preceding K10), but both generally shifted toward upper layers. This was particularly evident in thicker transplants of malignant clones. Staining for both keratins persisted "suprabasally" in invasive tissue masses, and this corresponded to their marked expression in solid carcinomas (after s.c. injection), seen by immunofluorescence and two-dimensional gel electrophoresis. Thus, notwithstanding some variations, differentiation potential was not significantly reduced in these clones disregarding levels of ras oncogene expression and malignant properties.

Topics: Cell Line; Cell Transformation, Neoplastic; Clone Cells; Cytoskeletal Proteins; Epidermal Cells; Fluorescent Antibody Technique; Genes, ras; Humans; Keratins; Morphogenesis; Phenotype; Transfection

Topics: Cell Transformation, Neoplastic; Dysgerminoma; Humans; Keratins; Male; Testicular Neoplasms

A differentiation inducer butyrate and a tumor promoter teleocidin had inhibitory effects on the proliferation of PLC/PRF/5 hepatoma. Both of these reagents stimulated the production of procollagen type III peptide, enhanced the cytokeratin assembly and altered the morphological appearance. Novobiocin, a topoisomerase II inhibitor, enhanced the cytokeratin assembly induced by butyrate but antagonized that induced by teleocidin without changing the expression and the phosphorylation state of cytokeratin proteins. In addition, novobiocin acted synergistically with butyrate but not with teleocidin in stimulating the procollagen production and the acetate uptake. These results suggest that butyrate and teleocidin induce cell differentiation via distinct signaling pathway and that novobiocin and butyrate can be used as subsidiary drugs in preventing the growth of hepatoma.

Topics: Acetylation; Butyrates; Butyric Acid; Carcinogens; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Cytoskeletal Proteins; Drug Synergism; Humans; Keratins; Liver Neoplasms; Lyngbya Toxins; Novobiocin; Peptide Fragments; Procollagen; Tumor Cells, Cultured

The L1 antigen was investigated as a marker of squamous differentiation in urothelium using a monoclonal antibody Mac387, and the results were compared with the expression of high molecular weight cytokeratins. L1 antigen was consistently demonstrated in all instances of partial and complete squamous metaplasia and in squamous carcinomas. In contrast, pure transitional cell carcinomas (except one with minor focal staining), adenocarcinomas and normal and hyperplastic urothelium did not label. In a few squamous carcinomas in situ, the pattern of labelling obtained with Mac387 was different from that seen in invasive squamous carcinomas and metaplasias. Compared with high molecular weight cytokeratins, the expression of L1 was more intense in areas of squamous differentiation. L1 expression, as identified by antibody Mac387, may therefore serve as a useful marker of squamous differentiation in urothelial lesions.

Topics: Antibodies, Monoclonal; Carcinoma, Squamous Cell; Carcinoma, Transitional Cell; Cell Adhesion Molecules, Neuronal; Cell Transformation, Neoplastic; Humans; Keratins; Leukocyte L1 Antigen Complex; Metaplasia; Urinary Bladder; Urinary Bladder Neoplasms

We report an undifferentiated sweat gland carcinoma of the vulva in an 80-year-old woman. The tumor, which was located in the right labium majus, resembled an epithelioid sarcoma histologically; it had a granulomatous appearance with multiple tumor nodules containing epithelioid tumor cells. The tumor also contained rhabdoid cells; a large cluster of them showed histological features indistinguishable from those of a malignant rhabdoid tumor. Immunohistochemically, the tumor cells reacted not only for epithelial markers such as cytokeratins, EMA, and CEA, which are known to be expressed by epithelioid sarcoma, but also for CA125 and with monoclonal antibodies recognizing sweat gland structures--namely, EKH5 and EKH6. For comparison, two epithelioid sarcomas and two extrarenal malignant rhabdoid tumors were also studied. Of these tumors, only one extrarenal rhabdoid tumor reacted with EKH5, and none reacted for CA125. Electron-microscopic examination of the present tumor showed the presence of discontinuous basal laminae and tonofibril-like structures as well as primitive cell junctions and interdigitating filopodia. From these findings, we conclude that the tumor was an undifferentiated sweat gland carcinoma mimicking an epithelioid sarcoma. Findings in this case support the idea of the diverse histogenesis of extrarenal malignant rhabdoid tumors and indicate that electron microscopy is important for differentiating epithelioid sarcoma from skin adnexal carcinoma.

Topics: Aged; Aged, 80 and over; Antibodies, Monoclonal; Antigens, Tumor-Associated, Carbohydrate; Carcinoembryonic Antigen; Carcinoma; Cell Transformation, Neoplastic; Diagnosis, Differential; Female; Humans; Immunohistochemistry; Keratins; Membrane Glycoproteins; Microscopy, Electron; Mucin-1; Sarcoma; Skin Neoplasms; Sweat Gland Neoplasms; Sweat Glands; Vulvar Neoplasms

Cutaneous squamous carcinoma with true glandular differentiation has only rarely been documented. Ten patients with such tumors are presented. There were six men and four women, aged 48 to 87 years. The tumors were located on the central face (eight), scalp (one), and hand (one) and consisted of minimally elevated, indurated, keratotic plaques, up to 6 cm in size. Microscopically, the neoplasms exhibited multifocal origin from the epidermis; deep, dispersed, infiltrative growth; perineural invasion; and stromal desmoplasia. Squamous differentiation was most marked superficially. Glandular differentiation was more obvious in deeper areas. Lumens typically developed within squamous nests and were often lined by cells with cytoplasmic vacuoles, some of which contained mucin. The neoplastic cells had obvious cytologic atypia and easily identified mitotic figures. Immunohistochemically, nine neoplasms studied contained carcinoembryonic antigen in glandular foci. Each patient had one or more surgical resections, and six also received radiation and/or chemotherapy. Five patients died with uncontrolled local recurrence, and two are alive with extensive disease and clinical evidence of regional lymph node involvement. Two individuals with small, superficial neoplasms that could be completely removed are disease free. One patient died of unrelated causes shortly after diagnosis. Cutaneous adenosquamous carcinoma is more aggressive than the usual carcinoma of the skin. It must be distinguished from the cytologically bland, microcystic adnexal (sclerosing sweat duct) carcinoma which is capable of recurring but rarely, if ever, proves fatal. The question of whether adenosquamous carcinoma is an epidermally derived squamous tumor with divergent differentiation or should be viewed as a newly recognized adnexal carcinoma remains to be resolved.

Topics: Adenocarcinoma; Aged; Aged, 80 and over; Antibodies, Monoclonal; Carcinoembryonic Antigen; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Male; Middle Aged; Mucins; S100 Proteins; Skin Neoplasms

Histologic diversity is intrinsic to salivary gland tumors, but it is a particular feature of polymorphous low-grade adenocarcinoma as denoted by the terminology. Application of immunohistochemistry and electron microscopy to three examples allowed the study of some aspects of tumor cell differentiation in this minor salivary gland lesion. In one case where the tumor cells were cytologically of one type, immunohistochemistry clearly identified both luminal and nonluminal tumor cells, but the latter showed no evidence of myoepithelial cell differentiation. A second case revealed differentiation only of luminal-type tumor cells, while a third example was largely differentiated as myoepithelial cells but again immunohistochemistry confirmed focal formation of duct-like structures by luminal epithelium. These cases show a considerable range of tumor cell heterogeneity as well as variations in their organization. This variation in differentiation characteristics underlies the histology of polymorphous low-grade adenocarcinomas, and likely occurs in other salivary gland tumors. To establish specific and reliable diagnostic criteria for such tumors requires awareness of this neoplastic process. The limited malignant potential and excellent survival of patients with polymorphous low-grade adenocarcinoma is apparently little affected by patterns of differentiation in this particular neoplasm.

Topics: Actins; Adenocarcinoma; Aged; Aged, 80 and over; Cell Differentiation; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Intermediate Filaments; Keratins; Male; Microscopy, Electron; Neoplasm Invasiveness; S100 Proteins; Salivary Gland Neoplasms; Salivary Glands, Minor; Vimentin

During a routine long-term drug safety study, lasting approximately 2 1/2 yr, male Wistar rats, treated with a prolactin-inhibiting compound, developed an excess of Leydig cell tumors (LCTs). Most tumors were typical for the rat but a small number showed an unusual variation and some appeared malignant. The variation consisted of glandular and/or tubular structures within the tumor mass which occasionally anastomosed and contained an eosinophilic periodic-acid Schiff (PAS) positive material. In a few of these variants, malignant features such as cellular atypia, capsular, and lymphatic invasion and necrosis were seen. No metastases were detected. Detailed morphological and immunohistochemical investigations were conducted in order to establish the cell of origin of these variants. Glandular/tubular structures were found to stain with varying intensity for vimentin and cytokeratin, but were always negative for beta-tubulin. The results indicated that the cell of origin of these LCT variants was indeed the Leydig cell and that glandular and/or tubular structures within LCTs represented a form of Leydig cell metaplasia.

Topics: Animals; Antibodies; Cell Transformation, Neoplastic; Enkephalin, Methionine; Epitopes; Glial Fibrillary Acidic Protein; Immunohistochemistry; Keratins; Leydig Cell Tumor; Male; Phosphopyruvate Hydratase; Rats; Rats, Inbred Strains; S100 Proteins; Sertoli Cells; Substance P; Synaptophysin; Testicular Neoplasms; Tubulin

A new permanent cell line (GRU-1) derived from the lymph-node metastasis of a human epithelioid sarcoma was established in tissue culture. Immunohistochemically, the original tumor had exhibited an intriguing potential for multidirectional differentiation with features of mesenchymal, epithelial and neural differentiation, evidenced by the co-expression of vimentin, cytokeratins and neurofilament proteins, respectively. This capability for multidirectional differentiation was fully preserved in the cultured cells. GRU-1 tumor cells proved to be uniformly positive for vimentin and a considerable proportion of the tumor cells exhibited a positive reaction for cytokeratins and neurofilament proteins. The neural markers neuron-specific enolase (NSE) and synaptophysin were observed in a small proportion of GRU-1 cells. Ultrastructurally, GRU-1 cells showed desmoplastic activity in vitro, being enmeshed by collagen fibrils. DNA distribution, as studied by flow cytophotometry, revealed DNA-diploidy (DNA index = 1) and a G0/G1-proportion of 70.5%. After heterotransplantation in nude mice, GRU-1 tumor cells expressed vimentin and cytokeratin only, whereas the neural markers could not be further demonstrated.

Topics: Adult; Animals; Cell Line; Cell Transformation, Neoplastic; DNA, Neoplasm; Female; Flow Cytometry; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Intermediate Filament Proteins; Keratins; Lymphatic Metastasis; Mice; Mice, Nude; Microscopy, Electron; Neoplasm Transplantation; Neurofilament Proteins; Sarcoma; Tumor Cells, Cultured; Vimentin

The expression of cytokeratin (CK) polypeptides was studied in 59 transitional cell carcinomas (TCC) of the urinary tract of different grade and stage. Using a panel of 14 polypeptide-specific monoclonal CK-antibodies we identified immunohistochemically 8 different CKs separately, ie, CKs 4, 7, 8, 10, 13, 14, 18, and 19, while in immunoblotting studies CK5 expression was detected indirectly by using the antibody RCK102, recognizing CK5 + 8. In low-grade TCCs (G1-G2), the CK distribution was comparable to that in normal urothelium, however with a variable expression of CK13 in the different tumors and a uniform distribution of CK7. In higher-grade TCCs (G3), a decrease in CK13 expression was observed, particularly in the areas of muscle invasion. Furthermore, the appearance and increasing expression of CK14 (not present in normal urothelium or G1 TCCs) with higher grade and stage was striking. With tumor progression changes in epitope configurations of CK8 and CK18 were detected, as concluded from immunohistochemical assays with the panel of monoclonal antibodies for each of these two CKs. In extreme cases this resulted in differential staining patterns of the invasive and noninvasive components within one tumor. In 7 of 32 G3 TCCs, some of which showed areas with evident squamous differentiation, a decrease in the expression of CK7 and/or CK8 was seen. We conclude that tumor progression in TCCs is associated with discrete changes of CK expression, which can be detected using monoclonal antibodies.

Topics: Antibodies, Monoclonal; Carcinoma, Transitional Cell; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Epitopes; Humans; Immunoblotting; Immunohistochemistry; Keratins; Neoplasm Staging; Peptides; Urinary Tract

Somatic cell hybrids between MCF-7 human breast cancer cells and normal immortalized human mammary epithelial cells have been obtained by polyethylene glycol-mediated cell fusion. The hybrid cells are suppressed in their ability to form tumors in nude mice, as well as in traits specific to the tumorigenic MCF-7 parent: growth factor independence, tumor necrosis factor sensitivity, and pS2 gene expression. In addition, they display other characteristics of the "normal" parent, including increased expression relative to the MCF-7 cells of the genes for the extracellular matrix component fibronectin, the intermediate filament keratin 5, and the angiogenesis inhibitor thrombospondin. The levels of keratins 8 and 18 also resemble those of the nontumorigenic parent. These results provide evidence for the existence of tumor suppressor gene products in immortal mammary epithelial cells. We propose a characteristic "suppressed" tumor cell phenotype, which encompasses altered cytoarchitecture, angiogenesis capabilities, and growth factor requirements.

Topics: Animals; Breast; Breast Neoplasms; Cell Division; Cell Fusion; Cell Transformation, Neoplastic; Epithelial Cells; Female; Humans; Hybrid Cells; Keratins; Mice; Mice, Nude; Subrenal Capsule Assay; Transfection; Tumor Cells, Cultured

A series of adrenal cortical adenomas (ACA) and carcinomas (ACC), as well as normal adrenal cortex have been studied by a panel of 11 antibodies to characterize antigenic changes that may distinguish these morphologically similar entities. Normal adrenal cortex and ACA express low-molecular weight cytokeratin intermediate filaments. However, none of the six primary or seven metastatic ACCs were found to express detectable levels of cytokeratins. In contrast, vimentin was seen in all ACCs studied and was heterogeneously expressed by ACAs. However, its expression was usually confined to stromal elements of the normal adrenal cortex. We conclude that adrenal cortical cells undergo characteristic changes in intermediate filament expression during the process of neoplastic conversion and malignant transformation. Undetectable expression of cytokeratins and strong expression of vimentin is associated with malignant adrenal cortical lesions. In addition, we examined the antigenic phenotype of a series of primary renal cell carcinomas (RCC). Renal cell carcinomas express cytokeratins, while ACCs do not. The majority of primary RCCs express Lewis blood group isoantigens (most commonly Lewis X), while ACAs and ACCs do not. The panel of antibodies described here may help to distinguish morphologically similar lesions of like histogenesis (ACAs vs. ACCs) and lesions of different histogenesis (adrenal vs. renal) on the basis of their composite antigenic phenotypes.

Topics: Adenoma; Adrenal Cortex; Adrenal Gland Neoplasms; Antigens; Carcinoma; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Isoantigens; Keratins; Kidney Cortex; Kidney Neoplasms; Vimentin

The relationship between the nuclear DNA content, the immunohistochemical findings, the clinical characteristics (tumor volume doubling time and survival) and the cytomorphologic features of small cell poorly differentiated squamous cell carcinoma of the lung was studied in ten cases. There were no significant correlations between the immunohistochemical stainings for neuron-specific enolase and keratin and the clinical characteristics in these cases. The DNA histogram patterns were classified as type I or type II, depending on the degree of dispersion of values. There was no relationship between the immunohistochemical findings and the DNA histogram patterns. Only the DNA histogram patterns were related to some of the clinical characteristics: patients with type II histograms had significantly shorter tumor volume doubling times than did patients with type I histograms. Such information may aid in distinguishing the small cell type of poorly differentiated squamous carcinoma from classic small cell carcinoma of the lung, with which it may be confused.

Topics: Aged; Aged, 80 and over; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; DNA, Neoplasm; Female; Humans; Immunohistochemistry; Keratins; Lung Neoplasms; Male; Middle Aged; Phosphopyruvate Hydratase; Prognosis

Cyclopentenone prostaglandins (PGs) such as delta 12-PGJ2 and PGA are potent inducers of growth inhibition in a variety of cultured cells, including epidermal cells. These PGs are actively transported into cells by a specific carrier on cell membrane and accumulate in cell nuclei with binding to nuclear protein. To clarify the mechanism of cytotoxicity of these PGs in epidermal cells, we examined the effects of delta 12-PGJ2 on protein synthesis and cytoskeleton in the PAM 212 transformed mouse epidermal cell line. Cycloheximide at 1 microgram/ml culture medium exhibited a protective effect on cell growth inhibition of PAM 212 cells by delta 12-PGJ2. The analysis of cell lysate protein patterns by SDS-polyacrylamide gel electrophoresis revealed that 12-h incubation with delta 12-PGJ2 increased the amount of 70 kD protein in PAM 212 cells. The amount of 70 kD protein in delta 12-PGJ2-treated cells was markedly decreased by cotreatment with cycloheximide. This 70 kD protein was also induced in PAM 212 cells with treatment at 43 degrees C for 90 min, indicating that this synthesized protein belongs to the heat shock protein. The addition of delta 12-PGJ2 to confluent PAM 212 cells resulted in the disappearance of action filament, as visualized by fluorescent labeled phallacidine, but in contrast, keratin filament appeared to be intact during 12-h incubation with delta 12-PGJ2 at a concentration of 5 micrograms/ml culture medium. These results suggest that the cytotoxicity of cyclopentenone PGs is at least in part due to induction of the synthesis of some protein(s), probably one of the heat shock proteins, and the damage to the actin filament in transformed cultured epidermal cells.

Topics: Actin Cytoskeleton; Actins; Animals; Antineoplastic Agents; Cell Transformation, Neoplastic; Cells, Cultured; Cycloheximide; Cytoskeleton; Epidermal Cells; Epidermis; Keratins; Mice; Mice, Inbred BALB C; Prostaglandin D2; Protein Biosynthesis

The early stages of the carcinogenic process induced by aflatoxin B1 (AFB1) in rat liver during 24 weeks of feeding and the resulting tumours have been studied with respect to cytokeratin (CK) expression. A previously uncharacterized monoclonal antibody, MRCTU/J1, has been shown to recognize rat CK18 and together with antibodies against human CK8, 18 and 19, has been used to examine the possible lineage of tumour cells and also to identify the altered foci that might be most relevant to tumorigenesis. Results suggested that AFB1-induced transformation in liver may occur in more than one cell type, since tumours with the normal hepatocyte CK pattern and those with bile duct or oval cell CK phenotype were identified. Additionally, hepatocytes with a bile duct CK phenotype appeared during the early stages of carcinogenesis. The in vivo pattern of CK expression also appeared to be maintained in one normal and one hepatoma-derived cell line. Overexpression of CKs (particularly of CK19) was a much more selective marker for altered foci, compared to gamma-glutamyltranspeptidase, and was more consistently expressed at high levels in tumours, suggesting that it might be a more reliable way of identifying those cells involved in the transformation process.

Topics: Aflatoxin B1; Aflatoxins; Animals; Antibodies, Monoclonal; Antibodies, Neoplasm; Cell Transformation, Neoplastic; Cross Reactions; Cytoskeleton; Intermediate Filaments; Keratins; Liver; Liver Neoplasms, Experimental; Male; Rats; Rats, Inbred F344; Tumor Cells, Cultured

We wished to assess the antigenic expression of primary lung tumors diagnosed as either carcinosarcoma or sarcoma in order to determine whether this information would be useful in distinguishing the two. We therefore immunohistochemically analyzed six pulmonary carcinosarcomas and five primary lung sarcomas for the presence of carcinoembryonic antigen (CEA), S100 protein, cytokeratin and vimentin using commercially available monoclonal and polyclonal antibodies on formalin fixed tissues. Six of six carcinosarcomas stained positively for cytokeratin while none of the sarcomas stained. In three carcinosarcomas both the carcinomatous and sarcomatous areas were positive while in three only the carcinomatous areas were positive. CEA staining was present in five carcinosarcomas and absent in all the sarcomas. CEA positivity was strong and not confined to those tumors with obvious gland formation. Staining for S100 protein was positive in two carcinosarcomas but only in those areas showing chondroid differentiation. Immunohistochemical staining for vimentin using two different monoclonal antibodies gave inconsistent results. We conclude that in differentiating between a carcinosarcoma and a sarcoma of the lung, immunohistochemical staining for both cytokeratin and CEA are useful with cytokeratin marginally preferable. The data indicate that carcinosarcoma of the lung, like that of the upper aerodigestive tract, expresses antigens suggesting both epithelial and mesenchymal differentiation.

Topics: Antigens, Neoplasm; Carcinoembryonic Antigen; Carcinosarcoma; Cell Transformation, Neoplastic; Diagnosis, Differential; Epithelium; Humans; Immunohistochemistry; Keratins; Lung Neoplasms; Mesoderm; S100 Proteins; Sarcoma; Vimentin

The cell differentiation properties of thirty-four sacrococcygeal teratomas (SCT) and their five recurrences were immunohistochemically studied for the expression of different classes of intermediate filament proteins, muscle actin (MA) and S-100 protein. Out of thirty-nine tumors twenty-three were SCTs with only mature tissue elements, seven immature teratomas, five pure endodermal sinus tumors (EST) and four ESTs or embryonal carcinomas (EC) combined with mature components. Cytokeratin positivity was found in all epithelial structures and sometimes also in smooth muscle and primitive mesenchymal cells. An intense cytokeratin immunoreactivity was observed in EC and EST components. Muscle markers, desmin and MA were present in smooth and striated muscle cells. Focal desmin positivity was also found in some epithelial structures in two cases. Glial tissue positive for glial fibrillary acidic protein (GFAP) was found in twenty-eight out of thirty-nine tumors. Some cases with no apparent glial tissue in hematoxylin and eosin staining showed glial differentiation as proved by GFAP positivity. In six out of eleven choroid plexus-like tissues GFAP positive cells were observed. S-100 protein showed an intense distribution of immunoreactivity outside neural tissue, and focal positivity was observed in malignant epithelial structures. Immunohistochemical markers did not reveal any prognostic significance in teratomas. Our findings, however, showed some aberrant features of cell differentiation from normal mature tissue components but closely parallel to those found in normal fetal development.

Topics: Actins; Cell Transformation, Neoplastic; Child; Desmin; Female; Follow-Up Studies; Glial Fibrillary Acidic Protein; Humans; Infant; Infant, Newborn; Keratins; Male; Mesonephroma; Neoplasms; Prognosis; S100 Proteins; Sacrococcygeal Region; Teratoma

The histogenesis of localized fibrous tumor of the pleura (LFTP) is controversial. We studied 12 LFTP's by light microscopy; by immunohistochemical staining for cytokeratin (CK), vimentin, muscle-specific actin, desmin, S-100 protein, epithelial membrane antigen (EMA) and factor VIII; by electron microscopy in 6 tumors; and by lung digestion for asbestos bodies in 4 cases. Three histologic patterns occurred in combination: 1) collagenous, 2) cellular and 3) hypocellular/myxoid. Hemangiopericytoma-like foci were prominent in the cellular areas of 9 tumors. Unusual features included diffuse small cells in 3 tumors, microcystic foci in 2, macrocystic areas in 5 and tumor giant cells in 4 tumors. Neoplastic cells in all patterns stained positively for vimentin and actin in 9 and 4 tumors, respectively, and were negative for all other markers. CK and EMA were identified in mesothelial and epithelial invaginations only. Ultrastructurally, neoplastic cells demonstrated intercellular junctions, intermediate or thin filaments, dense bodies and rough endoplasmic reticulum. Basal lamina was focally present in 5 tumors, while tonofilaments, desmosomes and short microvilli were observed in one case. Our results support the conclusion that LFTP is a neoplasm of the multipotential subserosal cell, and usually expresses mesenchymal (fibroblastic/myofibroblastic) differentiation. Coexpression of mesothelial features is rare. Lung asbestos body quantitation in 4 patients suggests that there is no association between LFTP and asbestos exposure.

Topics: Actins; Adult; Aged; Asbestos; Cell Transformation, Neoplastic; Desmin; Factor VIII; Female; Humans; Immunohistochemistry; Keratins; Lung; Lung Diseases; Male; Membrane Glycoproteins; Mesoderm; Mesothelioma; Microscopy, Electron; Middle Aged; Mucin-1; Pleural Neoplasms; Vimentin

We present four cases of a malignant thyroid tumor showing morphologic, immunocytochemical, and ultrastructural features of endothelial cell differentiation. The tumor cells had epithelioid features and displayed strong immunoreactivity for keratin. There was no evidence of follicular or C-cell differentiation in any instance. We interpreted these cases as keratin-positive epithelioid angiosarcomas. The findings presented here support the existence of primary malignant vascular tumors in the thyroid even in the presence of keratin positivity, a marker traditionally regarded as indicative of epithelial differentiation.

Topics: Aged; Aged, 80 and over; Cell Transformation, Neoplastic; Endothelium; Female; Hemangiosarcoma; Humans; Immunohistochemistry; Keratins; Lectins; Male; Microscopy, Electron; Middle Aged; Plant Lectins; Thyroid Neoplasms; Vimentin; von Willebrand Factor

We analyzed the normal/mutated allelic ratio of the Ha-ras-1 gene in mouse skin squamous cell carcinomas induced by initiation with dimethylbenz[a]anthracene and promotion with phorbol 12-myristate 13-acetate. DNA for these studies was obtained from short-term tumor cultures (24-72 hr) to eliminate the contribution of stromal and inflammatory cells to the sample. The allelotypic analysis was performed in 25 squamous cell carcinomas by quantitative radio-analysis of the Xba I restriction fragment length polymorphism as detected by BS9, a v-Ha-ras probe, and rehybridization of the Southern blots with probes for chromosomes 7 and 8. Approximately 85% of the tumors presented overrepresentation of the mutated allele in the form of 1 normal/2 mutated (12 tumors), 0 normal/3 mutated (4 tumors), 0 normal/2 mutated (3 tumors), and gene amplification (3 tumors). No tumor was found with a 2 normal/1 mutated allelic ratio. These results support our previous cytogenetic studies, indicating that trisomy of chromosome 7 is present in the majority of these tumors and show that nonrandom duplication of the chromosome carrying the mutated Ha-ras-1 allele appears to be a major mechanism by which the mutated gene is overrepresented.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Alleles; Animals; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Chromosome Mapping; Gene Amplification; Genes, ras; Immunoenzyme Techniques; Keratins; Mice; Mice, Inbred Strains; Models, Genetic; Mutation; Nucleic Acid Hybridization; Polymorphism, Restriction Fragment Length; Skin Neoplasms

To overcome the difficulty of assessing oncogene action in human epithelial cell types, such as thyroid, which have limited proliferative potential in culture, we have explored the use of temperature-sensitive (ts) mutants of simian virus 40 (SV40) early region to create conditionally immortalized epithelial cell lines. Normal primary cultures of human thyroid follicular cells were transfected with a plasmid containing the SV40 early region from mutant tsA58. Expanding epithelial colonies were observed after 2 to 3 months, all of which grew to greater than 200 population doublings without crisis. All showed tight temperature dependence for growth. After switch-up to the restrictive temperature (40.5 degrees C), no further increase in cell number was seen after 1 to 2 days. However, DNA synthesis declined much more slowly; the dissociation from cell division led to marked polyploidy. Viability was maintained for up to 2 weeks. Introduction of an inducible mutant ras gene into ts thyroid cells led, as expected, to morphological transformation at the permissive temperature when ras was induced. Interestingly, this was associated with a marked reduction in net growth rate. At the restrictive temperature, induction of mutant ras caused rapid cell death. These results demonstrate the utility of a ts SV40 mutant to permit the study of oncogene action in an otherwise nonproliferative target cell and reveal important differences in the interaction between ras and SV40 T in these epithelial cells compared with previously studied cell types.

Topics: Animals; Antigens, Viral, Tumor; Blotting, Northern; Cell Adhesion; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Clone Cells; Epithelial Cells; Gene Expression; Humans; In Vitro Techniques; Iodine; Karyotyping; Keratins; Mice; Mice, Nude; Mutation; Neoplasms, Experimental; Oncogene Protein p21(ras); Phenotype; RNA, Messenger; Temperature; Thyroglobulin; Thyroid Gland; Transfection

We studied the proliferation and differentiation of human laryngeal papillomas, which are benign tumors induced by human papillomaviruses. Immunofluorescent stains of tissues for a number of differentiation-specific proteins showed abnormal differentiation. Papilloma tissue fragments in vitro showed a slightly decreased fraction of proliferating cells that incorporated tritiated thymidine and a markedly reduced incorporation of tritiated uridine when compared with normal tissue. We propose that papillomavirus infection results in normal basal cell proliferation but abnormal terminal differentiation and that this abnormality significantly contributes to the hyperplasia of the papillomas.

Topics: Cell Division; Cell Transformation, Neoplastic; Filaggrin Proteins; Humans; Immunoblotting; Intermediate Filament Proteins; Keratins; Laryngeal Neoplasms; Neoplasm Proteins; Papilloma; Papillomaviridae; Protein Precursors; Staining and Labeling; Thymidine; Tumor Virus Infections; Uridine

A case of neuroendocrine lung tumor located beneath the pleura in a 71-year-old woman is reported. At autopsy, the tumor was found to have metastasized to the bones and liver without involving the hilar lymph nodes. Histologically, the tumor cells at the primary site and in the liver metastasis exhibited a carcinoid-like organoid structure, whereas pleomorphic giant cells were noted in the bone metastasis. The argyrophilic tumor cells were immunoreactive for neuron-specific enolase, chromogranin A, serotonin, calcitonin, calcitonin gene-related peptide, gastrin-releasing peptide, neuropeptide Y, gastrin, pancreatic polypeptide, glicentin, the alpha-subunit of human chorionic gonadotropin, keratin, epithelial membrane antigen, Leu M1 and carcinoembryonic antigen. Electron microscopy revealed abundant neurosecretory granules in the cytoplasm. This was considered to be a rare case of neuroendocrine lung tumor showing carcinoid-like histology at the primary site and large-cell transformation in bone metastasis.

Topics: Aged; Autopsy; Bone Neoplasms; Calcitonin; Calcitonin Gene-Related Peptide; Carcinoembryonic Antigen; Carcinoid Tumor; Cell Transformation, Neoplastic; Chorionic Gonadotropin; Chromogranin A; Chromogranins; Female; Giant Cell Tumors; Humans; Immunohistochemistry; Keratins; Lung Neoplasms; Membrane Glycoproteins; Microscopy, Electron; Mucin-1; Phosphopyruvate Hydratase; S100 Proteins; Serotonin

Continuous epithelial cell lines from individuals with cystic fibrosis (CF) and normal controls are required to understand the genetic and cellular defects in CF. We used retroviruses to transduce SV40 large T antigen into nasal epithelial cells. Transformed continuous cell lines were isolated that expressed epithelial markers, cytokeratin, and tight junctions. Northern blot analysis shows that all of the cell lines express the putative CF gene mRNA. Studies of transepithelial electrolyte transport show that CF and normal cell lines develop a transepithelial electrical resistance. Normal but not CF cell lines secreted Cl- in response to agonists that increase cellular levels of adenosine 3',5'-cyclic monophosphate (cAMP) (isoproterenol, forskolin, and a membrane-permeant analogue of cAMP) or in response to a tumor-promoting phorbol ester that activates protein kinase C. In contrast, the Ca2(+)-elevating agonist bradykinin and the Ca2+ ionophore A23187 stimulated secretion in both normal and CF cell lines. The continuous cell lines we have produced maintain their proper phenotypes and will serve as useful tools in understanding the pathophysiology of CF.

Topics: Antigens, Polyomavirus Transforming; Base Sequence; Biomarkers; Blood Proteins; Bradykinin; Calcimycin; Calgranulin A; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Chloride Channels; Chlorides; Colforsin; Cystic Fibrosis; Epithelial Cells; Epithelium; Genotype; Humans; Ion Channels; Keratins; Membrane Potentials; Membrane Proteins; Molecular Sequence Data; Nasal Mucosa; Oligonucleotide Probes; Phenotype; Polymerase Chain Reaction; Reference Values; Retroviridae; Simian virus 40; Tetradecanoylphorbol Acetate; Theophylline

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Child; Connective Tissue; Humans; Immunoenzyme Techniques; Keratins; Phosphopyruvate Hydratase; Rhabdomyosarcoma; S100 Proteins; Soft Tissue Neoplasms

We have previously shown that activation of ras oncogenes by mutation is a frequent early event in human thyroid neoplasia. Using amphotropic retroviral vectors to achieve gene transfer, we demonstrate here that human primary thyroid epithelial cells can be partially transformed by an activated cellular or viral Ha-ras oncogene, in the absence of a cooperating oncogene. The transformation event induced by ras involves temporary rescue from senescence for up to 20 rounds of cell division together with morphological alteration, growth factor independence and anchorage independence. It has therefore been possible to reconstruct in vitro a key early event in the genesis of human epithelial neoplasia.

Topics: Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Genes, ras; Genetic Vectors; Humans; Immunoblotting; Immunohistochemistry; Keratins; Mutation; Thyroglobulin; Thyroid Gland; Thyroid Neoplasms

We have characterized the cells that form the human oral epithelia by analyzing their patterns of keratin expression in culture and in transplants. Keratinocytes of all oral regions synthesized high levels of keratins K5/K14 and K6/K16,K17, as expressed by cells of all stratified squamous epithelia in culture. However, cells from different regions varied in their expression in culture of retinoid-inducible (K19 and K13) and simple epithelial (K7, K8 and K18) keratins. By these criteria, all oral cells could be classified as belonging to one of three intrinsically distinct subtypes: "keratinizing" (gingiva, hard palate), "typical nonkeratinizing" (inner cheek, floor of mouth, ventral tongue) and "special non-keratinizing" (soft palate), all of which differed from the epidermal keratinocyte subtype. Cells from fetal floor of mouth expressed a pattern of keratins in culture markedly different from that of adult floor of mouth cells but identical to that of the adult "special nonkeratinizing" subtype and similar to that of several oral squamous cell carcinoma lines. When cultures of oral keratinocytes were grafted to the dermis of nude mice, they formed stratified epithelial structures after 10 days. In some areas of the stratified structures, the basal layer recapitulated the K19 expression pattern of the oral region from which they had originated. Thus, regional differentiation of the oral epithelium is based on an intrinsic specialization of regional keratinocyte stem cells. Additionally, oral cell transformation either frequently involves reversion to the fetal keratin program or else oral cells that express this keratin program are especially susceptible to transformation.

Topics: Animals; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Gene Expression Regulation; Humans; Keratinocytes; Keratins; Mice; Mouth Mucosa; Transplantation, Heterologous

To investigate the antineoplastic activity of parvoviruses, proliferating normal human epidermal cells and a series of established keratinocyte cell lines derived from squamous cell carcinomas or transformed in vitro, were compared for the outcome of H-1 virus infection. All established keratinocyte cell lines were more sensitive to killing by H-1 virus than normal epidermal cells, although to varying extents. Using a step-wise procedure for malignant transformation in vitro, we found that sensitization of transformed epidermal cells to H-1 virus can be dissociated from the acquisition of a tumorigenic phenotype. Thus, spontaneously- or SV40-immortalized human keratinocytes were moderately and highly sensitive to H-1 virus, respectively, and could be made tumorigenic by Harvey-ras oncogene transfection without a major change in their susceptibility to the virus. The capacity of human keratinocytes for replicating and expressing H-1 virus DNA appears to be a revealer of cellular alterations that take place in at least some pathways to malignant transformation but that may be insufficient to confer a tumorigenic potential.

Topics: Cell Survival; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cytopathogenic Effect, Viral; DNA, Viral; Epidermis; Humans; Keratins; Parvoviridae; Tumor Cells, Cultured; Virus Replication

Morphological observations suggest that rate tracheal epithelial (RTE) cells undergo squamous differentiation when maintained in cell culture. The purpose of the studies presented here was to examine and define differentiation of cultured RTE cells with the help of markers previously shown to be specific for squamous differentiation. Furthermore, we wanted to determine whether neoplastic transformation of these cells causes significant disruption of their differentiation program. Our experiments showed that squamous differentiation occurs in normal primary RTE cell cultures. Epidermal transglutaminase (transglutaminase type I) activity increased approximately 20-fold in RTE cultures as a function of time. Cholesterol sulfate, another marker of squamous differentiation, increased only modestly with time. A significant number of cells formed cross-linked envelopes in cultures growth-arrested at a cell density of approximately 250 cells/mm2. However, no significant changes in keratin expression were detected. Neoplastically transformed RTE cells which exhibit a greatly increased growth capacity expressed the same three markers of squamous differentiation as normal RTE cells. However, transglutaminase type I activity was relatively low. The cross-linked envelope formation was independent of cell density in the transformed cells. Like in normal RTE cultures, cholesterol sulfate accumulation only increased moderately with increasing cell density. The keratin pattern of transformed RTE cell lines was identical to that of normal primary RTE cells. A well-differentiated squamous cell carcinoma derived from one of the neoplastic cell lines expressed in vivo keratin markers typical of keratinization (56 kd acidic keratin and 65-67 kd basic keratins). We draw the following conclusions. (i) The biochemical studies confirm that normal RTE cells undergo squamous differentiation. The pathway of terminal squamous differentiation is cell density dependent. (ii) In transformed RTE cells, growth as well as differentiation are less subject to regulation by cell density than in normal cells. (iii) Transformed RTE cells are differentiation competent; the main abnormality appears to be that in transformed cell populations proliferation and differentiation occur concomitantly.

Topics: Animals; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Cholesterol Esters; Epithelial Cells; Keratins; Male; Rats; Rats, Inbred F344; Trachea; Transglutaminases

Cultured human epidermal keratinocytes express a transglutaminase which appears in epidermis only during later stages of normal cell differentiation (Thacher and Rice, Cell 40:685, 1985). Using a monoclonal antibody affinity column, the keratinocyte transglutaminase has been purified approximately 1000-fold from non-ionic detergent extracts of the particulate fraction of a squamous cell carcinoma line. The prominent 92-kilodalton band in the purified material is recognized on Western Blots by a monoclonal antibody which also can immunoprecipitate the active enzyme. Further analysis of the 92-kilodalton band shows that it is apparently not antigenically related to tissue transglutaminase, or to the cytoplasmic enzyme expressed in cultured keratinocytes which is immunologically cross-reactive and of identical molecular weight to tissue transglutaminase. In monkey palmar and plantar epidermal homogenates significant transglutaminase activity appears to be membrane bound and chromatographs on anion-exchange as the keratinocyte enzyme does. It can be precipitated, at least in part, with the aid of specific monoclonal antibodies directed to keratinocyte transglutaminase. As determined immunohistochemically, the most likely membrane location of the enzyme in cultured cells is the plasma membrane, consistent with keratinocyte transglutaminase function in cross-linked envelope formation. Thigh, neonatal foreskin, psoriatic epidermis, and monkey esophagus are compared as to patterns of staining for keratinocyte transglutaminase. Cultured epidermal cells and psoriatic epidermis both show precocious expression when compared to normal epidermis, demonstrating that keratinocyte transglutaminase expression can be substantially modified during epidermal regenerative maturation or hyperproliferation.

Topics: Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Erythrocytes; Humans; Immunohistochemistry; Keratins; Transglutaminases; Tumor Cells, Cultured

The immunocytochemical method with monoclonal antibodies to human and murine epidermis basal-cell antigen (BCAg) has been used in studies on the disorders in keratinocyte differentiation in epidermal tumors and mycosis fungoides. The fluorescence intensity augmented and BCAg distribution in the cytoplasm grew more diffuse as the degree of the tumor cell differentiation lowered. In mycosis fungoides the BCAg reaction has been positive first in the suprabasal layers; as the disease progressed, the fluorescence has involved the entire epidermis. The BCAg test is a sensitive tool for estimating the epidermal cell maturity in processes associated with impaired differentiation of keratinocytes.

Topics: Antibodies, Monoclonal; Antigens, Differentiation; Antigens, Neoplasm; Biopsy; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epidermis; Humans; Immunohistochemistry; Keratins; Mycosis Fungoides; Skin Neoplasms

The mouse monoclonal antibody OSU 22-3 was prepared using cells from a squamous cell carcinoma (SCC) as an immunogen. This antibody reacts with an antigen found on squamous cell carcinomas but does not react with normal keratinocytes. This antibody and two antibodies that react with normal keratinocytes were used as markers of malignant and normal phenotypes. These markers were used to evaluate several spontaneous and carcinogen initiated SCC tumors and to identify the expression of an antigen associated with a malignant phenotype. A variety of subpopulations in carcinogen initiated tumors and spontaneous SCC tumors were noted. The subpopulations that reacted only with MoAb OSU 22-3 exhibited features of anchorage independent growth and cellular invasiveness, and formed progressively growing tumors in nude mice. Other SCC spontaneous tumor cell subpopulations reacted with the antibodies associated with normal keratinocytes. These cells did not proliferate in vitro and did not form tumors in the nude mouse. There were other carcinogen transformed cells which reacted with MoAb OSU 22-3 but not with the antibodies associated with normal keratinocytes. These cells exhibited anchorage independent growth and cellular invasiveness but did not form tumors in nude mice. We conclude from this work that human SCC tumors contain multiple cell populations. These cell populations have varied growth properties and express surface antigens that may indicate their malignant vigor. Carcinogen transformed keratinocytes do exhibit some of the characteristics of SCC tumor phenotypes but not the property of malignant progressively growing cells on a routine and consistent basis. This feature is transiently and inconsistently expressed in a surrogate host by populations prepared from spontaneous SSC tumors.

Topics: Animals; Antibodies, Monoclonal; Carcinogens; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Dose-Response Relationship, Drug; Epidermal Cells; Epidermis; Humans; Keratins; Lethal Dose 50; Male; Methylnitronitrosoguanidine; Mice; Mice, Nude; Neoplasm Staging; Phenotype; Skin Neoplasms; Tumor Cells, Cultured

Tumors transplanted into nude mice using the TTK-1 cell lines [TTK-1(E) and TTK-1(F)] derived from normal human early decidual tissue were studied morphologically. The epithelial-like cell line TTK-1(E) and the fibroblast-like cell line TTK-1(F) were maintained in culture through one hundred and ten subcultures since July 1979. Rapidly growing tumor nodules formed at the implantation sites. The incidence of tumor growth was 100% for both cell lines. Histologically the tumors were composed of poorly-differentiated cells arranged in a cord-like structure and showed typical malignant characteristics. Immunohistochemical studies, electron microscopy and immunocytochemical studies revealed that the tumors from the two cell lines differed in many respects. The tumors formed by TTK-1(E) showed epithelial characteristics and the tumors formed by TTK-1(F) showed both epithelial and mesenchymal characteristics. Therefore, TTK-1(E) might be useful as an in vitro model of endometrial cancer and TTK-1(F) as an in vitro model of both endometrial cancer and endometrial stromal tumor (containing mixed mesodermal tumor). These tumors will be valuable for future studies of the tumorigenicity and therapy of uterine malignant tumors. They may reflect the various functions of decidual tissue.

Topics: Animals; Cell Line; Cell Transformation, Neoplastic; Decidua; Epithelium; Female; Fibroblasts; Humans; Immunoenzyme Techniques; Keratins; Mesoderm; Mice; Mice, Inbred ICR; Mice, Nude; Models, Biological; Neoplasm Transplantation; Neoplasms, Experimental; Pregnancy; Uterine Neoplasms; Vimentin

Sodium butyrate affects cell differentiation in confluent epidermal keratinocyte cultures by considerably increasing the spontaneous formation of cross-linked envelopes in normal human keratinocytes (NHK). It also favors the development of envelope competence in the Simian virus-40 (SV-40)-transformed human foreskin keratinocyte line SV-K14. It completely abolishes the inhibitory effect of serum and retinoic acid on the expression of plasma membrane-associated transglutaminase. However, other markers of epidermal differentiation that are also under the control of retinoids such as keratins or the enzyme cholesterol sulfotransferase are not affected by butyrate. The level of the cellular retinoic acid binding protein (CRABP) is considerably increased in its presence. Butyrate does not interfere with the binding of retinoids to their cellular binding proteins. Our observations suggest that sodium butyrate stimulates cornified envelope formation via the induction of the plasma membrane-associated transglutaminase required for cornified envelope synthesis and, additionally, by abolishing the inhibitory effect of retinoids on the expression of this enzyme.

Topics: Butyrates; Butyric Acid; Carrier Proteins; Cell Line, Transformed; Cell Membrane; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Gene Expression Regulation; Humans; Keratins; Male; Receptors, Retinoic Acid; Retinoids; Serum Albumin, Bovine; Simian virus 40; Sulfotransferases; Transglutaminases

Stereological techniques have been used to study the vascularity in 110 sublingual keratosis lesions and 22 specimens of normal sublingual mucosa using an image analysis system "IBAS 1". The stereological parameters volume density (Vv), and surface density (Sv) were significantly increased in sublingual keratosis lesions as compared with those of normal sublingual mucosa. Lesions that eventually underwent malignant change showed the highest values and were highly significant (p less than .001) when compared with those which did not do so. The present results demonstrate that vascularity could be a supportive marker of impending malignant change in sublingual keratosis lesions.

Topics: Blood Vessels; Cell Transformation, Neoplastic; Female; Humans; Keratins; Leukoplakia, Oral; Male; Mouth Floor; Mouth Mucosa; Statistics as Topic

Topics: Abdominal Neoplasms; Adolescent; Adult; Antibodies, Monoclonal; Cell Transformation, Neoplastic; Child; Desmin; Diagnosis, Differential; Female; Fibroma; Humans; Keratins; Male; Neoplasms, Germ Cell and Embryonal

We have described clinical, histological immunohistochemical and ultrastructural features of four typical examples of epithelioid sarcoma. Smooth-muscle-type focal densities amid fine actin-like filaments in typical epithelioid cells often containing prominent rER have been observed, as well as a structure in the form of a densely granular cytoplasmic body. These features are suggestive of myofibroblastic differentiation. A tumour with light microscopy features typical of epithelioid sarcoma but of spindle-cell appearance is also described. Ultrastructurally this consists of myofibroblasts and it is suggested that this may be an unusual variant in which myofibroblastic differentiation is a major feature.

Topics: Adult; Aged; Cell Transformation, Neoplastic; Female; Fibroblasts; Humans; Immunohistochemistry; Keratins; Male; Microscopy, Electron; Middle Aged; Muscle, Smooth; Sarcoma; Skin Neoplasms; Vimentin

Differentiation in keratinocytes can be experimentally modulated by changing the culture conditions. When cultured under conventional, submerged conditions, the extent of cellular differentiation is reduced in the presence of low calcium medium and is enhanced in medium containing physiologic calcium concentrations. Moreover, cultures grown at the air-medium interface or on a dermal substrate, or both, differentiate even further. Herein we report the effect of culture conditions on lipid composition in normal human keratinocytes and three squamous carcinoma cell (SCC) lines that vary in their capacity to differentiate as assessed by cornified envelope formation. Under submerged conditions, the total phospholipid content was lower, triglyceride content higher, and phospholipid:neutral lipid ratio lower in direct correlation to the degree of differentiation in these cultures. When grown at the air-medium interface on de-epidermized dermis, evidence of further morphologic differentiation was found only for well-differentiated SCC cells and normal keratinocytes. Similarly, the phospholipid content remained high in poorly differentiated SCC cells and it decreased modestly in well-differentiated SCC cells and markedly in normal keratinocytes. In all cell lines the triglyceride content was increased and cholesterol content decreased when compared to parallel submerged cultures, but these differences were most pronounced in well-differentiated cell lines. Acylceramides and acylglucosylceramides were found only in normal keratinocytes and only under the most differentiation-enhancing conditions. These studies demonstrate differentiation-related changes in the lipid content of both normal and neoplastic keratinocytes.

Topics: Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Humans; Infant; Keratins; Lipids; Male; Skin Neoplasms; Tumor Cells, Cultured

Eighteen cases of primary thymic carcinoma were reviewed from the viewpoint of glandular differentiation. Squamous differentiation was evident in 14 cases (83%). Immunohistochemical study revealed secretory component (SC)-positive carcinoma cells in 12 cases (67%), most of which were also associated with squamous differentiation. Three of these 12 cases contained areas with a definite glandular or microcystic structure with occasional epithelial mucin, and were diagnosed as adenosquamous carcinoma. Review of patients' medical records revealed that thymic carcinomas with a glandular element were more often resectable at surgery, and had a much better prognosis than those without a glandular element. However, further study on larger number of cases is necessary to confirm this relationship. Because SC-positive epithelial cells do exist in the non-neoplastic thymus, the presence of a glandular component suggests another direction of morphological and/or functional differentiation of thymic carcinoma cells in addition to the well-known squamous differentiation.

Topics: Biomarkers, Tumor; Carcinoma; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Humans; Immunohistochemistry; Keratins; Prognosis; Thymus Gland; Thymus Neoplasms

Simian Virus 40 (SV40) transformation of primary cultures of human mammary epithelial cells has yielded a cloned epithelial-like cell line and a representative, single-cell subclone. Although apparently homogeneous, both cloned cell lines can also yield small numbers of three other cell types. The more-elongated cell type can be obtained directly by replating cells from the medium of the epithelial-like cell cultures or by picking and culturing single cells to form representative lines. Immunofluorescent and immunocytochemical analysis of these cell lines growing on plastic or as tumor-nodules in nude mice for epithelial membrane antigens, various cytokeratins, various actins, laminin, Type IV collagen, the common acute lymphoblastic leukemia antigen (CALLA), and a 135-kDa glycoprotein confirm the epithelial nature of the epithelial-like cells and suggest a myoepithelial origin for the more-elongated cell type. Ultrastructural analysis largely confirms the results, although the myofilamental bundles can be scanty in the growing myoepithelial-like cells. The other two cell types are possibly related to the keratinizing and casein-secreting cells seen in the epithelial tumor-nodules before and after mating the mice, respectively. The myoepithelial-like cells produce 5- to 17-fold more laminin, Type IV collagen, CALLA, and the 135-kDa glycoprotein than the epithelial cells, and all of these antigens are preferentially found on myoepithelial cells in vivo. It is suggested that the SV40-transformed epithelial cell is an immortalized form of human mammary stem cell which can differentiate in culture and in vivo to myoepithelial-like cells.

Topics: Actins; Animals; Antibodies, Monoclonal; Antigens, Differentiation; Antigens, Neoplasm; Antigens, Surface; Breast; Cell Differentiation; Cell Line, Transformed; Cell Transformation, Neoplastic; Cell Transformation, Viral; Collagen; Epithelial Cells; Epithelium; Fluorescent Antibody Technique; Humans; Immunohistochemistry; Keratins; Laminin; Membrane Glycoproteins; Mice; Mice, Nude; Mucin-1; Neoplasm Transplantation; Neoplasms, Experimental; Neprilysin; Simian virus 40; Stem Cells

Two monoclonal antibodies (MAb) specific for differentiation-related epidermal keratins have been developed. They represent specific molecular probes for different stages of epidermal differentiation. Antibody DE-K10 is chain-specific for cytokeratin polypeptide no. 10 (56.5 kD) expressed in all suprabasal layers of the epidermis. Antibody DE-SCK is specific for modified stratum corneum keratins and thus represents a marker for the terminal step of epidermal differentiation. Since the epitopes identified by both antibodies are preserved in formalin-fixed, paraffin-embedded tissue sections, these antibodies can be used for retrospective studies of differentiation in various pathological processes. We have used antibody DE-K10 to study the cytokeratin 10 expression in 26 stage II or III vulvar squamous cell carcinomas. Preliminary data suggest an increased risk of recurrence in cytokeratin 10 negative tumours.

Topics: Antibodies, Monoclonal; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epidermis; Female; Humans; Immunoblotting; Keratins; Middle Aged; Neoplasm Recurrence, Local; Vulvar Neoplasms

The oncogenic potential of human papillomavirus type 18 which is found in a significant number of cervical and penile cancer biopsies was tested in primary human keratinocytes derived from neonatal foreskin. Viral DNA and a gene for resistance to neomycin were introduced into these cells by calcium phosphate transfection. Selection of cells in G418 led to the isolation of resistant colonies which were propagated in culture. Four cell lines termed FE-A, FEH 18L, FEP18-5, and FEP18-11 have been maintained in culture for 1 1/2-2 years and were selected for further analysis. In all cases the viral DNA was integrated into the cellular genome and the early genes were transcribed, including RNA complementary to the E2, E6, and E7 open reading frames. Radioimmunoprecipitation showed that all cell lines synthesized the E6 and E7 proteins. However, none of the cell lines tested were tumorigenic. The differentiation capacity of these cells was analyzed by assessing their ability to proliferate clonally after exposure to 1.2 mM calcium chloride. All four cell lines were resistant to this stimulus and formed colonies upon return to regular growth medium whereas normal cells differentiated terminally. K6a and K14 keratin RNA expression was down-regulated in the HPV immortalized cell lines compared to primary human epithelial cells.

Topics: Blotting, Northern; Blotting, Southern; Cell Line; Cell Line, Transformed; Cell Transformation, Neoplastic; Cell Transformation, Viral; DNA, Viral; Humans; Keratinocytes; Keratins; Papillomaviridae; Radioimmunoprecipitation Assay; RNA, Viral; Transcription, Genetic; Transfection; Viral Proteins

We studied 23 cases of capillary hemangioblastoma (CHB) of the cerebellum with 17 immunohistochemical cell markers in an attempt to define the nature of the so-called "stromal" cells. These cases were compared to four cases of intracranial metastatic renal cell carcinoma (RCC), which may mimic CHB histologically. The 17 markers studied included vimentin (VIM), Factor VIII-related antigen (FVIIIR:Ag), blood group antigens A, B, and H, Ulex I lectin (Ulex), Alkaline Phosphatase (Alk P), neurofilament protein (NF), glial fibrillary acidic protein (GFAP), S-100 protein (S-100), nerve growth factor receptor (NGFR), muscle-specific actin (MSA), desmin (Des), monoclonal keratin (MKER, including Cam 5.2 and AE1/3), epithelial membrane antigen (EMA), and chromogranin (Chrom). No significant stromal cell staining was seen by markers for endothelial, epithelial, chromaffin, or smooth muscle origin. In some cases individual cells demonstrated positivity for GFAP (4/22) and S-100 protein (13/23); these cells were generally stellate and located near the periphery, and we conclude that these were the result of entrapment of surrounding cerebellum. No case demonstrated NF in stromal cells. However, nearly all cases of CHB showed stromal cell staining with VIM (19/22). In contrast, all of the cases of RCC showed significant staining for at least one marker of epithelial origin (3/4 for MKER and 4/4 for EMA). We conclude that the stromal cell of CHB is neither endothelial, neural, epithelial, pericytic, nor neuroendocrine in origin, and is instead of undifferentiated mesenchymal origin. The designation of this tumor as an "hemangioblastoma," although a misnomer, is firmly established in the literature and should probably be retained.

Topics: Actins; Alkaline Phosphatase; Cell Transformation, Neoplastic; Cerebellar Neoplasms; Chromogranins; Desmin; Diagnosis, Differential; Glial Fibrillary Acidic Protein; Hemangiosarcoma; Humans; Immunohistochemistry; Intermediate Filament Proteins; Isoantigens; Keratins; Lectins; Membrane Glycoproteins; Mesenchymoma; Mucin-1; Neoplasm Metastasis; Plant Lectins; Receptors, Cell Surface; Receptors, Nerve Growth Factor; S100 Proteins; Vimentin; von Hippel-Lindau Disease; von Willebrand Factor

A parasagittal meningioma of an eighty year old female patient showed by light and electron microscopy cystic architecture (forme humide) as well as nuclear vacuoles (indentations) and cytoplasmic inclusions. The latter are the known pseudopsammoma bodies or hyaline inclusions as demonstrated by light and electron microscopy. Light microcopy on paraffin sections and cytological smear preparations revealed, in addition to the cells of endotheliomatous meningioma and those containing the inclusions a third type with small granular cytoplasmic content. Electron microscopy showed characteristic features of meningioma such as folded double membranes, desmosomes and filaments and thus gave evidence of the meningiomatous nature of the tumor. By immunohistochemistry tumor cells in slightly focal distribution contained vimentin, whereas small clusters of cells with hyaline inclusions were strongly positive for cytokeratin. The dispersed cells of granular cytoplasmic content were positive for fibronectin. These findings, especially of the inclusion containing cytokeratin positive cell clusters may shed new light upon the concept of histogenesis and classification.

Topics: Aged; Aged, 80 and over; Biomarkers, Tumor; Cell Transformation, Neoplastic; Female; Fibronectins; Humans; Immunohistochemistry; Keratins; Meningeal Neoplasms; Meningioma; Microscopy, Electron; Vimentin

An infiltrating epithelial and spindle cell neoplasm developed in the breast of a 63-year-old female. An excisional biopsy was performed. Recurrence with rapid growth due to cyst development eventually resulted in more radical surgery. Interim fine needle aspirations had established its partially cystic nature. The unique microscopic appearance prompted the application of immunohistochemistry and electron microscopy. The tumour cells were found to exhibit characteristics denoting squamous and myoepithelial differentiation. Histopathological features of malignancy were absent. Our findings demonstrate the differentiation potential of breast epithelium. They are in concordance with the results of previous studies which delineate the histochemical and ultrastructural features of myoepithelial and establish the relationship of these cells to squamous metaplasia.

Topics: Actins; Breast Neoplasms; Carcinoma; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Keratins; Membrane Glycoproteins; Microscopy, Electron; Middle Aged; Mucin-1; Myoepithelioma; Vimentin

Rat liver T51B cells were maintained in the presence of low concentrations of Ni(II) derived from alpha Ni3S2 for 3-15 months in culture in order to monitor cytokeratin, differentiation, and transformation patterns. Nickel exposures caused irreversible, heritable juxtanuclear aggregates of cytokeratin CK55, which increased in size and complexity with prolonged nickel exposure, eventually resembling Mallory bodies and expressing glutamyltransferase. Altered cytokeratin expression was accompanied by induction of differentiation, with markers of both bile ductular cells and hepatocytes, such as induction of cytokeratin polypeptides CK39 and CK49, cell morphology, and cytokeratin filament network changes; whereas control cultures similarly maintained for long periods in culture remained unchanged. Altered cytokeratin expression was also accompanied by acquisition of transformation markers--loss of density dependence, progression toward calcium independence, and (benign) growth in nude mice. Observed cytokeratin aberrations may be a factor in nickel carcinogenesis, in view of the known affinity of the metal for cellular structural proteins, especially keratin, which play a role in maintenance of cell behavior.

Topics: Animals; Antibodies, Monoclonal; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Cytoskeleton; Electrophoresis, Polyacrylamide Gel; Epithelial Cells; Gene Expression; Keratins; Liver; Nickel; Rats; Spectrometry, Fluorescence

SVK14, an SV40-transformed human keratinocyte line, has previously been reported to be almost completely unable to differentiate, and indeed, to express a set of keratins characteristic of simple epithelia rather than the stratifying epithelium from which they were derived. We have recently shown that IGF I stimulation of SVK14 results in expression of keratin 14, a marker of stratifying epithelia, as well as expression of markers which are characteristic of differentiation in normal human keratinocytes such as involucrin and keratin 10. To study further the capacity of SVK14 to differentiate, we have cocultured SVK14 with a variety of fibroblastic cell lines with a view to examining whether the cocultured partner can promote or interfere with their differentiation. We have observed that SVK14, when cocultured with Swiss 3T3, form organized structures through specific cell-cell interactions in which SVK14 express keratins 14 and 5 and involucrin, while maintaining T-antigen expression. These results are interesting since they show coculturing of a transformed human keratinocyte cell line and a particular fibroblast line can result in induction of characteristics of stratifying epithelia in a cell line with characteristics of simple epithelia. This may be analogous to the epithelial-mesenchymal interactions seen during epithelial development in the very early embryo.

Topics: Animals; Antibodies, Monoclonal; Biomarkers; Cell Communication; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Fluorescent Antibody Technique; Humans; Keratinocytes; Keratins; Mice; Mitomycin; Mitomycins; Simian virus 40

Acetone-fixed frozen sections of 15 malignant melanomas of the skin with metastases were studied immunohistochemically for the presence of different types of intermediate filament proteins, synaptophysin, muscle cell actins, and desmoplakins. One of the melanomas was a primary toe tumor, and the others mainly regional lymph node metastases. The original diagnosis of melanoma was reconfirmed in each case, and the melanoma diagnosis of the metastases was verified by S100 protein immunostaining in all cases and by a monoclonal antibody to melanoma cells (NK1C3) in 7 cases. All melanomas were prominently vimentin-positive. In 10 of 15 cases, immunoreactive keratin could be demonstrated with antibody CAM 5.2. The presence of keratins was confirmed in selected cases with three other monoclonal antibodies including AE1, PKK1, and a monoclonal antibody specific for keratin number 18. Desmoplakin, another marker of epithelial differentiation, was not found in melanoma cells. Two melanomas contained neurofilament-positive tumor cells, which were however negative for synaptophysin. Desmin, muscle actins, and glial fibrillary acidic protein were not found in the neoplastic cells. On the basis of the present results one could conclude that the protein composition of the cytoskeleton of melanomas is more complex than has been previously thought and most importantly that melanomas may contain keratins.

Topics: Adult; Aged; Cell Transformation, Neoplastic; Female; Humans; Immunohistochemistry; Intermediate Filaments; Keratins; Male; Melanoma; Middle Aged; Skin Neoplasms

The epithelial properties of the cells in six cases of synovial sarcoma, five biphasic and one monophasic, were examined. Epithelial cells formed well demarcated glandular nests with a multi- or single-layered lining, poorly defined nest-like clusters, or appeared as scattered single plump cells in a spindle-cell stroma. Keratinization was seen in multi-layered epithelia, and was suggested in single-layered epithelia from the presence of some enlarged eosinophilic cells. Immunohistochemically, epithelia lining glands showed reactivity for both low- and high-molecular-weight (MW) cytokeratins, and the reactions for both were strongest in keratinized cells. Clustered or single plump cells showed low-MW cytokeratin reactivity. All epithelial cells were negative for the largest cytokeratin (no. 1), and occasionally stained positively for vimentin. Spindle-shaped stromal cells usually stained positively for vimentin only. Type IV collagen was distributed linearly around well circumscribed epithelia, but not around poorly defined epithelia or plump cells. Epithelial membrane antigen was distributed linearly along glandular spaces and irregularly or granularly in clustered plump cells. The immunophenotypic epithelialization in synovial sarcoma was restricted, but showed considerable correlation with the histological grade of differentiation.

Topics: Adult; Aged; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; Collagen; Desmin; Epithelium; Female; Humans; Keratins; Male; Membrane Glycoproteins; Middle Aged; Mucin-1; Phenotype; S100 Proteins; Sarcoma, Synovial; Skin Neoplasms; Vimentin

The epithelia lining the cyst of five cases of calcifying odontogenic cyst (COC) were evaluated immunohistochemically with the use of monoclonal antibodies (MoAb's) against keratin (PKK1, KL1, K4.62, K8.12) and vimentin, and polyclonal antisera agonist involucrin and filaggrin. Epithelial lining of COC was classified into 1) thin squamous-cell epithelium, 2) ameloblastoma-like, and 3) thin or 4) thick calcifying odontogenic epithelium. Foci consisting of ghost cells or calcified cells were categorized as calcifying epithelial odontogenic tumor (CEOT). Thin squamous-cell epithelium reacted with PKK1, KL1, K4.62, K8.12, and anti-vimentin MoAb's, thus demonstrating the co-expression of keratin and vimentin. Ameloblastoma-like cells showed positive staining with PKK1, KL1, and sometimes with anti-vimentin. Thick calcifying odontogenic epithelial lining showed stratification of cell layers, and the most strikingly reactive zone was the upper intermediate layer, which showed the presence of keratin, involucrin, and a small amount of filaggrin. Cells of this layer might be the most differentiated type of cells in COC. Undifferentiated odontogenic cells of COC masses were characterized by co-expression of keratin and vimentin, and by the absence of involucrin and filaggrin. All ghost cells were devoid of any immunostaining except for filaggrin, which was rarely positive, but eosinophilic or basophilic cells surrounding the ghost cells showed intense staining for all keratin proteins except vimentin.

Topics: Adolescent; Adult; Cell Transformation, Neoplastic; Epithelium; Female; Filaggrin Proteins; Humans; Immunohistochemistry; Intermediate Filament Proteins; Jaw Diseases; Keratins; Male; Odontogenic Tumors; Protein Precursors

Oval cells emerging in rat liver at the early period of 3-methyl-4-dimethylaminoazobenzene treatment constitute a mixed epithelial cell compartment with respect to alpha-fetoprotein (AFP) and cytokeratin differential expression, and include a subpopulation which exhibits a phenotype intermediate between ductular cells and hepatocytes (Germain et al., Cancer Res., 45:673-681, 1985). In the present study we have examined the developmental potential of ductular oval cells in primary culture and after in vivo transfer. The use of monoclonal and polyclonal antibodies directed against cytokeratins of Mr 39,000 (CK39), 52,000 (CK52), and 55,000 (CK55) and vimentin, and also monoclonal antibodies against exposed surface components of oval cells (BDS7) and normal hepatocytes (HES6) allowed us to establish the ductular phenotype of the oval cells. A highly enriched preparation of oval cells was obtained by perfusion/digestion of the liver with collagenase, treatment of the cell suspension with trypsin and DNase, selective removal of hepatocytes by panning using the anti-HES6 antibody, and cell separation by isopyknic centrifugation in a Percoll gradient. The procedure yielded about 8 x 10(7) cells, of which 95% expressed CK39, CK52, and BDS7, 84% gamma-glutamyl transpeptidase, and 5% albumin and AFP. The primary response of cultured oval cells to various combinations of growth and differentiation promoting factors was evaluated with respect to their capacity to initiate DNA synthesis as measured by [3H]thymidine labeling from day 1 to 3, and/or to produce albumin and AFP and express tyrosine aminotransferase. Culture in the presence of either serum or clot blood extract resulted in a low proliferative activity with less than 5% of the nuclei being labeled. Over a 5-day period, fusion of a large portion of the oval cells led to multinucleated cells. When the cells were cultured in the presence of an elaborate combination of supplements [minimum essential medium containing 1 mM pyruvate, 0.2 mM aspartate, 0.2 mM serine, 1 mM tyrosine, 1 mM proline, 1 mM phenylalanine and supplemented with 20% clot blood extract, 10 ng/ml oxidized bile acids, 17 microM bilirubin, 10 ng/ml cholera toxin, 1 microM dexamethasone, 2.5 micrograms/ml insulin, 50 mM beta-mercaptoethanol, and 5 micrograms/ml transferrin (medium MX)], the labeling index increased to around 30% and the level of cell fusion greatly decreased. The addition of dimethyl sulfoxide further enhanced the initiation of

Topics: Animals; Cell Separation; Cell Transformation, Neoplastic; Dimethyl Sulfoxide; gamma-Glutamyltransferase; Keratins; Liver; Male; Methyldimethylaminoazobenzene; Phenotype; Rats; Rats, Inbred F344; Tumor Cells, Cultured; Vimentin

In this study we have examined possible differentiation-dependent modulations in plasma membrane lipid properties in normal keratinocytes, SV-40 transformed keratinocytes (SVK14) and a number of squamous carcinoma (SCC) cells. In normal keratinocytes the lateral diffusion coefficient of plasma membrane lipids (D) differs significantly for cells cultured permanently under low and normal Ca2+-conditions (5.16 x 10(-9) and 3.27 x 10(-9) cm2/s, respectively). When differentiation is induced by exposing low Ca2+-cultured cells to normal Ca2+ concentrations D increases to 7.07 x 10(-9) cm2/s during the initial hours of differentiation followed by a gradual sustained decrease to values also observed in cells cultured permanently under normal Ca2+-conditions. In SCC and SVK14 cells a similar initial transient increase in lateral lipid mobility is observed upon initiation of differentiation, but, in contrast to normal keratinocytes, no sustained decrease in D is seen upon prolonged culturing under normal Ca2+ conditions. The results indicate that the deficiency of the transformed cells to respond to Ca2+-induced differentiation might involve transformation-dependent alterations in membrane structure and function.

Topics: Calcium; Carcinoma, Squamous Cell; Cell Differentiation; Cell Membrane; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cells, Cultured; Culture Media; Epidermal Cells; Epidermis; Humans; Keratins; Lipid Bilayers; Membrane Lipids

The cheek pouch of the Syrian hamster is an excellent tissue for the experimental induction of oral cancer by carcinogenic chemicals. Lysate prepared from a cell line (HCPC-1) derived from one of these hamster oral tumors greatly increased the growth of these oral tumor cells in vitro. We now show that the mitogenic substance, transforming growth factor alpha (TGF-alpha), is present in all of the chemically transformed hamster oral tumors examined (in vitro and in vivo). In no adult normal tissue of the Syrian hamster can we detect expression of TGF-alpha. TGF-alpha could be partly or wholly responsible for the mitogenic activity detected in the lysate of the chemically transformed hamster oral keratinocytes. Both normal and chemically transformed hamster oral keratinocytes express the receptor to epidermal growth factor. The consistent detection of TGF-alpha and epidermal growth factor receptor mRNAs in these hamster oral tumor cells suggests that an autocrine growth mechanism might be operative. This hamster cheek pouch oral cancer model can be used for the molecular analysis of how TGF-alpha and epidermal growth factor receptor might be involved in the malignant transformation of epithelial tissues.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Cloning, Molecular; Cricetinae; DNA Replication; Epidermal Cells; Epidermal Growth Factor; Epidermis; ErbB Receptors; Growth Substances; Keratins; Mouth Neoplasms; Peptides; Transforming Growth Factors

Monospecific antikeratin antisera and specific complementary DNA probes were used to analyze expression of keratin genes in newborn mouse skin and skin papillomas and carcinomas by indirect immunofluorescence, immunoblotting, and in situ hybridization. Tumors were induced by initiation with 7,12-dimethylbenz[a]anthracene and promotion with 12-O-tetradecanoylphorbol-13-acetate. Type I epidermal keratin K14 protein (Mr 55,000) is found in all living layers of the newborn skin but is most abundant in the lower strata. K1 (Mr 67,000) and K10 (Mr 59,000) proteins are predominantly suprabasal and K1 is processed in the stratum corneum. Transcripts for K14 were confined largely to the basal cell layer by in situ hybridization. Transcripts for K1 and K10 were highly expressed in suprabasal cells including the granular cell layer. In benign tumors, distribution of K14 protein is similar to that in newborn skin, while the abundance of K1 and K10 appears to be somewhat reduced although the tissue distribution remains suprabasal. Transcription of K14 is aberrant in benign tumors and transcripts persist throughout much of the suprabasal cell layers. Transcripts of K1 and K10 are normally distributed in papillomas but grain density is less intense than in newborn epidermis. Keratin expression in carcinomas is highly disturbed. K14 protein and transcripts are highly expressed in all strata in carcinomas while protein and transcripts for K1 and K10 are essentially absent. These results suggest that papilloma cells fail to respond to or generate signals to regulate K14 expression in the differentiating suprabasal cell layers and may not fully express their suprabasal cell keratins. Carcinomas fail to express suprabasal cell keratins and this is regulated at the transcriptional level. The loss of suprabasal keratin expression may provide a marker for malignant conversion in the mouse skin carcinogenesis model.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Animals, Newborn; Carcinoma; Cell Transformation, Neoplastic; Female; Gene Expression Regulation; Genes; Keratins; Mice; Mice, Inbred Strains; Nucleic Acid Hybridization; Papilloma; Skin; Skin Neoplasms; Tetradecanoylphorbol Acetate; Transcription, Genetic

Dedifferentiated chondrosarcoma is a biphasic tumour, comprising well-differentiated chondrosarcoma and an anaplastic non-cartilaginous sarcoma juxtaposed but distinct from each other. Two cases of dedifferentiated chondrosarcoma, one primary and one recurrent, demonstrated muscle differentiation when studied with monoclonal antibodies to muscle specific actin, desmin and myoglobin. One of the tumours was also positive for cytokeratin, identified by AE1/AE3 and CAM 5.2 antibodies. Our findings are consistent with the concept that these tumours are capable of diverse patterns of morphological and immunophenotypic differentiation.

Topics: Adult; Anaplasia; Bone Neoplasms; Cell Transformation, Neoplastic; Chondrosarcoma; Female; Humans; Immunohistochemistry; Keratins; Middle Aged; Muscles; Pelvic Bones; Ribs

Ten examples of giant cell carcinoma of the lung were examined by immunohistochemistry for expression of keratin and vimentin intermediate filaments and for epithelial membrane antigen (EMA). Six cases were also examined electron microscopically. Keratin expression and, to a lesser extent, EMA immunoreactivity were reduced in comparison with better differentiated forms of lung carcinoma. Vimentin expression was increased, often taking the form of strong paranuclear staining. This may correspond to dense paranuclear aggregates of intermediate filaments seen ultrastructurally. Desmosomes were absent or sparse in most tumours. We propose that giant cell carcinoma arises by a process of dedifferentiation. The resulting loss of epithelial features gives rise to neoplastic cells which have features in common with some forms of sarcoma.

Topics: Adenocarcinoma; Antigens; Carcinoma; Cell Differentiation; Cell Transformation, Neoplastic; Desmosomes; Humans; Keratins; Lung Neoplasms; Membrane Glycoproteins; Microscopy, Electron; Microvilli; Mucin-1; Vimentin

To facilitate understanding of the mechanisms underlying pulmonary diseases, including lung cancer and cystic fibrosis, we have transformed and characterized cultures of human tracheal epithelial cells. Cells were transfected by calcium phosphate precipitation with a plasmid containing a replication-defective simian virus 40 (SV40) genome. Colonies of cells with enhanced growth potential were isolated and analyzed for transformation- and epithelial-specific characteristics. Precrisis cells were observed to express the SV40 large tumor antigen, produce cytokeratins, have microvilli, and form tight junctions. After crisis, cells continued to express the SV40 large tumor antigen as well as epithelial-specific cytokeratins and to display the apical membrane microvilli. Apical membrane Cl channels were opened in postcrisis cells exposed to 50 microM forskolin. These channels showed electrical properties similar to those observed in primary cultures. The postcrisis cells have been in culture for greater than 250 generations and are potentially "immortal." In addition to providing a useful in vitro model for the study of ion transport by human airway epithelial cells, the cells can be used to examine stages of neoplastic progression.

Topics: Cell Line, Transformed; Cell Transformation, Neoplastic; Cell Transformation, Viral; Chlorides; Defective Viruses; Epithelium; Humans; Ion Channels; Keratins; Oncogenes; Simian virus 40; Trachea

The activated ras oncogene that is present in Harvey sarcoma virus is able to induce malignant transformation of pure cultures of mouse primary keratinocytes. Malignant transformation of these cells is demonstrated by their ability to form carcinomas when grafted back onto syngeneic animals. However, expression of the malignant phenotype by the ras-transformed keratinocytes is drastically inhibited by the presence of normal dermal fibroblasts. This inhibitory effect depends on the ratio of fibroblasts to keratinocytes. It can be observed with mitomycin C-treated growth-arrested dermal fibroblasts and not with other cells, such as normal keratinocytes or established fibroblasts. Thus, a cellular environment approximating normal tissue can suppress tumor formation triggered by a single oncogene.

Topics: Animals; Animals, Newborn; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Fibroblasts; Genes, ras; Harvey murine sarcoma virus; Keratins; Mice; Mice, Inbred BALB C; Sarcoma Viruses, Murine; Skin

We report here the isolation and characterization of three antisera, each of which is specific for a single keratin from one of the three different pairs (K1/K10, K14/K5, K16/K6) that are differentially expressed in normal human epidermis and in epidermal diseases of hyperproliferation. We have used these antisera in conjunction with monospecific cRNA probes for epidermal keratin mRNAs to investigate pathways of differentiation in human epidermis and epidermal diseases in vivo and in epidermal cells cultured from normal skin and from squamous cell carcinomas in vitro. Specifically, our results suggest that: (a) the basal-specific keratin mRNAs are down-regulated upon commitment to terminal differentiation, but their encoded proteins are stable, and can be detected throughout the spinous layers; (b) the hyperproliferation-associated keratin mRNAs are expressed at a low level throughout normal epidermis when their encoded proteins are not expressed, but are synthesized at high levels in the suprabasal layers of hyperproliferating epidermis, coincident with the induced expression of the hyperproliferation-associated keratins in these cells; and (c) concomitantly with the induction of the hyperproliferation-associated keratins in the suprabasal layers of the epidermis is the down-regulation of the expression of the terminal differentiation-specific keratins. These data have important implications for our understanding of normal epidermal differentiation and the deviations from this process in the course of epidermal diseases of hyperproliferation.

Topics: Amino Acid Sequence; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Humans; Immune Sera; Immunoassay; Immunohistochemistry; Keratins; Molecular Sequence Data; Nucleic Acid Hybridization; Psoriasis; RNA, Messenger; Skin Neoplasms; Tumor Cells, Cultured

The hamster cheek pouch is an excellent target tissue for the experimental study of oral carcinogenesis. In the course of searching for molecular alterations during the malignant transformation process, the necessity for a molecular marker for cellular proliferation became apparent. In this report, we show that the cellular level of the histone H3 mRNA is valid as a molecular index of proliferation for cycling cell populations. H3 is known to be proliferation dependent for its expression in cultured animal cells. This study shows that H3 retains its cell-cycle-dependent expression in chemically transformed oral keratinocytes. The onset of H3 mRNA synthesis couples to the onset of DNA synthesis (S-phase). The cellular level of H3 mRNA therefore is proportional to the fraction of cells in the S-phase of the cell cycle. This conveniently allows us to correlate, in asynchronized cell populations, the expression of cellular genes to their proliferation rates. We demonstrate the usefulness of this proliferation marker by presenting data that different chemically induced oral carcinomas, but not normal cheek pouch tissues, contain readily detectable levels of c-Ki-ras proto-oncogene mRNA. Probing the same RNA blot to quantitate H3 mRNA levels allowed us to conclude that the high levels of c-Ki-ras mRNA in tumor tissues was likely due to the increased growth rate of the tumor tissues and not due to the deregulated expression of this cellular-proto-oncogene.

Topics: Animals; Blotting, Northern; Carcinoma, Squamous Cell; Cell Cycle; Cell Division; Cell Transformation, Neoplastic; Cricetinae; Epidermis; Gene Expression Regulation; Histones; Interphase; Keratins; Mouth Neoplasms; Proto-Oncogenes; RNA, Messenger; RNA, Neoplasm; Tumor Cells, Cultured

Normal rat oral keratinocytes have been transformed with the carcinogen 4-nitroquinoline N-oxide (4NQO) in vitro. The morphology, growth characteristics, ability to grow without anchorage and tumorigenicity in athymic mice was examined in 12 selected cell lines. Each of the lines could be assigned to one of two general groups. The first group of cell lines although showing some morphological signs of transformation and the ability to be subcultured beyond passage 15 were not anchorage independent or able to form tumours in athymic mice. The second group of cell lines showed distinct signs of morphological transformation, could be serially subcultured without 3T3 feeder cells, were anchorage independent and tumorigenic in athymic mice. Anchorage independence was more common at higher passages and with increased 4NQO treatment and correlated well with a decreased reliance on 3T3 feeder cell support. The anchorage-independent phenotype was closely associated with the ability to form tumours in athymic mice. This same sequence of phenotypic changes has been demonstrated in rat oral keratinocytes after 4NQO treatment in vivo indicating that during carcinogenesis, cell populations progress through the same stages whether proliferation occurs in vitro or in vivo. There is some evidence to suggest, however, that the time interval between stages may be altered when carcinogenesis takes place in vitro.

Topics: 4-Nitroquinoline-1-oxide; Animals; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Keratins; Male; Mouth Mucosa; Mouth Neoplasms; Nitroquinolines; Rats; Rats, Inbred Strains

The cellular heterogeneity of cutaneous tumors from nine patients with type 1 (von Recklinghausen's) neurofibromatosis was studied using several antigenic markers with special reference to focal heterotopic differentiation and interindividual variation. Furthermore, cells which actively express the genes for type I and III collagens and fibronectin, the major components of the abundant extracellular matrix of neurofibromas, were localized using in situ hybridizations. In eight of nine cases, the S-100 protein positive cells, i.e. Schwann-like cells, composed 60 to 80% of the total cell population. However, in one case, only about 40% of the cells were S-100 protein positive. The latter tumor was studied with respect to perineurial cell differentiation and stained with a mixture of two antibodies, directed against the S-100 protein and type IV collagen. In Schwann cells, the staining reaction for S-100 protein was observed in the nuclear region, whereas the staining reaction for type IV collagen was located peripherally, corresponding to the basement membrane zone covering the cells. The stromal cells which showed only the peripheral staining profile were considered to be neoplastic perineurial cells. Distinct structures with epithelial, endothelial, or smooth muscle cell differentiation were present within the benign tumors, as detected by immunostaining for cytokeratin, epithelial membrane antigen, factor VIII-related antigen and desmin, respectively. In situ hybridizations revealed a clearly detectable expression of type I procollagen genes in less than 10% and type III procollagen gene in less than 5% of the total cell population. Active synthesis of fibronectin was limited to the vascular walls, when examined by in situ hybridization, and antibodies to cellular fibronectin localized to the same areas. However, antibodies to plasma fibronectin produced a uniform staining reaction throughout the tumors suggesting that most of the fibronectin in neurofibromas is plasma-derived. The latter observation suggests that neurofibroma cells are freely accessible to various plasma proteins, including growth factors, which may influence the growth characteristics of these lesions.

Topics: Antibodies, Monoclonal; Antigens; Antigens, Neoplasm; Cell Transformation, Neoplastic; Collagen; Desmin; DNA Probes; Endothelium; Epithelium; Epitopes; Extracellular Matrix; Factor VIII; Fibronectins; Gene Expression Regulation; Humans; Immunoenzyme Techniques; Immunohistochemistry; Keratins; Mast Cells; Membrane Glycoproteins; Mucin-1; Muscles; Mutation; Neurofibromatosis 1; Nucleic Acid Hybridization; Phagocytes; Procollagen; RNA, Messenger; S100 Proteins; Schwann Cells; Skin Neoplasms; von Willebrand Factor

Changes in the intermediate filament composition of rat kidney proximal tubule cells in culture have been investigated. The data suggest that differentiated tubular epithelial cells do not express vimentin, but vimentin expression is induced when the cells begin to proliferate in culture. The cultured cells are positive for both cytokeratins and vimentin by immunofluorescence microscopy. The data support the concept that the intermediate filament composition of proximal tubule epithelial cells can be altered during proliferation induced by nephrotoxic chemicals or by neoplastic transformation.

Topics: Animals; Blotting, Western; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Epithelium; Fluorescent Antibody Technique; Intermediate Filament Proteins; Keratins; Kidney Tubules, Proximal; Kinetics; Male; Molecular Weight; Rats; Rats, Inbred Strains; Vimentin

Malignant cell lines related to mouse lung alveologenic carcinoma have been established from urethane-induced tumors and after in vitro spontaneous transformation of preneoplastic cell lines. Both the chemically and spontaneously transformed cell lines formed invasive, poorly differentiated carcinomas with secondary lung deposits when implanted subcutaneously in immune-suppressed mice. They differed from the related preneoplastic cell line in coordinately exhibiting anchorage-independent growth, reduced epidermal growth factor receptor activity and absence of pericellular fibronectin. These data suggest that similar molecular events may occur in type 2 pneumocyte-related cells in order to generate mouse lung alveologenic adenomas and carcinomas by both spontaneous and chemical carcinogen induction mechanisms. A reduced level of pericellular fibronectin was also demonstrated in an in situ compressive urethane-induced mouse lung adenoma. Loss of pericellular fibronectin may therefore be an early and persistent phenotypic alteration during transformation to the alveologenic adenoma and carcinoma.

Topics: Adenoma; Animals; Cell Line, Transformed; Cell Transformation, Neoplastic; Female; Fibronectins; Keratins; Lung Neoplasms; Mice; Mice, Inbred BALB C; Pulmonary Alveoli; Urethane

The p117 keratinocyte cell line was derived in culture from chemically induced mouse papillomas. The benignly transformed nature of these cells was demonstrated by their ability to re-form benign papillomas when grafted back onto the animal. Retroviral vectors were used to introduce into the p117 cells three distinct oncogenes: v-Ha-ras, p53, and neu. All three oncogenes were able to induce tumorigenic conversion of the p117 keratinocytes when assayed by subcutaneous injection into nude mice. However, grafting the oncogene-transformed cells onto the back of the mouse revealed important differences in the ability of the three oncogenes to induce a fully malignant phenotype. While the ras-transformed papilloma cells formed aggressive carcinomas, p53 and neu transformation yielded an intermediate phenotype, with formation of large exophytic tumors, not yet invasive but with highly dysplastic features remarkably similar to those of in situ carcinomas. These findings establish a homologous, genetically modifiable cell system in which various stages of malignant transformation can be directly compared.

Topics: Animals; Carcinogenicity Tests; Cell Line, Transformed; Cell Transformation, Neoplastic; Epidermal Cells; Epidermis; Genes, ras; Keratins; Mice; Mice, Nude; Oncogenes; Papilloma; Skin Neoplasms; Skin Transplantation

Murine papilloma cell lines 308 and SP-1 have been used as recipients for transfected oncogenes to investigate malignant conversion. These cell lines express an activated c-rasHa gene with a codon 61 mutation and produce squamous papillomas when transplanted as skin grafts onto nude mice. They are not tumorigenic by subcutaneous injection. Both papilloma cell lines were stably transfected with plasmid DNA containing either a rearranged murine plasmacytoma-derived c-myc (minus exon 1), adenovirus 5 E1A, FBJ v-fos or a human c-fos/FBJ v-fos chimera, using cotransfection with the neomycin resistance gene contained in pSV2neo to select for transformants. Southern and northern blotting analysis confirmed the uptake and expression of exogenous DNA in both G418-selected cell lines and in the derived tumors. Unlike the E1A- and myc-containing plasmids, both fos constructs caused malignant conversion in either cell line, as defined by the squamous cell carcinoma histology of tumors from grafted cells and the development of carcinomas after subcutaneous injection into athymic nude mice. Immunofluorescence analysis for specific keratin gene expression indicated that tumors derived by introduction of either of the fos oncogenes were devoid of staining for K1, a 67 kDa epidermal keratin that is expressed in papillomas but not in squamous carcinomas. Tumors from E1A, myc, or pSV2neo transfectants expressed K1, although in a focal distribution. The malignant phenotype induced by the fos oncogene constructs was not associated with the ability to form agar colonies in vitro or to express gamma-glutamyl transpeptidase in the tumors. Since both 308 and SP-1 were sensitive to the fos oncogene for malignant conversion and insensitive to E1A or myc, it is possible that fos may cooperate with the endogenous-activated c-rasHa gene to convert these cells to malignancy. However, since gamma-glutamyl transpeptidase activity is found in the majority of chemically induced mouse skin carcinomas that possess an activated c-rasHa gene, fos activation may not be a common pathway for spontaneous malignant conversion.

Topics: Animals; Biomarkers, Tumor; Cell Division; Cell Line; Cell Transformation, Neoplastic; Female; Fluorescent Antibody Technique; gamma-Glutamyltransferase; Immunoblotting; Keratins; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Oncogenes; Papilloma; Plasmids; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins p21(ras); Transfection

A deficiency in DNA repair, manifest as enhanced chromatid radiosensitivity during the G2 phase of the cell cycle, together with a proliferative stimulus such as that provided by active oncogenes may be necessary and sufficient for the malignant neoplastic transformation of human keratinocytes in culture. Normal epidermal keratinocytes established as continuous cell lines by transfection with pSV3-neo or infection with adeno 12-SV40 hybrid virus developed enhanced G2 chromatid radiosensitivity after 18 passages in culture. In contrast to cells from primary or secondary culture, these cells could be transformed to malignant neoplastic cells by infection with Kirsten murine sarcoma virus containing the Ki-ras oncogene or in one line by the chemical carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine; both of these agents produced a marked proliferative response. Cytological heterogeneity and karyotypic instability characterized the cells during their progression to neoplasia. These results are interpreted in terms of a mechanism for neoplastic transformation.

Topics: Animals; Carcinogens; Cell Transformation, Neoplastic; Cells, Cultured; Chromatids; Chromosome Aberrations; DNA Repair; Humans; Interphase; Keratins; Mice; Oncogenes; Radiation Tolerance; Skin Neoplasms; Transfection

Human keratinocytes and fibroblasts isolated from foreskin were transformed by transfection with recombinant human papillomavirus type 16 (HPV16) DNA. The transformed cells exhibited an extended (fibroblasts) or indefinite (keratinocytes) life-span compared with that of normal controls. In addition, HS27, a human fibroblast cell line previously transfected with origin-defective simian virus 40, was successfully transfected. HPV16 sequences were stably maintained in the cells, and extensive amplification and rearrangements occurred with continuous culturing. Moreover, both fibroblasts and keratinocytes expressed several specific HPV16 mRNAs. Because HPV16-transfected cells had viral transcripts and because transfection with the vector alone did not extend the life-span of the cells, it is likely that the virus was responsible for the indefinite life-span. Transfected fibroblast and keratinocyte lines will be useful for investigating the molecular biology of HPV16 and the interactions between the viral DNA and the human genome. Moreover, transfected keratinocytes provide a model for analyzing the effects of HPV16 on the differentiation properties of human epithelial cells.

Topics: Cell Transformation, Neoplastic; Cells, Cultured; DNA, Viral; Fibroblasts; Humans; Infant, Newborn; Keratins; Male; Papillomaviridae; Skin; Transfection

Topics: Cell Transformation, Neoplastic; Cells, Cultured; Child; Chromatography, High Pressure Liquid; Epidermis; Fibronectins; Glycosaminoglycans; Heparitin Sulfate; Humans; Infant, Newborn; Keratins; Male; Sulfates

Phorbol ester tumour promoters like phorbol-12-myristate-13-acetate (PMA) and serum-derived factors inhibit growth and induce terminal differentiation in normal human and mouse keratinocytes but have a much reduced effect on their transformed counterparts. These observations may be relevant to potency of PMA and wounding as tumour promoters in mouse epidermis. Since some serum factors produced during wounding are thought to exert their effects through the production of diacylglycerol (DAG-the proposed physiological ligand for the phorbol ester receptor) from phospholipids by the activation of phospholipase C (PC) we have compared the effects of PC with PMA in cultures of normal and transformed human keratinocytes. The addition of PC from Clostridium perfringens (0.1-3.0 units/ml) to the culture medium of normal human keratinocytes produced similar morphological changes to PMA and also mimicked the effects of the phorbol ester on cloning efficiency and cornified envelope formation. Most importantly PC, like PMA, had a very weak effect on the human squamous cell carcinoma lines SCC-12B and SCC-15. All the effects of PC were abolished by boiling the enzyme. These results are discussed in relation to the proposed role of serum factors in tumour promotion by deep skin wounding and their mechanistic relationship to phorbol esters.

Topics: Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epidermis; Humans; Keratins; Kinetics; Skin; Tetradecanoylphorbol Acetate; Type C Phospholipases

Prekeratin of simple epithelia with m.m. 55 kD (PK55) was found in all the studied tumorigenic and non-tumorigenic liver epithelial cell lines of the IAR series by means of indirect immunofluorescent methods in combination with corresponding monoclonal antibodies. The most prominent expression was observed in some tumorigenic cell lines. Expression of PK55 was reversible--the cells lost prekeratin in low density cultures. It has been found that the synthesis of prekeratins with m.m. 49 kD (PK49) and 40 kD (PK40) began on reaching higher cell densities than those needed for PK55 synthesis in IAR6-7 line. The PK40 appeared in cells spread on the substratum, while the PK49 was observed in upper poorly spread cells of ridges in multilayered dense cultures. Thus, the synthesis of prekeratins is not constitutive at least for some types of epithelial cells. Specific cell-to-cell interactions are presumably needed for each particular prekeratin synthesis induction.

Topics: Animals; Antibodies, Monoclonal; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Epithelial Cells; Epithelium; Fluorescent Antibody Technique; Gene Expression Regulation; Intermediate Filament Proteins; Keratins; Liver; Molecular Weight; Protein Precursors; Rats; Time Factors

Cultured keratinocytes and squamous carcinoma cells provide a useful model system for studying the processes involved in the regulation of differentiation, as the differentiation capacity of the cells can be modulated experimentally by changing the extracellular calcium concentration. Furthermore, the squamous carcinoma cell lines exhibit a defect in their differentiation capacity which they express to different extents. In this paper, the effect of external lipoproteins has been studied on lipid synthesis in normal keratinocytes and three squamous carcinoma cell (SCC) lines which showed a decreasing capacity to differentiate in the order of normal keratinocytes greater than SCC-12F2 greater than SCC-15 greater than SCC-4. The ability of the cells to form cornified envelopes was taken as a measure of differentiation capacity. The rate of total lipid synthesis as well as the phospholipid-neutral lipid ratio decreased in the order SCC-4 greater than SCC-15 greater than SCC-12F2 greater than or equal to normal keratinocytes, clearly correlating with the differentiation capacity of the cells. Because of the high rate of phospholipid synthesis and the low rate of ceramide synthesis, it is concluded that, under these in vitro conditions used, the maturation of keratinocytes proceeds to a lesser extent than that seen under in vivo conditions. In proliferating cells, in which the low-density lipoprotein (LDL) receptor is operative to a high extent, the rate of lipogenesis, especially that of neutral lipids, responded dramatically to changes of extracellular lipoprotein concentration. In the presence of lipoproteins a marked decrease of cholesterol and triacylglycerol synthesis and an increase of cholesterol ester synthesis has been observed. On the other hand, in differentiating cells lipogenesis appeared to be independent of extracellular lipoproteins, due to the absence of the LDL uptake mechanism, the only exception being the synthesis of triacylglycerols, the rate of which could be modulated to a certain extent by extracellular lipoproteins. The results presented here demonstrate a close inverse relationship between the regulation of lipogenesis by extracellular lipoproteins and the ability of the cells to differentiate.

Topics: Calcium; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cholesterol Esters; Epidermal Cells; Epidermis; Humans; Keratins; Lipids; Lipolysis; Lipoproteins; Microscopy, Phase-Contrast; Triglycerides

We have previously described a human keratinocyte line, NM1, which had been carried for more than 400 doublings, was trisomic for chromosome 8, and appeared to make a number of structural proteins characteristic of keratinocytes. This line has now been carried for more than 800 doublings and grows with the same vigor. It reaches confluence in 7 to 10 days and can be grown without a feeder layer for more than 15 passages. Its karyotype has remained 47,XY, +8. The current NM1 cells make readily detectable amounts of 67 kd and 48 kd keratins, and it has been established that the previously poorly resolved 58 kd band actually consists of 58 kd and 59 kd bands. We have also found that the apparent 56 kd band consists of the 56 kd and 56.5 kd bands. A unique basic polypeptide precursor of the cornified envelope has been discovered in the NM1 line. Although similar in charge to one in normal cells it is lower in molecular weight.

Topics: Carcinogens; Cell Line; Cell Transformation, Neoplastic; Chromosomes, Human, Pair 8; Humans; Keratins; Skin; Trisomy

Human foetal keratinocytes were transfected with a recombinant plasmid (pSV6-1) which contained an origin defective SV40 genome. The resulting transformed cell line had many properties in common with previously described SV40-transformed keratinocytes, including expression of simple epithelial-type keratins. It was non-tumourigenic in nude mice at early passages, forming small benign cysts, however, after approximately 46 in vitro passages, these transformed keratinocytes formed invasive squamous cell carcinomas in athymic nude mice. Several in vitro changes were associated with this acquisition of tumourigenicity (a) an alteration in cellular morphology, (b) development of a cytogenetically marked clone and (c) loss of cell surface fibronectin. The loss of fibronectin was also observed in vivo; cysts formed by SV6-1 Bam/HFK produced human fibronectin whereas tumours did not, although both tumours and cysts were laminin- and keratin-positive. These results indicate that the spontaneous development of secondary events in immortalised human cells may lead to the acquisition of a malignant phenotype.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Transformed; Cell Transformation, Neoplastic; Cell Transformation, Viral; Epidermis; Fibronectins; Fluorescent Antibody Technique; Humans; Karyotyping; Keratins; Male; Mice; Mice, Nude; Neoplasm Transplantation; Simian virus 40; Skin Neoplasms; Transfection

The transcriptional promoter of the candidate E6-E7 transforming gene region of human papillomavirus (HPV)-16 (P97) was active in transiently transfected cervical carcinoma cells when linked to the HSV-1 tk or bacterial cat genes. Sequences 5' to P97 contain a short enhancer element responding to cellular factor(s) in uninfected human foreskin keratinocytes and in cervical carcinoma cells, but not in human or animal fibroblasts. The E2 trans-activator products of HPV-16 or of the related bovine papillomavirus (BPV)-1 further elevated HPV-16-driven transcripts in co-transfections, and required the presence of E2-binding ACC(N)6GGT cores in cis. A 'short E2' C-terminal repressor gene product (sE2) of HPV-16 or the BPV-1 sE2 repressor not only inhibited viral E2 trans-activation, but also suppressed enhancer response to keratinocytic factors. Suppression by the sE2 products was abolished by deletion of the E2-binding cores in cis or by a mutation in the sE2 DNA binding domain. The keratinocyte-dependent enhancer is likely to contribute to the epithelial cell tropism of HPV-16, and may direct persistent E6-E7 gene transcription in response to cellular factors in cervical carcinoma cells in which the viral E2 genes are inactive.

Topics: Animals; Cell Line; Cell Transformation, Neoplastic; Enhancer Elements, Genetic; Epidermal Cells; Female; Gene Expression Regulation; Genes; Genes, Viral; Humans; Keratins; Papillomaviridae; Promoter Regions, Genetic; Transcription, Genetic; Transfection; Uterine Cervical Neoplasms

Keratins are alpha-type fibrous polypeptides which basically compose 10 nm thick or intermediate-sized filaments (IF) in almost all epithelial cells and tissues. Their patterns of expression in normal and malignant upper digestive tract squamous epithelium were monitored by high resolution gel electrophoresis, immunoblotting, and immunohistochemical techniques. Uninvolved epithelia, in all instances, were found to express keratins 4, 5, 6 and 13, 14 the members of the high molecular weight basic (type II or Type B) and of the low molecular weight acidic (type I or type A) subfraction, respectively. Cancers of squamous epithelial cell origin retain keratin synthesis. However, their overall patterns of keratin expression appeared aberrant when compared with those of normal epithelia. In particular, these differences result from highly proliferative tumour cells unable in most cases to synthesize keratins 4/13, a type II/type I keratin pair which specifically indicates in squamous primarily non-keratinizing epithelia completely, i.e. terminally differentiated (suprabasal or spinous) cells. The patchwise expression of acidic keratin 13 in related primaries confirms their heterogeneous phenotype, and may be explained, in part, by cancer cells no longer resistant to terminal differentiation as a result perhaps of an altered micro-environment and/or in response to various effects mediated by vitamin A. We discuss some problems pertinent to the biochemical analysis of keratin polypeptides in normal and involved epithelial tissues, and relate to the controversial question whether specific keratin members may actually candidate for markers of malignancy.

Topics: Carcinoma, Squamous Cell; Cell Division; Cell Transformation, Neoplastic; Epithelium; Head and Neck Neoplasms; Humans; Immunoenzyme Techniques; Keratins; Neoplasm Staging

Human epidermal cells, despite being 'immortalized' or 'transformed' by combinations of either oncogenic virus (SV40, adenovirus 12 or Kirsten murine sarcoma virus) or chemical carcinogen (N-methyl-N'-nitro-N-nitrosoguanidine or 4-nitroquinoline-1-oxide) exhibited similar keratins (although quantitatively reduced) to that of control cells when grown in vitro. However, athymic nude mouse tumors derived from such cells exhibited suppression of the 52, 56 and 58 kd (basic type II) keratins and a predominance of small-sized (40-48 or 50 kd) (acidic type I) keratins. The synthesis of these specific keratins was resumed following re-establishment of cell lines in culture. These results suggest that the changes in keratin protein profiles frequently exhibited by human carcinomas represent a component of the pleomorphic transformed phenotype which can be uncoupled from neoplastic growth.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Epidermal Cells; Epidermis; Humans; Keratins; Male; Methionine; Mice; Mice, Nude; Molecular Weight; Neoplasm Transplantation; Simian virus 40; Skin

An immunohistochemical survey of the distribution of keratin was studied in chemically induced carcinomas of the submandibular glands of mice. Initial signs of premalignant changes were degranulation of granular convoluted tubule cells and deposition of keratin protein in small limited areas of the degranulated cells. There was a gradual increase in the area showing keratin staining in altered tubule cells. Duct-like and cystic structures stained intensely for keratin, as did squamous metaplastic epithelial cells. Induced carcinomas were variably keratinized. Basal layers of cells of squamous-cell carcinomas displayed weak keratin staining, and spinous tumor cells and parakeratotic tumor cells showed somewhat increased levels of keratin staining. Some desquamated keratotic tumor cells stained intensely for keratin. Just as the localization of epidermal and nerve growth factors and lectin-binding histochemistry have been used in studying tumorigenesis in the mouse submandibular gland, immunohistochemically detected keratin proved to be a useful marker of tumor cells of ductal segment origin.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Epithelium; Histocytochemistry; Immunoenzyme Techniques; Keratins; Male; Metaplasia; Mice; Mice, Inbred Strains; Precancerous Conditions; Salivary Gland Neoplasms; Submandibular Gland; Submandibular Gland Neoplasms

Using the well-differentiated human squamous carcinoma cell line LICR-LON-HN-5 as an immunogen we have produced a monoclonal antibody (32a) that reacts with the keratohyalin granular layer of the normal epidermis. We present here results showing the distribution of the epitope recognized by this antibody in human tissues in vivo and in vitro, as demonstrated using immuno cytochemical staining techniques at the light and ultrastructural levels. Expression of the determinant first appears at 18 weeks of fetal development, localized in cells associated with the hair germ. In hyperplastic epidermis the staining pattern is altered, apparently linked with a switch from orthokeratotic to parakeratotic keratinization. In primary squamous cell carcinomas and in xenografts formed by the squamous carcinoma cell line LICR-LON-HN-5 the keratinized elements are stained. Poorly differentiated tumours are not stained, indicating that antibody may be useful as a marker of terminal differentiation in vivo. When grown on collagen gels both human epidermal keratinocytes and the squamous carcinoma cell line show staining of the more differentiated cells which appears to be associated with the keratinohyalin granules, indicating that this antibody may be of value in studies aimed at the control of squamous differentiation.

Topics: Antibodies, Monoclonal; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Epitopes; Humans; Keratins; Skin; Skin Neoplasms

For six patients with partial or total laryngectomies with extensive mucosal hyperplasias, we generated DNA histograms from multiple mucosal sites using Feulgen-stained tissue sections and microspectrophotometric microscopy. Aneuploid DNA histograms were identified in the mucosa of all six specimens, indicating that neoplastic transformation had occurred. The histologic characteristics of neoplastic change included thickened or hyperplastic epithelium, surface maturation or keratinization, often a proliferation of small, immature basallike cells in the depths of the epithelium, and evidence of abnormal epithelial maturation as evidenced by focal areas of cytoplasmic keratinization in the lower portions of the mucosa. We think this histologic expression of intraepithelial neoplasia is more common than the "classic" form of carcinoma in situ with full mucosal replacement by proliferating immature basallike cells. Keratin is a common reaction in laryngeal mucosa, and its presence on the surface or in the depths of the epithelium does not militate against the diagnosis of severe intraepithelial neoplastic transformation.

Topics: Aged; Carcinoma; Carcinoma in Situ; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; DNA, Neoplasm; Epithelium; Humans; Hyperplasia; Keratins; Laryngeal Mucosa; Laryngeal Neoplasms; Male; Middle Aged

Efforts to investigate the progression of events that lead human cells of epithelial origin to become neoplastic in response to carcinogenic agents have been aided by the development of tissue culture systems for propagation of epithelial cells. In the present study, nontumorigenic human epidermal keratinocytes immortalized by adenovirus 12 and simian virus 40 (Ad 12-SV40) were transformed by treatment with the chemical carcinogens N-methyl-N'-nitro-N-nitrosoguanidine or 4-nitroquinoline-1-oxide. Such transformants showed morphological alterations and induced carcinomas when transplanted into nude mice, whereas primary human epidermal keratinocytes treated with these chemical carcinogens failed to show any evidence of transformation. This in vitro system may be useful in assessing environmental carcinogens for human epithelial cells and in detecting new human oncogenes.

Topics: 4-Nitroquinoline-1-oxide; Adenoviruses, Human; Animals; Cell Line; Cell Transformation, Neoplastic; Cell Transformation, Viral; Epidermal Cells; Humans; Keratins; Methylnitronitrosoguanidine; Mice; Mice, Nude; Neoplasm Transplantation; Nitroquinolines; Oncogenes; Simian virus 40; Skin Neoplasms

Immunomorphological examination of the distribution of three keratins in cultured rat liver-derived epithelial cell lines of the IAR series was performed in order to find out the effects of neoplastic evolution on the expression of these epithelium-specific markers. Specific monoclonal antibodies were used to reveal various intermediate filament proteins: keratins with molecular masses of 55, 49 or 40 kD (K55, K49 or K40), and vimentin. The expression of keratins was negligible in sparse and dense cultures of non-transformed lines, which had typical epithelial morphology. The examined keratins were also absent in the sparse cultures of transformed lines, which have lost partially or completely the morphological features of epithelia. However, cells in dense cultures of most transformed lines contained numerous keratin filaments. It is suggested that the paradoxical increase of keratin expression after transformation is due to increased saturation density of transformed cultures; this high density favours the expression. As shown by the experiments with culture wounding, the effects of density are local and reversible. While K55 was present in all the cells of dense cultures, the expression of the other two keratins was dependent on the cell position within these cultures. It is suggested that the expression of the latter two keratins, besides high cell density, also requires the presence (K40) or the absence (K49) of cell-substratum contacts. Possible mechanisms of the effects of cell density on the expression of keratins are discussed.

Topics: Animals; Cell Adhesion; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Cytoskeleton; Epithelium; Fluorescent Antibody Technique; Intermediate Filaments; Keratins; Liver; Rats; Vimentin

A clonal continuous cell line consisting of pre-differentiated lung epithelial cells was established from a Syrian golden hamster fetus on day 15 of gestation. Immunocytological techniques revealed cytoplasmic filaments positive to antikeratin antibody and positive to antivimentin antibody in these cells. Supplementation of the medium with epidermal growth factor, insulin, other types of hormones and vitamin A markedly enhanced cell growth and DNA synthesis. Simultaneously, the cell differentiation characterized by mucus-like glycoprotein production was greatly stimulated. Removal of vitamin A diminished cell growth and this differentiation. The cell system was found to be useful for studying the relationship between in vitro cell transformation and differentiation. Additionally, studies to define general chemical toxicities in this cell system by evaluating several parameters such as cell survival, sialic acid content and enzyme leakage into the medium are in progress.

Topics: Animals; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cricetinae; Cytoskeleton; DNA; Female; Fetus; Fluorescent Antibody Technique; Glucosamine; Keratins; Lung Neoplasms; Mesocricetus; Mucus; Vimentin; Vitamin A

The clinicopathological features of 32 thymomas were reviewed and tumours were staged according to their degree of invasion. Their antigenic profiles were studied using monoclonal antibodies to cytokeratins (CAM 5.2 and DAKO-CK1), HNK-1 (Leu 7), and HLA-DR (TAL-IB5). Stage I (non-invasive) tumours were mainly of the spindle cell (SC) or predominantly lymphocytic (PL) types, whilst all the predominantly epithelial (PE) tumours were either locally invasive (stage II) or showed more extensive spread (stage III). Neoplastic epithelial cells all expressed cytokeratin, but varied in their degree of positivity. CAM 5.2 was more uniformly positive with cells at the periphery of tumour nodules and lining tubulo-cystic areas staining most strongly. DAKO-CK1 gave less uniform positivity but highlighted areas of medullary differentiation. HNK-1 was variably expressed in all tumour groups but was found more often in the invasive tumours (73 per cent stage III, 62 per cent stage II, 50 per cent stage I), particularly those of PE or mixed (M) type. In general, TAL-IB5 expression was lost in the more invasive thymomas. Focal medullary differentiation in tumours suggests a common origin for cortical and medullary epithelium, indicating that sub-division of tumours into cortical or medullary types is not valid. Immunohistochemistry may usefully complement clinical and macroscopic findings in the assessment of malignancy in thymoma.

Topics: Adolescent; Adult; Aged; Antibodies, Monoclonal; Antigens, Neoplasm; Cell Transformation, Neoplastic; Child; Child, Preschool; Female; Humans; Immunoenzyme Techniques; Infant; Keratins; Male; Middle Aged; Neoplasm Staging; Thymoma; Thymus Gland; Thymus Neoplasms

The development of malignant tumors in carcinogen-treated mouse skin appears to involve several genetic changes. Genetic changes which initiate the process are believed to induce alterations in the normal pattern of epidermal differentiation, resulting in the formation of benign tumors, i.e., epidermal papillomas. Subsequent changes appear to be required for the malignant conversion of papillomas to epidermal, squamous-cell carcinomas. Activation of the rasHa gene occurs frequently in chemically induced benign skin papillomas as well as squamous cell carcinomas and thus may represent one mechanism to achieve the initiation step. In the present study, we analyzed several cell lines derived from chemically induced mouse skin papillomas for the presence of transforming oncogenes by transfection of their DNA into NIH 3T3 cells. These papilloma cell lines exhibit an altered differentiation program, i.e., the ability to proliferate under culture conditions favoring terminal differentiation. When DNA from six separate cell lines was tested in the NIH 3T3 transfection assay, active transforming activity was not detected. However, when the EJ rasHa gene was introduced into three of the papilloma cell lines by DNA transfection, transfectants showed an enhanced capacity to proliferate under differentiating culture conditions and formed rapidly growing, anaplastic carcinomas in nude mice. Our findings suggest that in some papilloma cells, a genetic change distinct from rasHa activation may produce an altered differentiation program associated with the initiation step, and this genetic alteration may act in a cooperating fashion with an activated ras gene to result in malignant progression.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Keratins; Mice; Mice, Inbred Strains; Mice, Nude; Neoplasm Transplantation; Oncogenes; Papilloma; Skin

Cells of the normal colonic mucosa express several types of cytokeratins, the characteristic intermediate filament proteins of epithelial cells. An immunohistochemical study was designed to examine the expression of two distinct groups of cytokeratins, recognized by monoclonal antibodies AE1 and AE3, in the colonic mucosa and to compare the findings with those obtained with a large number of polypoid lesions (adenomatous and hyperplastic) and carcinomas of the colon. AE1 and AE3 immunostaining was found in the surface epithelium and upper portions of the crypts of Lieberkühn (functional zone) of normal colonic mucosa, whereas the lower portions of the crypts (proliferative compartment) were unreactive with both AE1 and AE3. Polypoid lesions of the colonic mucosa can be placed into two categories based on their patterns of cytokeratin expression. Solitary tubular adenomas and hyperplastic polyps are composed of AE1 and AE3 nonexpressing cells with only few, patchy areas of AE1 and AE3 expressing cells present within glands and in the surface epithelium. In contrast, villous adenomas show strong AE1 and AE3 reactivity throughout the glands. Furthermore, tubular and villous adenomas, and even histologically normal mucosa in patients with familial polyposis, show AE1/AE3 expression throughout the glands and surface epithelium. Colonic carcinomas show a predominance of AE1/AE3 expressing cells. Thus, cytokeratins recognized by monoclonal antibodies AE1 and AE3 represent molecular markers of cellular maturation in the normal colonic mucosa, that are expressed in colonic carcinomas and, in addition, serve as markers that distinguish colonic mucosa and adenomas with a high risk for development of cancers from those with a lower risk.

Topics: Adenocarcinoma; Adenoma; Adenomatous Polyposis Coli; Antibodies, Monoclonal; Cell Differentiation; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Epithelium; Humans; Hyperplasia; Intestinal Mucosa; Keratins

Expression of the myc and fos genes has been monitored in mouse primary keratinocytes after induction of terminal differentiation by calcium or tetradecanoylphorbol acetate (TPA). myc RNA levels in growing cells are very high and remain elevated even at late times after calcium-induced differentiation. Thus, keratinocytes provide the first example of normal primary cells with persistent c-myc expression irrespective of their proliferative or differentiated state. fos expression is also relatively unaffected by addition of calcium. In contrast to calcium, TPA-induced differentiation is accompanied by dramatic changes in proto-oncogene expression: marked c-fos induction and considerable although transient decrease in c-myc expression. These effects might be important for the keratinocyte response to TPA: TPA treatment of a keratinocyte cell line (RBK) resistant to this substance has no effect on c-myc expression and leads only to minimal c-fos induction. In these cells full fos induction can still be triggered by addition of fresh medium. Thus, the fos gene in normal keratinocytes is inducible through at least two independent mechanisms, only one of which has been lost during derivation of the TPA-resistant cell line.

Topics: Animals; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Genetic Variation; Keratins; Mice; Mice, Inbred BALB C; Oncogenes; RNA, Neoplasm

Lung tumors of various types, fixed in 4% formaldehyde-1% glutaraldehyde, were stained for keratin proteins. The results were compared with previous ultrastructural evidence of intermediate filament bundles (IFBs), presumed to be keratin. Electron and light microscopic methods were largely complimentary and the results were in agreement in 79% of cases. Light microscopy was superior for demonstrating keratinizing foci containing numerous well-developed IFBs, whereas electron microscopy was superior when keratin filaments were sparsely distributed in cells throughout a tumor. Fetal and adult bronchial specimens were also studied. Normal adult bronchus, fixed in aldehydes, was unreactive but keratin was observed in similarly fixed bronchi that showed epidermoid metaplasia and/or intraepithelial carcinoma. Keratin was demonstrated in normal adult bronchi fixed in ethanol. Keratin was not observed in fetal lung until the 14th week of gestation, when it appeared in basal cells and a few columnar cells of the larger bronchi. Thereafter, keratin progressively appeared in the more distal branches. As specimens from gestations of less than 14 weeks were fixed in aldehydes, the apparent lack of immunoreactivity may have been artifactual. Nevertheless, keratin was demonstrable in aldehyde-fixed fetal bronchi at 16 and 23 weeks' gestation.

Topics: Adult; Carcinoma, Bronchogenic; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Cytoskeleton; Fetus; Histocytochemistry; Humans; Infant, Newborn; Keratins; Lung Neoplasms; Phenotype

We have compared the responses of normal human cervical keratinocytes (HCE) to diethylstilboestrol (DES), and the promoting agents, phorbol-12-myristate-13-acetate (PMA) and mezerein using the loss of cloning efficiency as a measure of terminal differentiation in vitro. Dose-response studies showed that normal HCE are growth inhibited by chronic exposure to DES at concentrations greater than or equal to 2.5 X 10(-5) M, to PMA at concentrations greater than 10(-8) M and mezerein at concentrations greater than 10(-9) M. Compared to acetone controls, promoter or DES-treated cells exhibited a 10- to 12-fold increase in cornified-envelope formation. Normal HCE exhibit a heterogeneous response to PMA in that 85-90% of colony-forming cells lose their colony-forming ability after a 24-h exposure to 10(-6) M PMA. The PMA-resistant subpopulation, PMAR, remains constant and is not reduced even after 96 h chronic exposure to PMA. In contrast, the colony-forming ability of normal HCE is almost totally suppressed after 24 h exposure to 10(-6) M mezerein. After 24 h incubation with 5 X 10(-5) M DES, 20% of normal HCE are capable of colony formation but this resistant fraction is eliminated after 96 h chronic exposure. Cornified-envelope formation was negligible in malignant cervical keratinocytes grown in the presence of DES or promotors and these cells were characterised by a very large PMAR fraction - 85 - 90% of cells retained colony-forming ability after exposure to 10(-6) M PMA for 24 h. Furthermore, 90-100% of malignant cervical keratinocytes retained their colony-forming capacity after exposure to 10(-6) M mezerein. However, colony-forming ability declined steadily in the presence of 5 X 10(-5) M DES and after 96 h only a tiny fraction, 1% of malignant cervical keratinocytes could form colonies on replating. The mechanisms by which DES inhibits growth and induces cornified-envelope formation in HCE would appear to be distinct from those activated by PMA and mezerein.

Topics: Carcinogens; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Cervix Uteri; Clone Cells; Diethylstilbestrol; Diterpenes; Female; Humans; Keratins; Kinetics; Phorbols; Terpenes; Tetradecanoylphorbol Acetate

Two new mouse epidermal cell lines have been isolated and characterized as target cells for three chemical carcinogens. The ability to grow these cells at low density (approximately 5 clonogenic cells/cm2) has permitted more precise quantitation of chemical carcinogen-induced changes in epidermal differentiation. The cell lines, designated 291 and 271c, retain the property previously observed in primary cultures of mouse epidermal cells, that is the regulation of terminal differentiation by extracellular Ca2+ ion. Altered response to extracellular Ca2+ after carcinogen treatment of these cells is the basis of the assay endpoint. Other normal properties demonstrated by these cells are keratin immunofluorescence patterns, ability to form cornified envelopes in response to Ca2+ and a lack of tumorigenicity. Both of the lines have high cloning efficiencies (up to 20%) and characteristic epidermal morphology. Their chromosome number, however, is near tetraploid. Dose response studies indicated an increase in colonies with altered response to Ca2+ proportional to the dose of three chemical carcinogens: DMBA 0.001-0.5 microgram/ml X 24 h, MCA 0.01-5 micrograms/ml X 24 h and MNNG 0.01-0.2 micrograms/ml X 1 h. The optimized assay protocol has provided a reproducible means of quantitating carcinogen-altered epidermal cells relative to carcinogen dose, and of isolating cell clones for studies of altered differentiation in carcinogenesis and chemotherapy.

Topics: Animals; Calcium; Carcinogens; Cell Transformation, Neoplastic; Chromosome Aberrations; Clone Cells; Epidermis; Keratins; Mice; Mice, Inbred BALB C

Topics: Animals; Calcium; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Chromosome Aberrations; Keratins; Mice; Neoplasm Transplantation; Phenotype; Precancerous Conditions; Skin Neoplasms

Carcinosarcomas are rare tumors of the upper aerodigestive tract, consisting of both carcinomatous and sarcomatous tissue. The larynx and oral cavity are most frequently involved. There has been much controversy regarding the histological nature and clinical course of these tumors. We report a case of carcinosarcoma of the floor of mouth in a 76 year old man who presented with a large pedunculated sublingual mass. There was no evidence of regional or systemic metastatic disease. After local excision, he was followed for one year without evidence of recurrence or metastasis. A review of the literature is presented, with an attempt to clarify clinically relevant aspects of nomenclature, pathogenesis, and clinical course.

Topics: Aged; Carcinoma, Squamous Cell; Carcinosarcoma; Cell Transformation, Neoplastic; Epithelium; Humans; Immunoenzyme Techniques; Keratins; Male; Microscopy, Electron; Mouth Floor; Mouth Mucosa; Mouth Neoplasms; Neoplasms, Multiple Primary

The generation of hydrogen peroxide (H2O2) and the derivative free hydroxyl radical (. OH) in cultures of mouse cells grown in the presence of visible light and ambient oxygen was shown previously to be implicated in chromatid damage. Furthermore, chromosome alterations appear to be associated with the spontaneous neoplastic transformation of mouse cells in culture. An attempt was made in this study to reduce the incidence of chromosomal aberrations and delay or prevent the onset of spontaneous neoplastic transformation of freshly isolated mouse cells, both fibroblasts and epidermal keratinocytes, by adding catalase to the culture medium, shielding the cultures from wavelengths less than 500 nm and providing a gas phase of 0-1% O2. These conditions significantly decreased the incidence of chromosomal aberrations in both cell types, and in fibroblasts prevented tumourigenicity in non-irradiated syngeneic mice, and increased latent periods for tumour development in X-irradiated mice. The epidermal keratinocytes were particularly resistant to spontaneous neoplastic transformation under all conditions tested. These observations on the protective effect of extracellular catalase suggest that H2O2, a normal metabolite, and/or the derivative .OH can directly or indirectly produce genetic damage and neoplastic transformation in mouse fibroblasts.

Topics: Animals; Catalase; Cell Line; Cell Transformation, Neoplastic; Chromatids; Chromosome Aberrations; Chromosomes; Epidermis; Fibroblasts; Free Radicals; Hydrogen Peroxide; Keratins; Light; Mice; Oxygen; Time Factors

We have examined 18 primary malignant lung tumours categorized as either carcinosarcoma, blastoma or spindle-cell carcinoma according to accepted criteria. Two monoclonal antibodies to keratins, CAM 5.2 and LP 34, were used to determine whether the non-epithelial or spindle-cell components of each tumour showed evidence of keratin expression. By this means the epithelial nature of the five tumours classified as spindle-cell carcinomas was confirmed. In all four pulmonary blastomas and in five of nine carcinosarcomas, the sarcomatous elements failed to stain for keratin but in the remaining four carcinosarcomas there was focal staining. The histogenesis of these tumours is discussed and it is suggested that the sarcomatous component of a carcinosarcoma may be derived from malignant epithelial cells by a process of mesenchymal metaplasia with a switch in intermediate filament type. It remains uncertain whether blastomas are derived from both endoderm and mesoderm, or from either one of these tissues, with one component representing complete metaplastic transformation.

Topics: Aged; Antibodies, Monoclonal; Carcinoma; Carcinosarcoma; Cell Transformation, Neoplastic; Female; Humans; Intermediate Filaments; Keratins; Lung Neoplasms; Male; Middle Aged; Neoplasms, Germ Cell and Embryonal

Topics: Animals; Basement Membrane; Breast; Breast Neoplasms; Cell Transformation, Neoplastic; Epithelium; Female; Humans; Keratins

Topics: Cell Transformation, Neoplastic; Humans; Immunologic Techniques; Keratins; Peptides; Skin; Skin Neoplasms

Antibodies to intermediate filament proteins were used to study different cell layers in normal human transitional epithelium, 16 human transitional cell carcinomas, and two cell lines derived from human bladder carcinomas. Conventional rabbit antisera to human skin keratins stained all layers of the transitional epithelium from bladder, ureter, and kidney. A slightly higher staining intensity was found in the basal and superficial layers as compared with the intermediate cell layers. A monoclonal antibody to cytokeratin 18 (RGE 53), however, stained only the superficial cell layer of transitional epithelium, the so-called umbrella cells. In well-differentiated (grade I) transitional cell carcinomas, RGE 53 stained only the superficial cells of papillary structures. In higher grade papillary tumors, RGE 53 also stained cells within the basal and intermediate layers, whereas in high-grade, invasive tumors almost all tumor cells were RGE 53 positive. These results show that monoclonal antibodies to cytokeratins can provide both an indication of processes involved in neoplastic progression of bladder tumors and a means of studying the molecular relationship of the tumor cells to normal cells.

Topics: Animals; Antibodies, Monoclonal; Antibody Specificity; Antigen-Antibody Reactions; Carcinoma, Transitional Cell; Cell Transformation, Neoplastic; Epithelium; Histocytochemistry; Humans; Immune Sera; Intermediate Filament Proteins; Keratins; Rabbits; Urinary Bladder Neoplasms

The various liver cell populations emerging during the transitory reappearance of alpha-fetoprotein (AFP) in serum of 3'-methyl-4-dimethylaminoazobenzene-ingesting rats were analyzed in situ and in vitro on isolated cell preparations in terms of their cytokeratin and AFP expression using single and double indirect immunofluorescence microscopy. A polyclonal guinea pig antibody raised against cow hoof prekeratin, which recognized a Mr 52,000 cytokeratin, was found to react with bile ductular epithelial cells and oval cells but not with hepatocytes. A monoclonal antibody against a Mr 55,000 cytokeratin reacted not only with bile ductular and oval cells but also with hepatocytes. In contrast, a polyclonal antibody against porcine eye lens vimentin reacted with sinusoidal cells and stroma cells. To assess further the heterogeneity of the emerging cell populations, liver cells were isolated after 4 weeks of treatment and fractionated according to cell size and ploidy level into 4 fractions (I to IV) by velocity sedimentation at 1 X g. A cell-type analysis using AFP and albumin as functional markers revealed the presence of AFP-producing cells in Fraction IV at a mean velocity equivalent to that of newborn diploid rat hepatocytes, whereas most of the albumin-producing cells were distributed in Fractions I to III at velocities similar to those of adult tetraploid rat hepatocytes. A similar analysis based on the differential expression of Mr 52,000 and Mr 55,000 cytokeratins and vimentin in bile ductular and other diploid epithelial cells, hepatocytes, and mesenchymal cells showed that large cells in Fractions I to III were tetraploid hepatocytes, whereas viable cells present in Fraction IV were diploid epithelial cells and mesenchymal cells in proportion of 62 and 38%, respectively. These cell populations could be resolved further by changing the sedimentation time. A subsequent examination of the Mr 55,000 cytokeratin-containing diploid epithelial cells in Fraction IV using double immunofluorescence microscopy resolved three cell populations with respect to Mr 52,000 cytokeratin and AFP expression, namely, two cell populations expressing either protein marker and a third one containing both markers. These results suggest a ductular origin of oval cells and a possible relation to immature hepatocytes.

Topics: alpha-Fetoproteins; Animals; Antibodies, Monoclonal; Bile Ducts; Cell Differentiation; Cell Transformation, Neoplastic; Epithelium; Keratins; Liver; Liver Neoplasms; Male; Molecular Weight; Rats; Rats, Inbred F344

MSK-C3H-NU, a cloned mouse epidermal keratinocyte cell line, was established from the epidermis of C3H/HeN mammary tumor virus-positive nude mice. Although it has lost its diploid chromosome number, the cell line is nontumorigenic, has been stable during serial subcultivations for over 2 years, and has retained some differentiated biological characteristics of normal keratinocytes. MSK-C3H-NU cells were cultured in growth medium containing 7,12-dimethylbenz(a)anthracene. After 2 months, colonies exhibited marked changes in cell morphology, cell arrangement, and keratinization pattern that appeared. The transformation frequency per 10(5) survivors in 7,12-dimethylbenz(a)anthracene-treated (10, 100, and 500 ng/ml) subgroups was 0, 119, and 1370 for Experiment I and 3.9, 238, and 2500 for Experiment II, respectively. Most of these transformed cells became malignant and formed tumors in nude mice. Histologically, the tumors were well-differentiated, keratinizing, squamous cell carcinomas showing papillary growths. In 7,12-dimethylbenz(a)anthracene-treated subgroups, cells from colonies that retained the original morphological characteristics did not form tumors in animals, and in control groups, no cell population showed tumorigenicity. In the MSK-C3H-NU cell system, the morphological alterations seem to be strongly associated with malignant conversion.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Chromosomes; Clone Cells; Keratins; Mice; Mice, Inbred C3H; Mice, Nude; Skin Neoplasms

Human lung tumor cell lines established from the major histological types of lung cancer were examined by immunofluorescent staining techniques for their patterns of intermediate filament (keratin, vimentin, and neurofilament triplet protein) expression. All cell lines examined, both small cell lung carcinoma (SCLC) and non-SCLC (squamous cell carcinoma, adenocarcinoma, large cell carcinoma, and mesothelioma) contained keratin, consistent with their epithelial derivation. These lung carcinoma cell lines also expressed vimentin, the characteristic intermediate filament of mesenchymal cells in vivo. In light of the proposed neuroectodermal origin of SCLC, cell lines were also studied for neurofilament expression. Two of four SCLC tumor cell lines, as well as non-SCLC cell lines, showed no reactivity with antibodies to neurofilament triplet protein. Two of the SCLC cell lines stained weakly with anti-neurofilament antibody. Examination of specific keratin patterns in human lung tumor cell lines by selective immunoprecipitation with keratin antiserum and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that small-sized keratin proteins (Mr 44,000 to 52,000) were present in cell lines derived from SCLC and non-SCLC types of lung cancer. Tumor cell lines exhibiting squamous differentiation by light microscopic criteria (i.e., intracellular keratin, intercellular bridging, "pearl" formation, and/or individual cell keratinization) also displayed a preponderance of intermediate-sized keratins (Mr 57,000 and 59,000) and exhibited another feature of terminal keratinocyte differentiation (cross-linked envelope formation). Mesothelioma cell lines had varying keratin profiles. The presence of keratin proteins in all SCLC cell lines examined argues against a neuroectodermal origin for these tumors and is consistent with the notion that these tumors arise from a common bronchial "stem cell," similar to that from which other types of bronchogenic carcinomas arise.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Cytoskeleton; Fluorescent Antibody Technique; Humans; Intermediate Filament Proteins; Keratins; Lung Neoplasms; Mesothelioma; Mice; Microfilament Proteins; Molecular Weight; Protein Precursors; Vimentin

Seventeen cases of verrucous carcinoma of the oral cavity were reviewed. It was found that cytologic features generally associated with viral modification were observed in 15 of these cases. This finding suggests that viruses may play some role in the pathogenesis of verrucous carcinoma. The hypothesis that an opportunistic, persistent virus may act in concert with frank carcinogens to promote the development of verrucous carcinoma is discussed.

Topics: Adult; Aged; Animals; Carcinoma, Papillary; Cell Nucleus; Cell Transformation, Neoplastic; Cell Transformation, Viral; Epithelium; Female; Humans; Hyperplasia; Keratins; Male; Middle Aged; Mouth Neoplasms; Tumor Virus Infections

Recent investigations have begun to dissect the number and nature of genetic alterations associated with cancer cells. In the present study, primary human epidermal keratinocytes acquired indefinite life-span in culture but did not undergo malignant conversion in response to infection with a hybrid of adenovirus 12 and simian virus 40. Addition of Kirsten murine sarcoma virus, which contains a K-ras oncogene, to these cells induced morphological alterations associated with the acquisition of neoplastic properties. These findings demonstrate the malignant transformation of human primary epithelial cells in culture and support a multiple-step process for neoplastic conversion.

Topics: Adenoviruses, Human; Animals; Cell Transformation, Neoplastic; Epithelial Cells; Humans; Keratins; Kirsten murine sarcoma virus; Mice; Mice, Inbred C3H; Mice, Nude; Sarcoma Viruses, Murine; Sarcoma, Experimental; Simian virus 40; Skin

Normal human keratinocytes are able to stratify, form cornified squames, and terminally differentiate in tissue culture. These properties are frequently impaired by malignant transformation. In the present paper, we show that, in addition, transformation by SV40 results in the coordinate reexpression of a whole set of fetal characters. Moreover, a comparison of two SV40-transformed human keratinocyte cell lines, one still showing a certain degree of stratification and terminal differentiation (HE-SV) and the other almost completely unable to differentiate (SVK14), suggests that the impairment of differentiation and the intensity of reexpression of fetal markers are correlated. Particularly, a set of three keratin polypeptides, absent in adult stratified epithelia but normally found in the nonstratified fetal epidermis, is present in much larger amounts in SVK14 cells than in HE-SV cells. On the other hand, the inability of SVK14 cells, in contrast to HE-SV cells, to form cornified envelopes seems to be due to the inability of those cells to accumulate involucrin.

Topics: Basement Membrane; Cell Differentiation; Cell Transformation, Neoplastic; Cells, Cultured; Epidermis; Female; Humans; Keratins; Laminin; Molecular Weight; Peptides; Phenotype; Simian virus 40; Skin

For the purpose of clarifying cellular differentiation of epithelioid sarcoma, studies based on various methods were performed. Enzyme histochemical studies showed that epithelioid sarcoma tumor cells have characteristics intermediate between epithelial cells and the large plump cells of synovial sarcoma-incomplete epithelial differentiation. For alkaline phosphatase and adenosine triphosphatase particularly, positive cells and negative cells coexisted, as in the large plump cells of synovial sarcoma. Immunohistochemical studies for alpha 1-antitrypsin, alpha 1-antichymotrypsin, vimentin, and keratin also showed that epithelioid sarcoma tumor cells are very similar to the large plump cells of synovial sarcoma and have incomplete epithelial differentiation. For example, the examinations of serial sections and double staining methods revealed that keratin-positive cells are always vimentin-positive in epithelioid sarcoma and in the monophasic area of synovial sarcoma. Electron-microscopically, bundles of intermediate filaments and filopodia toward the intercellular lumen were observed, as in the monophasic area of synovial sarcoma. The results of enzyme-histochemical and immunohistochemical studies of non-neoplastic synovial lining cells, performed here for the first time, are also discussed.

Topics: Adenosine Triphosphatases; Adult; Alkaline Phosphatase; Cell Transformation, Neoplastic; Female; Fibroma; Humans; Keratins; Male; Microscopy, Electron; Middle Aged; Sarcoma; Sarcoma, Synovial; Staining and Labeling; Vimentin

In summary, two in vitro systems with rat tracheal epithelial cells were used to demonstrate that initiated respiratory tract epithelial cells can be promoted or enhanced to transform at a higher frequency by exposure to a noncarcinogenic tumor promoter. The latter two studies suggest possible mechanisms for this enhancement: 1) TPA amplifies the chromosomal instability brought about by initiating doses of carcinogen and 2) TPA alters gene expression and cellular differentiation such that it causes the initiated cell to escape the normal differentiation and senescence process thereby enhancing the proliferative lifespan of initiated cells.

Topics: Aneuploidy; Animals; Cell Transformation, Neoplastic; Cells, Cultured; DNA; Keratins; Methylnitronitrosoguanidine; Rats; Rats, Inbred F344; Tetradecanoylphorbol Acetate; Tracheal Neoplasms

Small cell bronchial carcinoma (SCC) xenografts with differing sensitivity to cyclophosphamide (CY) were investigated using a variety of techniques. Two xenografts (HX78 and HX88) were relatively sensitive to CY, one xenograft (HX72) was inherently resistant to CY and a fourth xenograft (HX78Cy) was a CY induced resistant subline of HX78 and was unstable when maintained without CY exposure. Conventional light microscopy, cytology and electron microscopy examination of the xenografts revealed appearances consistent with SCC. An antikeratin antibody demonstrated the presence of keratin in the inherent and the induced resistant xenografts (and in the unstable revertant) but not in the two sensitive xenografts; the presence of keratin suggested squamous differentiation. Monolayer culture morphology of the sensitive HX78 and the unstable revertant was anchorage independent with the cells forming floating aggregates; the induced resistant subline of this xenograft (HX78Cy) showed, by contrast, flattened, angular adherent cells. Beta HCG production was detected in the monolayer culture supernatant of sensitive HX78 cells; the level of production of beta HCG was increased in the induced resistant HX78Cy cells. Karyotype and flow cytometry studies were also performed. The morphological responses of small cell lung cancer to treatment are discussed.

Topics: Animals; Carcinoma, Small Cell; Cell Transformation, Neoplastic; Cells, Cultured; Chorionic Gonadotropin; Cyclophosphamide; Drug Resistance; Flow Cytometry; Humans; Karyotyping; Keratins; Lung Neoplasms; Mice; Neoplasm Transplantation; Transplantation, Heterologous

Keratin composition has been widely used as a biochemical marker of differentiation in normal epithelia, cell culture systems and tumours of epithelial tissues. We have been developing a model system for the study of human squamous epithelial cell differentiation, and among a panel of monoclonal antibodies we have generated for analysing this system are two antibodies recognizing subsets of epidermal keratins. The two antibodies, designated LICR-LON-16a and LICR-LON-29b, were raised to the human squamous carcinoma cell line LICR-LON-HN-5, and we describe here their biochemical and immunocytochemical characterization. Antibody 16a reacts with only epidermal basal cells in normal human skin and shows specificity for the 45 and 46 kdalton keratins. Antibody 29b stains all living layers of the epidermis, and reacts with a broad range of ketain polypeptides, (45-56 kdaltons) in immunoblotting analyses. We have investigated the alterations of cellular staining that occur in chronic hyperproliferative skin diseases and carcinomas and compared this with the staining of multilayered cultures of normal keratinocytes and the HN-5 cell line. We show that in squamous cell carcinomas and in HN-5 cell xenografts 16a and 29b stain only the well-differentiated cell types. Furthermore we found that the basal cell specificity of 16a was lost in all of the hyperproliferative skin lesions examined including psoriasis and eczema. This transition to suprabasal staining pattern was also seen in the cultures of normal keratinocytes and HN-5 cells. We conclude that aberrant keratin synthesis or abnormal post-translational processing of keratins associated with an increased rate of cell turnover could account for the altered expression of the epitope recognized by antibody 16a.

Topics: Antibodies, Monoclonal; Antibody Specificity; Carcinoma, Basal Cell; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Epitopes; Humans; Keratins; Skin; Skin Neoplasms

We have studied the effects of the potent tumour promoter phorbol, 12-myristate, 13-acetate (PMA) and two non-promoting hyperplasiogenic compounds ethyl phenylpropriolate (EPP) and the divalent cation ionophore A23187 on the terminal differentiation of normal and transformed human keratinocytes using the loss of cloning efficiency and the formation of cornified envelopes as markers of the differentiated state. PMA induced terminal differentiation in a far greater proportion of normal keratinocytes than it did in the squamous cell carcinoma line SCC-27 but EPP and the calcium ionophores A23187 and Br-X537A had no such differential effect, possibly explaining the poor promoting ability of the last three compounds.

Topics: Alkynes; Calcimycin; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Clone Cells; Humans; Hyperplasia; Keratins; Phorbols; Skin; Tetradecanoylphorbol Acetate

A cell population highly enriched in human epidermal basal cells has been obtained and characterized by using antibodies specific for various cell types in the epidermis. Quantitative two-dimensional gel electrophoretic analysis (isoelectric focusing) of [35S]methionine-labeled polypeptides from basal cells and simian virus 40-transformed keratinocytes showed that the basal cells synthesize very low amounts (less than 0.02% of the total protein) of the nuclear, transformation-sensitive protein cyclin as compared to the transformed cells, which synthesize this protein constitutively (0.15% of the total protein). Very low levels of cyclin were observed in total human epidermis, and preliminary studies of two basaliomas have shown a significant synthesis of this protein in these tumors. Immunofluorescence studies using antibodies to proliferating cell nuclear antigen that immunoprecipitate cyclin confirmed the above observations at least in the case of the cultured cells. Taken together, these results support the notion that cyclin may be a central component of the pathway(s) that controls cell proliferation.

Topics: Antigens, Neoplasm; Breast; Cell Transformation, Neoplastic; Electrophoresis, Polyacrylamide Gel; Female; Fluorescent Antibody Technique; Humans; Keratins; Nucleoproteins; Peptide Fragments; Proliferating Cell Nuclear Antigen; Simian virus 40; Skin Physiological Phenomena

The various epithelial cells of the lower respiratory tract and the carcinomas derived from them differ markedly in their differentiation characteristics. Using immunofluorescence microscopy and two-dimensional gel electrophoresis of cytoskeletal proteins from microdissected tissues we have considered whether cytokeratin polypeptides can serve as markers of cell differentiation in epithelia from various parts of the human and bovine lower respiratory tract. In addition , we have compared these protein patterns with those found in the two commonest types of human lung carcinoma and in several cultured lung carcinoma cell lines. By immunofluorescence microscopy, broad spectrum antibodies to cytokeratins stain all epithelial cells of the respiratory tract, including basal, ciliated, goblet, and alveolar cells as well as all tumor cells of adenocarcinomas and squamous cell carcinomas. However, in contrast, selective cytokeratin antibodies reveal cell type-related differences. Basal cells of the bronchial epithelium react with antibodies raised against a specific epidermal keratin polypeptide but not with antibodies derived from cytokeratins characteristic of simple epithelia. When examined by two-dimensional gel electrophoresis, the alveolar cells of human lung show cytokeratin polypeptides typical of simple epithelia (nos. 7, 8, 18 and 19) whereas the bronchial epithelium expresses, in addition, basic cytokeratins (no. 5, small amounts of no. 6) as well as the acidic polypeptides nos. 15 and 17. Bovine alveolar cells also differ from cells of the tracheal epithelium by the absence of a basic cytokeratin polypeptide. All adenocarcinomas of the lung reveal a "simple-epithelium-type" cytokeratin pattern (nos. 7, 8, 18 and 19). In contrast, squamous cell carcinomas of the lung contain an unusual complexity of cytokeratins. We have consistently found polypeptides nos. 5, 6, 8, 13, 17, 18 and 19 and, in some cases, variable amounts of cytokeratins nos. 4, 14 and 15. Several established cell lines derived from human lung carcinomas (SK-LU-1, Calu -1, SK-MES-1 and A-549) show a uniform pattern of cytokeratin polypeptides (nos. 7, 8, 18 and 19), similar to that found in adenocarcinomas. In addition, vimentin filaments are produced in all the cell lines examined, except for SK-LU-1.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Adenocarcinoma; Animals; Carcinoma, Squamous Cell; Cattle; Cell Line; Cell Transformation, Neoplastic; Diagnosis, Differential; Humans; Keratins; Lung; Lung Neoplasms

The complete tumour promoter phorbol, 12-myristate, 13-acetate (PMA) induces terminal differentiation in the majority of normal cultured human and mouse keratinocytes but a subpopulation exists which is resistant to this effect (PMAR). We have compared with PMA the effects of mezerein (Mez) and phorbol, 12-retinoate, 13-acetate (PRA) on the ability of normal and transformed human and mouse keratinocytes to terminally differentiate in an attempt to elucidate why the latter two compounds are inefficient complete tumour promoters but are effective as second-stage promoters when given after PMA in the two-stage promotion regimen. Both PMA and Mez increased cornified envelope formation in a similar way in normal and transformed keratinocyte cultures inducing a 20- to 25-fold increase over the solvent controls in normal keratinocytes but only a 2-fold increase in line SCC-27 (a cell line derived from a human squamous cell carcinoma). However, while quantitative dose response studies of the effect of phorbol esters on colony forming ability revealed a proportion of normal human and mouse keratinocytes which were resistant to PMA, no normal keratinocytes were resistant to Mez or PRA. In contrast, cell lines derived from papillomas and squamous cell carcinomas showed a resistant fraction of similar size with all three compounds. Furthermore, when Mez or PRA were mixed with PMA the survival of line SCC-27 was the same as when the cultures were treated with the compounds individually indicating that the keratinocytes which were resistant to PRA or Mez were also the PMAR subpopulation. A non-tumorigenic subclone of line SCC-12 (clone F.2), previously shown to possess all known properties of transformed keratinocytes except defective terminal differentiation in suspension culture responded to PMA and Mez in a similar way to normal keratinocytes, suggesting that resistance of the PMAR subpopulation to second-stage promoters requires the expression of a defect in the keratinocyte terminal differentiation programme.

Topics: Carcinogens; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Diterpenes; Drug Resistance; Epidermal Cells; Epidermis; Humans; Keratins; Phorbol Esters; Phorbols; Terpenes; Tetradecanoylphorbol Acetate

Previous studies have indicated that some Simian-virus-40-transformed human epidermal keratinocytes (SV40-HE) undergo significant changes in their growth and differentiated properties. To better understand the significance of these changes, we have characterized the keratins of SV40-HE cells by one- and two-dimensional immunoblot analysis using the subfamily-specific AE1 and AE3 monoclonal antikeratin antibodies. The results indicate that our SV40-HE cells have lost the Mr 58,000 (No. 5), Mr 56,000 (No. 6), Mr 50,000 (No. 14/15), Mr 48,000 (No. 16), and Mr 46,000 (No. 17) keratins that are expressed by cultured normal human keratinocytes. Instead, these cells express mainly Mr 52,000 (No. 8), Mr 45,000 (No. 18), and Mr 40,000 (No. 19) keratins, a set highly characteristic of simple epithelial cells. Furthermore, our SV40-HE cells have ceased to express involucrin, another marker for keratinocytes, and have a greatly diminished ability to undergo in vitro stratification. These results suggest that epidermal cells can sometimes lose their keratinocyte features as a consequence of viral transformation. This finding may have important implications regarding the mechanisms of epithelial differentiation and tumorigenesis and in the use of keratinocyte markers for tumor diagnosis.

Topics: Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Epithelial Cells; Humans; Keratins; Molecular Weight; Protein Precursors; Simian virus 40; Skin

Topics: Animals; Cell Transformation, Neoplastic; Etretinate; Female; Humans; Keratins; Male; Middle Aged; Neoplasm Recurrence, Local; Papilloma; Skin Neoplasms; Tretinoin; Urinary Bladder Neoplasms

Complementary DNA (cDNA) clones constructed to the 55, 59 and 67 kilodalton (K) keratins, the major keratins synthesized in newborn mouse epidermis, were used as molecular hybridization probes to examine the expression of these genes in newborn epidermis and normal and malignantly transformed epidermal cells in culture. Transcripts of these three keratin genes are abundant in newborn epidermis. However, primary cultures of epidermal cells contain very low levels of these RNAs. The decreased expression of these keratin genes in primary cells appears to be due to factors within the culture system. Unlike primary-cell cultures, the malignantly transformed cell line Pam 212 synthesizes keratin proteins and mRNAs similar to newborn epidermis, including the 67 K keratin. However, synthesis of the 67 K keratin in Pam 212 cells is modulated by culture factors. Keratin gene expression in another Pam line, 321, differs from that of Pam 212 cells in that decreased expression of these three keratin genes occurs. These results indicate that keratin genes that are normally expressed in vivo in epidermis may be expressed in malignant epidermal cells under conditions that do not permit expression of these genes in nonmalignant primary epidermal cells.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Epidermal Cells; Gene Expression Regulation; Keratins; Mice; Mice, Inbred BALB C; Molecular Weight; Transcription, Genetic

A human papillomavirus type 1 (HPV-1)/pBR322 recombinant plasmid was constructed consisting of two complete HPV-1 genomes in a head-to-tail arrangement, inserted into the BamHI site of pBR322. To obtain established human keratinocytes keratinocytes carrying dimeric HPV-1, origin-minus simian virus 40 (SV40) DNA was used as a dominant transforming marker in co-transfection experiments performed with cultured human foetal keratinocytes. Using the Southern blotting technique, HPV-1 DNA was detected in one out of five SV40-transformed human keratinocyte cell lines which were obtained in this way. Further blotting experiments using SV40, pBR322 and HPV-1 DNA as probes revealed patterns of hybridization which were consistent with co-integration of SV40 DNA with between two and four copies of the HPV-1 genome. The HPV-1 sequences in this cell line were virtually non-methylated and were transcribed at a lower level than SV40 mRNAs in the same cultured cells. A low level of monomeric HPV form I DNA was detected by blotting of undigested total cellular DNA. No evidence for a stimulation of form I HPV DNA replication could be obtained by blotting analysis of total DNA extracted from keratinizing cysts, which were formed by subcutaneous inoculation of these cells into nude mice.

Topics: Cell Transformation, Neoplastic; Cells, Cultured; Cloning, Molecular; DNA Replication; DNA, Recombinant; DNA, Viral; Female; Fetus; Genes, Viral; Humans; Keratins; Nucleic Acid Hybridization; Papillomaviridae; Pregnancy; Simian virus 40; Skin

We have studied the action of phorbol, 12-myristate, 13-acetate (PMA) on human keratinocytes grown with lethally-irradiated 3T3 cells using medium supplemented with hydrocortisone, cholera toxin and epidermal growth factor. Normal keratinocyte cultures show a heterogeneous response to PMA; 90-93% of the colony-forming cells lose their colony-forming ability and form cornified envelopes when treated for 24 h with doses of 100 nM or less, but the remainder are resistant to doses of 1000 nM. The resistant cells are the precursors of the sensitive ones and heterogeneity is restored to those cells and their progeny after 8 days culture in the absence of PMA. Cultures of 3 squamous cell carcinoma lines, a SV40-transformed human keratinocyte line, and three clones of these lines were found to contain 3-17 times more PMA-resistant keratinocytes than the normal strains, and the size of the PMA-resistant fraction in each line was inversely related to the competence of that line to lose colony-forming efficiency when placed in suspension culture (which is the first detectable change in an ordered programme of events resembling terminal differentiation of the keratinocyte). The number of cells with cornified envelopes in surface cultures of normal human keratinocytes increased from approximately 3% in control cultures to approximately 70% in those treated for 6 days with 100 nM PMA. The transformed human keratinocyte cultures showed a 3-25-fold smaller increase in cornified envelope formation when treated with 100 nM PMA, and the increase in envelope formation by each line when exposed to this dose of PMA was related to the competence of that line to lose cloning efficiency in suspension culture. No relationship was found between the ability of any human keratinocyte strain or line we studied to metabolically inactivate PMA and their resulting response to the compound. The results are discussed in relation to the mechanism of action of PMA as a promoter of epidermal carcinogenesis.

Topics: Animals; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Child; Cholera Toxin; Epidermal Growth Factor; Female; Fetus; Humans; Hydrocortisone; Keratins; Male; Mice; Phorbols; Pregnancy; Simian virus 40; Skin; Skin Physiological Phenomena; Tetradecanoylphorbol Acetate

We have studied the changes in the surface ultrastructure of human epidermal keratinocytes after infection by the oncogenic virus, SV40. The surfaces of uninfected cells show patterns of dense microvilli and microridges in varying proportion with microridges predominating in the center of the developing keratinocyte colony and on the fully mature keratinocytes (squames) and the microvilli being more prevalent on the immature proliferative cells at the colony periphery. As the transformation process progressed over time highly microridged surfaces became less numerous. After many months in culture cell surfaces were found to exhibit more sparse, predominantly villous ultrastructure. The surface characteristics of the infected cells in long term culture were similar to those observed in a cell line derived from a squamous cell carcinoma.

Topics: Carcinoma, Squamous Cell; Cell Membrane; Cell Transformation, Neoplastic; Cells, Cultured; Humans; Infant, Newborn; Keratins; Male; Microscopy, Electron, Scanning; Microvilli; Simian virus 40; Skin

Eight cell lines exhibiting resistance to Ca2+ induced terminal differentiation were derived from primary mouse epidermal cultures and their properties analyzed. The lines developed either spontaneously (2 lines) or after exposure of primary cultures to carcinogens or carcinogens and tumor promoter. All but one of the lines were of epithelial or epitheloid morphology but 3 of the 8 lines lacked desmosomes, keratin filaments and immunoprecipitable keratin proteins, and thus could not be defined as keratinocytes. Two of the 5 keratinocyte lines were tumorigenic in syngeneic Balb/c newborns after 4 months in medium containing 1.2 mM Ca2+, and 3 lines remained non-tumorigenic even after 11 months in 1.2 mM Ca2+. All three of the non-keratinizing lines were tumorigenic. Tumorigenic potential of the 5 keratinocyte lines did not correlate with ploidy (as determined by DNA content), transglutaminase activity or growth in soft agar. However, the 2 tumorigenic keratinocyte lines contained cells which stained intensely red for gamma glutamyl transpeptidase activity, while the non-tumorigenic keratinocyte lines did not. Only those lines lacking desmosomes and keratin filaments grew in soft agar, but these lines were negative for gamma glutamyl transpeptidase activity. Ploidy and transglutaminase activity did not correlate with tumorigenicity in these non-keratinizing lines. These results show that cell lines derived from cultured mouse epidermal cells and selected on the basis of their resistance to Ca2+ induced terminal differentiation may be preneoplastic. Furthermore the association of additional markers with malignant change in these cell lines depended on whether or not the cells were keratinizing.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Benz(a)Anthracenes; Calcium; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Drug Resistance; Keratins; Methylnitronitrosoguanidine; Mice; Microscopy, Electron, Scanning; Skin; Skin Neoplasms; Skin Physiological Phenomena

Human fetal lung tissue was cultured on dead, sterile pigskin as a substrate and treated with 3-methylcholanthrene. Several cell lines were obtained. One of the lines produced a tumor in nude mouse and showed anchorage-independent growth in soft agar. The tumor cells were stained positive for keratin, indicating their epithelial origin.

Topics: Antibodies, Neoplasm; Cell Transformation, Neoplastic; Cells, Cultured; Female; Fetus; Humans; Keratins; Lung Neoplasms; Methylcholanthrene

Murine keratinocytes, isolated by flotation trypsinization of skin, can be separated into five groups by centrifugation through Percoll, a colloidal silica gradient. Within each group a good correlation was found between density, plating efficiency, morphological appearance, DNA synthesis, and degree of keratinization/cornification. This method can be applied equally well to fetal, newborn, or adult keratinocytes and should be useful in a variety of studies including isolation of subpopulations of pathological cell types, work on chalones and hyperplastic diseases such as psoriasis, and in vitro transformation studies.

Topics: Animals; Cell Differentiation; Cell Transformation, Neoplastic; Centrifugation, Density Gradient; DNA; Epidermal Cells; Keratins; Mice; Povidone; Silicon Dioxide; Skin

In this study, we report the isolation of a Mr 52,000 keratin from a nontumorigenic bladder epithelial cell line and its localization, by immunofluorescence, in bladder frozen sections at different stages of neoplastic progression and in various carcinogen-transformed bladder epithelial cells and human bladder carcinoma-derived cell lines. The results showed that: (a) the Mr 52,000 keratin is present in basal epithelial cells of the normal bladder but absent from the intermediate and superficial epithelial cell layers; (b) hyperplastic bladder lesions, induced in mice after the administration of butyl-(4-hydroxybutyl)-nitrosamine, resulted primarily from the proliferation of cells in the basal compartment; (c) butyl-(4-hydroxybutyl)nitrosamine-induced bladder carcinomas displayed differential staining with the anti-Mr 52,000 keratin antiserum reflecting divergent differentiated status within a tumor and a subpopulation of cells with reduced overall keratin expression; (d) primary cultures of bladder carcinomas revealed a subpopulation of cells with limited filamentous keratin as was observed in vivo; (e) mouse bladder epithelial cell lines transformed in culture by a chemical carcinogen showed a loss or reduction in expression of the Mr 52,000 keratin; (f) two human bladder carcinoma cell lines displayed very limited expression of the Mr 52,000 keratin. Although the loss or reduction in expression of a specific keratin is unlikely to be responsible for transformation, it may contribute to the heterogeneity in the differentiated state and morphology of bladder epithelial cells during neoplastic progression both in vivo and in culture.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Cell Line; Cell Transformation, Neoplastic; Cricetinae; Epithelium; Fluorescent Antibody Technique; Humans; Immune Sera; Keratins; Mice; Microscopy, Phase-Contrast; Molecular Weight; Rats; Urinary Bladder; Urinary Bladder Neoplasms

Topics: Aging; Cell Division; Cell Survival; Cell Transformation, Neoplastic; Cells, Cultured; Cyclic AMP; Epithelial Cells; Gene Expression Regulation; Humans; In Vitro Techniques; Keratins; Neoplasms

Topics: Animals; Cell Differentiation; Cell Division; Cell Line; Cell Nucleus; Cell Transformation, Neoplastic; Cytoskeleton; Desmosomes; Epidermal Cells; Epidermis; Keratins; Kinetics; Mice; Mice, Inbred C3H; Organoids

Rabbit bladder epithelium, grown on collagen gels and exposed to the chemical carcinogen benzo[a]pyrene, produced nontumorigenic altered foci as well as tumorigenic epithelial cell lines during 120-180 d in culture. Immunofluorescence studies revealed extensive keratin filaments in both primary epithelial cells and benzo[a]pyrene-induced altered epithelial foci but showed no detectable vimentin filaments. The absence of vimentin expression in these cells was confirmed by two-dimensional gel electrophoresis. In contrast, immunofluorescence staining of the cloned benzo[a]pyrene-transformed rabbit bladder epithelial cell line, RBC-1, revealed a reduction in filamentous keratin concomitant with the expression of vimentin filaments. The epithelial nature of this cell line was established by the observation that cells injected into nude mice formed well-differentiated adenocarcinomas. Frozen sections of such tumors showed strong staining with antikeratins antibodies, but no detectable staining with antivimentin antibodies. These results demonstrated a differential expression of intermediate filament type in cells at different stages of neoplastic progression and in cells maintained in different growth environments. It is apparent that the expression of intermediate filaments throughout neoplastic progression is best studied by use of an in vivo model system in parallel with culture studies.

Topics: Animals; Benzo(a)pyrene; Benzopyrenes; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Cytoskeleton; Epithelium; Fluorescent Antibody Technique; Keratins; Muscle Proteins; Rabbits; Urinary Bladder; Vimentin

The effect of 5-halogenated deoxyuridines on the keratinization of an in vitro-transformed rat bladder epithelial cell line, BES20B, was studied by a cover slip culture method established previously. 5-Bromodeoxyuridine (BrdU) and 5-iododeoxyuridine (IdU) both showed dose-dependent inhibition of the keratinization. The complete inhibition of keratinization was achieved by BrdU at concentrations higher than 5 microgram/ml. At these concentrations, BrdU reduced the saturation density of the test cells on a cover slip. Removal of BrdU from the culture medium restored the cell growth as well as the keratinization of BrdU-treated cells, suggesting that the effect of BrdU is reversible. The effect of BrdU was abolished by the addition of excess thymidine. A possible mechanism of the anti-keratinization effect of BrdU is discussed in comparison with that of vitamin A.

Topics: Animals; Binding, Competitive; Bromodeoxyuridine; Cell Line; Cell Transformation, Neoplastic; DNA Replication; Epithelium; Floxuridine; Idoxuridine; Keratins; Rats; RNA, Neoplasm; Thymidine; Urinary Bladder Neoplasms; Vitamin A

Human foreskin epithelial cells were transformed to an anchorage-independent state of growth (in soft agar) and neoplasia (invasion of chick embryonic skin in vitro). Aflatoxin B1, N-methyl-N'-nitro-N-nitrosoguanidine, propanesultone, beta-propiolactone, or ultraviolet absorbance at 254 nm were used successfully as carcinogens. These foreskin epithelial cells, like human foreskin fibroblasts, were readily transformed when treated in S phase but, unlike the transformed fibroblasts, expression of cellular neoplasia did not require an extended period of time in culture. The invasive features of the transformed human epithelial cells in chick embryonic skin in vitro simulated squamous cell carcinoma.

Topics: Animals; Carcinogens; Carcinoma, Squamous Cell; Cell Cycle; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Chick Embryo; Epithelium; Humans; Keratins; Male; Neoplasms, Experimental

Topics: Animals; Argininosuccinate Synthase; Carcinogens; Cell Division; Cell Line; Cell Transformation, Neoplastic; Citrulline; Cricetinae; Cytoskeleton; Desmosomes; Epidermal Cells; Epidermis; Fibroblasts; Keratins; Ligases; Mesocricetus

Cellular retinol-binding protein (CRBP) in cancers and in vitro-transformed epithelial cells of the ACI/N rat urinary bladder was assayed with radiolabeled retinol by sucrose density gradient centrifugation. CRBP was detected in abundance in tumor tissues of all 5 transplantation bladder cancer lines. However, this protein was below the limits of detection in bladder cancer cells of culture lines that had originated from the same primary cancers as the corresponding transplantation lines. CRBP was detected in small quantity in in vitro-transformed epithelial cells of a culture line but it was absent in another line of similar origin. The addition of retinol to the culture medium prevented the in vitro keratinization of CRBP-positive cells but it had no effect on the keratinization of CRBP-negative cells.

Topics: Animals; Cell Line; Cell Transformation, Neoplastic; Keratins; Neoplasm Transplantation; Neoplasms, Experimental; Rats; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Transplantation, Homologous; Urinary Bladder; Urinary Bladder Neoplasms

Topics: Animals; Cell Differentiation; Cell Transformation, Neoplastic; Female; Guinea Pigs; Histocytochemistry; Humans; Immune Sera; Immunologic Techniques; Keratins; Molecular Weight; Mouth Mucosa; Peptides; Skin; Skin Neoplasms

The ultrastructural study of the senile epidermis without an apparent lesion, in the phase of cicatrization of a traumatic lesion and in a state of hyperkeratosic transformation in comparison with those performed in the normal epidermis, reveals decrease in the number of clear keratinocytes (only compensated in the process of cicatrization) it is in concord with the most recent theory of the aging of cells of Gelfant and Smith. The increase of clear suprabasal cells in the epidermis degenerative process with a constant of the percentage of clear and dark basal cells suggest a liberation of bloqued cells, which constituted probably the direct source of neoplasic differentiation.

Topics: Aged; Aging; Cell Transformation, Neoplastic; Epidermis; Humans; Keratins; Keratosis; Middle Aged; Wound Healing

The intermediate-sized filaments present in epidermal keratinocytes derived from mouse skin and in an established cell line (HEL) derived from spontaneous transformation of murine keratinocytes grown in vitro, have been examined by immunofluorescence microscopy, using antibodies directed against subunit proteins of different classes of intermediate-sized filaments, as well as by electron microscopy and gel electrophoresis of cytoskeletal preparations highly enriched in intermediate-sized filaments. The keratinocytes derived from neonatal skin, which are capable of only limited replication in vitro, show only a single type of intermediate-sized filaments, i.e., the tonofibril-like arrays of filaments containing prekeratin. HEL cells, which proliferate indefinitely in vitro, retain the tonofilament-like structures typical of differentiated epidermal cells but in addition display intermediate-sized filaments of the vimentin type, i.e., the filament system typically found in mesenchymal and mesenchyme-derived cells. We discuss the possibility that (i) the advent of vimentin-type filaments in epidermal cells in culture is related either to the transformed state or the in vitro growth conditions as such and (ii) other differentiated epithelial cells proliferating in vitro may have more than one system of intermediate-sized filaments.

Topics: Animals; Cell Differentiation; Cell Division; Cell Line; Cell Transformation, Neoplastic; Cells, Cultured; Cytoskeleton; Epidermis; Fluorescent Antibody Technique; Keratins; Mice; Peptides; Protein Precursors; Skin

In vitro malignant transformation of fetal rat keratinizing epidermal cells from inbred SD rats after benzo[a]pyrene (BP) treatment was analyzed from various biologic viewpoints. BP treatment directly and indirectly effected changes in cell growth characteristics, i.e., temperature dependence for growth, in vitro keratinization, chromosome structure, and the ability to form colonies on plastic substrate, on 0.57% agar medium layer, and in 0.33% soft agar medium. BP-treated cells at 30 degrees C remained in the premalignant stages and showed shifts in chromosome structure toward the hypodiploid range and parakeratotic changes in their keratinization process. However, the cells failed to form colonies even on a plastic substrate. BP-treated cell lines that adapted to temperatures of 35 and 37.5 degrees C remained in the premalignant stages; however, they acquired the ability to form colonies on plastic substrates during subcultivation. Malignantly transformed colonies appeared in these cell lines. In vitro keratinization processes were classified into nearly normal (diffuse lamellar, focal lamellar, and parakeratotic), intermediate, and atypical subtypes (columnar, spherical, and single-cell type). Cells of atypical keratinization subtypes and some of the intermediate subtypes formed squamous cell carcinomas in syngeneic hosts. Malignantly transformed cells showed shifts in chromosome structure toward the hypotetraploid range and colony formation on the 0.57% agar medium layer. However, they failed to form colonies in 0.33% soft agar medium. With the use of changes in biologic characteristics of the cells as indicators, fetal rat keratinizing epidermal cells in culture were classified into five stages. The appearance of stage III cells seemed to be the first key step in their malignant transformation.

Topics: Animals; Benzopyrenes; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Chromosome Aberrations; Clone Cells; Keratins; Neoplasm Transplantation; Neoplasms, Experimental; Rats; Rats, Inbred Strains; Skin Neoplasms; Transplantation, Isogeneic

In vitro and in vivo transformed epithelial cells of the rat urinary bladder keratinized well on a coverslip culture. Keratinization proceeded more rapidly in in vitro-transformed cells than in in vivo-induced bladder cancer cells. The grade of keratinization was estimated from the absorption spectrum of a papanicolaou-stained specimen which afforded two peaks derived from keratinized cells and viable cells. Using this semiquantitative method, effect of various drugs on keratinization was studied on 3 lines of in vitro-transformed epithelial cells. Vitamin A and its analogs prevented the keratinization in 2 out of 3 lines at dosage over 1 micrograms/ml, but other drugs such as vitamin C, E, and K, steroid hormones, cyclic AMP, polyamines, polyanions, and dimethyl sulfoxide were ineffective for preventing keratinization. Amino acid compositions of keratinized cells and viable cells were not fundamentally different.

Topics: Amino Acids; Animals; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Keratins; Male; Neoplasms, Experimental; Rats; Urinary Bladder Neoplasms; Vitamin A

Topics: Adolescent; Cell Transformation, Neoplastic; Humans; Ichthyosis; Infant, Newborn; Keratins; Melanoma; Nevus; Nevus, Pigmented; Skin Neoplasms; Warts

After serial 20-minute trypsinization, skin from near-term fetal MRC rats was separated into epidermis attached to basal lamina and dermis with trapped hair follicles. Fragmentation of the basal lamina by trypsinization caused the epidermal basal cell layer to form cell clumps of various sizes, whereas dermal mesenchymal cells dissociated into single cells, leaving a fibrous network with hair follicles. Clumps of eopdermal basal cells were then isolated with few contaminating hair follicles. At 30-32.5 degrees C, a cornification process caused the clumps to form thickened sheets of nearly pure keratinizing epidermal cells, which were isolated in petri dishes and grew to fill the dishes. The mode of cell differentiation in these sheets resembled an in vivo cornification process; the sheets showed maximum growth at 30 degrees C and failed to survive at 35 degrees C or above. Benzo[a]pyrene (BP) treatment accelerated the growth of cell sheets and enabled them to adapt to 35 and 37.5 degrees C. Keratinizing epidermal cells treated with BP at 35 and 37.5 degrees C were serially subcultured at these temperatures and lost their original dependence on lower temperatures for growth. The cell sheets were still capable of in vitro keratinization; however, some qualitative and quantitative changes were observed.

Topics: Animals; Benzopyrenes; Cell Differentiation; Cell Division; Cell Separation; Cell Transformation, Neoplastic; Cells, Cultured; Culture Techniques; Keratins; Mitosis; Rats; Skin; Temperature; Transplantation, Homologous; Trypsin

The fine structural morphology of dyskeratotic and dysplastic keratinocytes in human oral epithelium was investigated by light microscopy as well as by transmission and scanning-electron microscopy. On the epithelial-connective tissue border, marked changes are seen in the form of polymorphous microinvasive cytoplasmic processes of basal keratinocytes, structural alterations of the basement membrane and rarefaction of the subepithelial connective tissue. Dyskeratotic keratinocytes with abnormal tonofilament configuration are phagocytized by dermal macrophages and are transported to the lamina propria. Ultrastructural signs of atypia are found in the nucleus, nucleolus, cytoplasmic organelles and mitotic apparatus. Furthermore, multiple alterations of the plasma membrane, decrease in numbers of junctional complexes and acantholytic widened intercellular spaces are observed. Intracytoplasmic lumina are formed by endocytotic invagination of desmosome-studed plasma membrane regions at the cell surface. Despite an inverse relationship between the degree of keratinization and the glycogen content of the epithelium at the subcellular level, large amounts of glycogen are found in some keratinocytes. The epithelial surface is formed by hyper-, para-, or orthokeratosis, or shows individual cell keratinization, alteration of the disintegration process and defective keratin synthesis. The ultrastructural analysis of dysplastic keratinocyte populations reveals some morphological criteria common with invasive squamous cell carcinoma, which may be important in the early diagnosis and prognosis of malignant transformation of epithelial dysplasia.

Topics: Cell Membrane; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Glycogen; Humans; Intercellular Junctions; Keratins; Leukoplakia; Macrophages; Mouth Mucosa; Organoids; Phagocytosis

Topics: Carcinoma, Squamous Cell; Cell Differentiation; Cell Transformation, Neoplastic; Humans; Keratins; Lymph Nodes; Lymphatic Metastasis; Microtomy; Mouth Neoplasms; Neck Dissection

Topics: Animals; Cell Transformation, Neoplastic; Endometrium; Epithelium; Female; Hyperplasia; Hypertrophy; Intrauterine Devices; Keratins; Metaplasia; Ovarian Diseases; Ovary; Polyps; Rats; Time Factors; Uterine Diseases; Uterus

Topics: Adenocarcinoma; Animals; Carcinoma, Squamous Cell; Cell Line; Cell Transformation, Neoplastic; Connective Tissue Cells; Cricetinae; Cytopathogenic Effect, Viral; Embryo, Mammalian; Keratins; Neoplasm Transplantation; Neoplasms, Experimental; Sarcoma, Experimental; Simian virus 40; Skin; Transplantation, Homologous; Virus Cultivation