tetracycline and Osteosarcoma

Anatomic pathology has produced considerable knowledge about environmental teratogens and carcinogens. A special disease registry established by a pathologist provided details of the association between oral contraceptives and hepatic neoplams. Pathologists were also involved in establishing in the link between diethylstilbestrol use and clear-cell adenocarcinomas of the vagina. An area of particular interest has been gender and ethnic differences in the incidence of certain diseases. Pathologists further make use of animal studies to investigate the pathogenesis of human tumors. Finally, stored serum or tissue is often used by pathologists to help diagnose diseases retrospectively. Human skin fibroblasts grown in culture and stored have been especially valuable for laboratory research. This chapter briefly highlights some of the milestones in the detection of enviromental effects through anatomic pathology.

Topics: Adenocarcinoma; Animals; Ataxia Telangiectasia; Contraceptives, Oral; Diethylstilbestrol; Disease Models, Animal; Environment; Ethnicity; Female; Geography; Herpesvirus 4, Human; Humans; Liver Neoplasms; Lymphoproliferative Disorders; Osteosarcoma; Pathology, Clinical; Radiation, Ionizing; Sex Factors; Tetracycline; Thorium Dioxide; Tooth Discoloration; Vaginal Neoplasms; X Chromosome

Nonstructural protein 5A (NS5A) is a membrane-associated essential component of the hepatitis C virus (HCV) replication complex. An N-terminal amphipathic alpha helix mediates in-plane membrane association of HCV NS5A and at the same time is likely involved in specific protein-protein interactions required for the assembly of a functional replication complex. The aim of this study was to identify the determinants for membrane association of NS5A from the related GB viruses and pestiviruses. Although primary amino acid sequences differed considerably, putative membrane anchor domains with amphipathic features were predicted in the N-terminal domains of NS5A proteins from these viruses. Confocal laser scanning microscopy, as well as membrane flotation analyses, demonstrated that NS5As from GB virus B (GBV-B), GBV-C, and bovine viral diarrhea virus, the prototype pestivirus, display membrane association characteristics very similar to those of HCV NS5A. The N-terminal 27 to 33 amino acid residues of these NS5A proteins were sufficient for membrane association. Circular dichroism analyses confirmed the capacity of these segments to fold into alpha helices upon association with lipid-like molecules. Despite structural conservation, only very limited exchanges with sequences from related viruses were tolerated in the context of functional HCV RNA replication, suggesting virus-specific interactions of these segments. In conclusion, membrane association of NS5A by an N-terminal amphipathic alpha helix is a feature shared by HCV and related members of the family Flaviviridae. This observation points to conserved roles of the N-terminal amphipathic alpha helices of NS5A in replication complex formation.

Topics: Amino Acid Sequence; Amino Acids; Animals; Cattle; Cell Line, Tumor; Cell Membrane; Circular Dichroism; Conserved Sequence; Diarrhea Viruses, Bovine Viral; Electroporation; GB virus A; GB virus B; Humans; Molecular Sequence Data; Osteosarcoma; Peptides; Protein Biosynthesis; Protein Structure, Secondary; Protein Structure, Tertiary; Tetracycline; Transfection; Viral Nonstructural Proteins

We recently developed a new tetracycline-inducible gene switch employing the tetracycline operator-containing hCMV major immediate-early promoter and the tetracycline repressor, tetR, rather than the previously used tetR-mammalian cell transcription factor fusion derivatives.. The present study demonstrates that this tetR-mediated transcription repression system can function as a powerful gene switch for On-and-Off regulation of therapeutic gene expression in ex vivo gene transfer protocols. Firstly, for achieving regulated gene expression in a localized tissue environment, R11/OEGF cells, a stable line that expresses hEGF under the control of the tetR-mediated transcription repression switch, were transplanted into porcine full-thickness wounds enclosed by wound chambers.. By topically applying tetracycline in wound chambers at various concentrations or at different time points post-transplantation, the levels and timing of hEGF expression in transplanted wounds could be reversibly regulated by tetracycline. Over 3000-fold induction in hEGF expression was achieved in the local wound microenvironment. Secondly, R11/OEGF cells were intramuscularly injected into NCr outbread nude mice to test the efficacy of intermittent systemic gene delivery of a soluble peptide(s).. Basal circulating hEGF was undetectable and induced up to at least 1,500-fold after administration of tetracycline. Furthermore, the timing and duration of hEGF expression could be finely adjusted by the presence or the absence of tetracycline in the drinking water.

Topics: Animals; Anti-Bacterial Agents; Cell Line, Tumor; Cytomegalovirus; Epidermal Growth Factor; Female; Gene Expression; Gene Transfer Techniques; Humans; Male; Mice; Mice, Nude; Osteosarcoma; Promoter Regions, Genetic; Skin; Sus scrofa; Tetracycline; Transgenes; Wounds and Injuries

The goal of this study was to design improved regulatable lentivirus vector systems. The aim was to design tetracycline (tet)-regulatable lentivirus vectors based on the Tet-on system displaying low background expression in the absence of the doxycycline (DOX) inducer and high transgene expression levels in the presence of DOX.. We constructed a binary lentivirus vector system that is composed of a self-inactivating (SIN) lentivirus vector bearing inducible first- or second-generation tet-responsive promoter elements (TREs) driving expression of a transgene and a second lentivirus vector encoding a reverse tetracycline-controlled transactivator (rtTA) that activates transgene expression from the TRE in the presence of DOX.. We evaluated a number of different rtTAs and found rtTA2S-M2 to induce the highest levels of transgene expression. Regulated transgene expression was stable in human breast carcinoma cells implanted into nude mice for up to 11 weeks. In an attempt to minimize background expression levels, the chicken beta-globin cHS4 insulator element was cloned into the 3' long terminal repeat (LTR) of the transgene transfer vector. The cHS4 insulator element reduced background expression but expression levels following DOX addition were lower than those observed with vectors lacking an insulator sequence. In a second strategy, vectors bearing second-generation TREs harboring repositioned tetracycline operator elements were used. Such vectors displayed greatly reduced leakiness in the absence of DOX and induced transgene expression levels were up to 522-fold above those seen in the absence of DOX.. Inducible lentivirus vectors bearing insulators or second-generation TREs will likely prove useful for applications demanding the lowest levels of background expression.

Topics: Animals; Breast Neoplasms; Cell Line; Cell Line, Tumor; Cell Transplantation; Chickens; Doxycycline; Female; Fibroblasts; Gene Expression Regulation; Genetic Vectors; Globins; HeLa Cells; Humans; Kidney; Lentivirus; Mice; Mice, Nude; Mice, SCID; Osteosarcoma; Promoter Regions, Genetic; Skin; Tetracycline; Trans-Activators; Transgenes; Transplantation, Heterologous

The regulatory subunit of protein kinase CK2, designated CK2beta, exists both free in cells and in complexes with the CK2 catalytic subunits. Growing evidence suggests that CK2beta has functions dependent and independent of the CK2 catalytic subunits. There have been indications that CK2beta has functions associated with DNA damage responses and in the control of cell proliferation. For example, transient and stable constitutive overexpression of CK2beta in mammalian cells was previously shown to perturb cell cycle progression and to attenuate proliferation. To systematically investigate the molecular mechanisms responsible for these effects of CK2beta on cell proliferation, we generated human osteosarcoma U2OS cell lines with tetracycline-regulated expression of CK2beta. Increased expression of CK2beta results in increases in total cellular CK2 activity, but no changes in cell cycle profiles or proliferation. Furthermore, following exposure to ultraviolet radiation, p53 induction was identical regardless of the levels of CK2beta in cells. Mouse 3T3-L1 cells stably transfected with CK2beta also showed no alterations in cell proliferation. The differences between these results and those previously reported emphasize the complex nature of CK2beta and its cellular functions. Furthermore, these results indicate that increased expression of CK2beta is not by itself sufficient to effect alterations in cell proliferation.

Topics: 3T3 Cells; Animals; Binding Sites; Casein Kinase II; Catalytic Domain; Cell Cycle; Cell Division; DNA Damage; Enzyme Induction; Humans; Macromolecular Substances; Mice; Osteosarcoma; Protein Serine-Threonine Kinases; Protein Subunits; Tetracycline; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Ultraviolet Rays

To study the effects of XW630 on bone formation in overiectomized (OVX) rats and in human osteoblast-like cell line TE85.. Bone histomorphometric analysis was performed with undecalcified bone sections and tetracycline intraperitoneally labeling.. Compared with that of OVX rats, the static data of trabecular bone volume (TBV)/total tissue volume (TTV), TBV/sponge bone volume (SBV) and mean trabecular plate density (MTPD) were enhanced while mean trabecular plate spacing (MTPS) decreased after treated with XW630 for 13w. The dynamic data of single-labeled surface [Sfract(s)], double-labeled surface [Sfract(d)], Sfract(d+1/2s), trabecular osteoid surface (TOS) and bone formation rate in tissue level (Svf) were increased and osteoid maturation period (OMP) shortened in XW630 group. In osteoblast-like cells, both 3H-thymidine incorporation and cell count increased after treated with XW630 for 48. Treated with XW630 for 12 approximately 18h, inducible nitric oxide synthase (iNOS) activity and cGMP content increased in time-dependent manners.. XW630 enhanced bone activation frequency and increased trabecular connectivity, stability, and strength. The cellular mechanism related to effects of XW630 on bone formation in ovariectomized rats.

Topics: Animals; Bone Neoplasms; Cell Division; Cyclic GMP; Estrone; Female; Femur; Humans; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Osteoblasts; Osteogenesis; Osteosarcoma; Ovariectomy; Piperazines; Rats; Rats, Wistar; Tetracycline; Tetracyclines; Tumor Cells, Cultured

Implant surface roughness and chemical composition, as well as other factors, affect the ability of osteogenic cells to form bone adjacent to an implant. The same principles may also apply to the tooth root and some reports have shown that surface modification of the root may lead to improved restoration of the periodontal apparatus. The most common of these surface modification techniques involves demineralization with citric acid or treatment with tetracycline to expose collagen fibrils. In addition, during normal bone remodeling, osteoclasts demineralize the extracellular matrix, leaving resorption pits and exposed collagen fibrils. In this study, the effect of different dentin surface-preparation techniques on osteoblasts were compared.. Slices of sperm whale dentin were mechanically polished and surfaces were treated with tetracycline-HCl (TCN) or were cultured with mouse bone marrow cells to create a surface with osteoclast (OC) resorption pits or left untreated. Profilometry, x-ray photoelectron spectroscopy (XPS), and scanning electron microscopy (SEM) were used to evaluate the 3 different dentin surfaces. MG63 osteoblast-like cells were cultured on the 3 different surfaces and the effect of dentin surface preparation technique on MG63 cell proliferation (cell number), differentiaton (alkaline phosphatase specific activity of isolated cells and cell layer lysates; osteocalcin production), and local factor production (transforming growth factor (TGF)-beta1 and prostaglandin E2 (PGE2) compared.. Profilometry showed the polished and TCN surfaces were smooth with comparable Ra values, whereas the OC surfaces were slightly rougher due to resorption pits which covered 3.7% of the surface. XPS measurements showed that TCN treatment reduced the Ca and P content of the surface, indicating that it had dissolved the mineral. Osteoclast-resorption also reduced the Ca and P content, but to a lesser extent. MG63 cell proliferation on polished dentin and tissue culture polystyrene was equivalent. In contrast, cells grown on the TCN- and OC-treated surfaces exhibited increased proliferation. No effect of surface treatment on cell alkaline phosphatase activity was observed, but activity in the cell layer lysates was increased on the TCN- and OC-treated surfaces. Osteocalcin production was reduced on all dentin surfaces, but the greatest reduction was found on the TCN-treated surface. Production of both TGF-beta1 and PGE2 was increased on the treated surfaces. All effects were greatest in cultures grown on the TCN-treated dentin.. These data indicate that demineralization of the dentin surface promotes proliferation of osteoblasts and early differentiation events like production of alkaline phosphatase and autocrine mediators such as PGE2 and TGF-beta1. However, later differentiation events like osteocalcin production are decreased. Osteoclast-mediated bone resorption elicits similar responses; less than 4% of the dentin surface resulted in approximately 75% of the response caused by TCN treatment. These observations suggest that greater attention should be paid to the effects of osteoclastic resorption in designing methods for enhancing bone and cementum formation adjacent to root surfaces.

Topics: Alkaline Phosphatase; Analysis of Variance; Animals; Calcium; Cell Count; Cell Differentiation; Cell Division; Collagen; Dentin; Dinoprostone; Electron Probe Microanalysis; Extracellular Matrix; Humans; Mice; Microscopy, Electron, Scanning; Osteoblasts; Osteocalcin; Osteoclasts; Osteosarcoma; Phosphorus; Protein Synthesis Inhibitors; Tetracycline; Transforming Growth Factor beta; Tumor Cells, Cultured; Whales

To study the effects of 2-[3-estrone-N-ethyl-piperazine-methyl] tetracycline (XW630) in human osteoblast-like cell line TE85.. [3H]Thymidine incorporation and cell count for cell proliferation, radioimmunoassay for cyclic GMP (cGMP) content, and monitoring the conversion of [3H]arginine for inducible nitric-oxide synthase (iNOS) activity assay.. After treatment with XW630 for 48 h, [3H]thymidine incorporation and cell numbers increased by 62.7% and 96.9%, respectively. NG-monomethyl-L-arginine (L-NMMA, an NOS inhibitor) induced a concentration-dependent inhibitory effect on the proliferation after treatment for 48 h. The inhibitory effect was prevented partially by XW630 (1.0 nmol.L-1). After treatment with XW630 for 12-48 h, iNOS activity and cGMP concentration increased in time-dependent manners.. XW630 stimulated cell proliferation, enhanced iNOS activity and cGMP content in human osteoblast-like cell line TE85.

Topics: Bone Neoplasms; Cell Division; Cyclic GMP; Estrogens, Conjugated (USP); Estrone; Humans; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Osteoblasts; Osteosarcoma; Piperazines; Tetracycline; Tetracyclines; Tumor Cells, Cultured

With the cloning of DNA encoding the trans-dominant negative mutant form of the HSV-1 origin-binding protein UL9, UL9-C535C, under the control of the tet operator-bearing hCMV major immediate-early promoter (pcmvtetO), this article demonstrates that the tetR-mediated mammalian transcription repression switch (Yao et al., Hum. Gene Ther. 9:1939-1950, 1998) can be converted to a novel HSV-1-specific viral replication switch. Using this viral replication switch, the plaque-forming efficiency of infectious HSV-1 DNA can be reversibly regulated by tetR over 100-fold in transient viral infection assays. Moreover, while less than 0 PFU/ml of HSV-1 was detected from tetR-expressing cells transfected with infectious HSV-1 DNA and plasmid pcmvtetOUL9-C535C in the presence of tetracycline, close to 1000 PFU/ml of HSV-1 was produced when similar experiments were carried out in the absence of tetracycline. The tetracycline treatment led no reduction in HSV-1 synthesis in cells transfected with infectious HSV-1 DNA alone. Taken together, given that the UL9-C535C-associated antiviral activity can be silenced in the context of this HSV-1 replication switch, the establishment of this reversible switch would allow construction of a new generation of HSV-1 recombinants able to inhibit its own replication as well as replication of wild-type virus.

Topics: Animals; Base Sequence; Cell Line; Chlorocebus aethiops; Cytomegalovirus; DNA-Binding Proteins; Gene Expression Regulation, Viral; Genes, Regulator; Genetic Vectors; Herpesvirus 1, Human; Models, Biological; Molecular Sequence Data; Osteosarcoma; Promoter Regions, Genetic; Repressor Proteins; Tetracycline; Transfection; Vero Cells; Viral Proteins; Viral Vaccines; Virus Replication

This article describes the first (to our knowledge) tetracycline-inducible regulatory system that demonstrates that the tetracycline repressor (tetR) alone, rather than tetR-mammalian cell transcription factor fusion derivatives, can function as a potent trans-modulator to regulate gene expression in mammalian cells. With proper positioning of tetracycline operators downstream of the TATA element and of human epidermal growth factor (hEGF) as a reporter, we show that gene expression from the tetracycline operator-bearing hCMV major immediate-early enhancer-promoter (pcmvtetO) can be regulated by tetR over three orders of magnitude in response to tetracycline when (1) the reporter was cotransfected with tetR-expressing plasmid in transient expression assays, and (2) the reporter unit was stably integrated into the chromosome of a tetR-expressing cell line. This level of tetR-mediated inducible gene regulation is significantly higher than that of other repression-based mammalian cell transcription switch systems. In an in vivo porcine wound model, close to 60-fold tetR-mediated regulatory effects were detected and it was reversed when tetracycline was administered. Collectively, this study provides a direct implementation of this tetracycline-inducible regulatory switch for controlling gene expression in vitro, in vivo, and in gene therapy.

Topics: Animals; Base Sequence; Chlorocebus aethiops; Cytomegalovirus; Epidermal Growth Factor; Female; Gene Expression Regulation; Gene Transfer Techniques; Genes, Immediate-Early; Genes, Reporter; HeLa Cells; Herpes Simplex Virus Protein Vmw65; Humans; Molecular Sequence Data; Operator Regions, Genetic; Osteosarcoma; Recombinant Fusion Proteins; Repressor Proteins; Swine; TATA Box; Tetracycline; Transcription, Genetic; Tumor Cells, Cultured; Vero Cells; Wounds and Injuries

WT1 encodes a zinc finger transcription factor that is expressed in the developing kidney and the inactivation of which leads to Wilms' tumor, a pediatric kidney cancer. We have recently shown that inducible expression of WT1 in osteosarcoma cells triggers programmed cell death, an effect that is associated with transcriptional repression of the endogenous epidermal growth factor receptor. We now show that WT1-mediated apoptosis is preceded by induction of the cyclin-dependent kinase inhibitor p21, associated with G1 phase arrest. This effect is only demonstrated by WT1 isoforms with an intact DNA binding domain, and it is associated with increased expression of endogenous p21 mRNA. WT1-mediated induction of p21 is independent of p53, another tumor suppressor gene known to regulate p21 expression. In the kidney, p21 is expressed in differentiating glomerular podocytes along with WT1. We conclude that induction of p21 expression may contribute to WT1-dependent differentiation pathways in the kidney and potentially to the function of WT1 as a tumor suppressor gene.

Topics: Anti-Bacterial Agents; Blotting, Western; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; DNA-Binding Proteins; Flow Cytometry; G1 Phase; Genes, p53; Genes, Wilms Tumor; Humans; Kidney; Mutation; Neoplasm Proteins; Osteosarcoma; RNA, Messenger; Tetracycline; Transcription Factors; Transfection; Tumor Cells, Cultured; Tumor Suppressor Protein p53; WT1 Proteins

The effect of overexpression of p21waf1 on drug sensitivity was studied in an osteosarcoma cell line (SaOs-2) lacking both p53 and functional retinoblastoma protein using a tetracycline (TC)-inducible expression system. p21waf1 expression was barely detectable in SaOS-2 cells incubated in the presence of TC. After TC withdrawal, high levels of p21waf1 were induced in these cells. These p21waf1-induced cells showed increased sensitivity to doxorubicin, tomudex, and methotrexate as compared to uninduced cells; this condition is associated with increased apoptosis. Expression of p21waf1 reduced cyclin A-associated kinase activity and, surprisingly, resulted in inhibition of phosphorylation of E2F-1 and increased E2F-1 binding activity. An S-G2 cell cycle arrest/delay and an increase in expression of E2F-responsive genes (dihydrofolate reductase and thymidylate synthase) was correspondingly observed. Overexpression of p21waf1 in cells lacking functional retinoblastoma protein may mediate sensitivity to anticancer drugs by inhibiting E2F-1 phosphorylation, which may contribute to increased S-G2 cell cycle delay and increased cell susceptibility to apoptosis.

Topics: Anti-Bacterial Agents; Antibiotics, Antineoplastic; Antimetabolites, Antineoplastic; Apoptosis; Carrier Proteins; Cell Cycle; Cell Cycle Proteins; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinases; Cyclins; DNA-Binding Proteins; Doxorubicin; E2F Transcription Factors; E2F1 Transcription Factor; Gene Expression Regulation, Neoplastic; Genes, p53; Humans; Methotrexate; Osteosarcoma; Phosphorylation; Quinazolines; Retinoblastoma Protein; Retinoblastoma-Binding Protein 1; Tetracycline; Tetrahydrofolate Dehydrogenase; Thiophenes; Thymidylate Synthase; Transcription Factor DP1; Transcription Factors; Tumor Cells, Cultured

Chromosomal translocations resulting in chimaeric transcription factors underlie specific malignancies, but few authentic target genes regulated by these fusion proteins have been identified. Desmoplastic small round-cell tumour (DSRT) is a multiphenotypic primitive tumour characterized by massive reactive fibrosis surrounding nests of tumour cells. The t(11;22)(p13;q12) chromosomal translocation that defines DSRT produces a chimaeric protein containing the potential transactivation domain of the Ewing-sarcoma protein (EWS) fused to zinc fingers 2-4 of the Wilms tumour suppressor and transcriptional repressor WT1 (refs 2,3). By analogy with other EWS fusion products, the EWS-WT1 chimaera may encode a transcriptional activator whose target genes overlap with those repressed by WT1 (ref. 4). To characterize its functional properties, we generated osteosarcoma cell lines with tightly regulated inducible expression of EWS-WT1. Expression of EWS-WT1 induced the expression of endogenous platelet-derived growth factor-A (PDGFA), a potent secreted mitogen and chemoattractant whose promoter contains the many potential WT1-binding sites. Native PDGFA was not regulated by wild-type WT1, indicating a difference in target gene specificity between this tumour suppressor and its oncogenic derivative. PDGFA was expressed within tumour cells in primary DSRT specimens, but it was absent in Wilms tumours expressing WT1 and Ewing sarcomas with an EWS-Fli translocation. We conclude that the oncogenic fusion of EWS to WT1 in DSRT results in the induction of PDGFA, a potent fibroblast growth factor that contributes to the characteristic reactive fibrosis associated with this unique tumour.

Topics: Binding Sites; Blotting, Northern; DNA-Binding Proteins; Early Growth Response Protein 1; Gene Expression Regulation, Neoplastic; Heterogeneous-Nuclear Ribonucleoproteins; Humans; Immediate-Early Proteins; Immunohistochemistry; In Situ Hybridization; Oncogene Proteins, Fusion; Osteosarcoma; Platelet-Derived Growth Factor; Promoter Regions, Genetic; Recombinant Proteins; Ribonucleoproteins; RNA-Binding Protein EWS; Tetracycline; Transcription Factors; Transcription, Genetic; Translocation, Genetic; Tumor Cells, Cultured

It is well established that induction of the p53 tumor suppressor protein in cells can lead to either cell cycle arrest or apoptosis. To further understand features of p53 that contribute to these cell responses several p53-null Saos2 and H1299 cell lines were generated that express wild-type or mutant forms of p53, or the cyclin-dependent kinase inhibitor p21/WAF1, under a tetracycline-regulated promoter. Our results show that the cellular level of p53 can dictate the response of the cell such that lower levels of p53 result in arrest whereas higher levels result in apoptosis; nevertheless, DNA damage can heighten the apoptotic response to p53 without altering the protein level of p53 in cells. We also demonstrate that arrest and apoptosis are two genetically separable functions of p53 because a transcriptionally incompetent p53 can induce apoptosis but not arrest, whereas induction of p21/WAF1, which is a major transcriptional target of p53, can induce arrest but not apoptosis. Finally, we show that a full apoptotic response to p53 requires both its amino and carboxyl terminus, and our data suggest that there is synergism between transcription-dependent and -independent functions of p53 in apoptosis. Thus, there are multiple independent cellular responses to p53 that together may account for the extraordinarily high frequency of p53 mutations in diverse types of human tumors. The implications of these results are discussed and a model is proposed.

Topics: Apoptosis; Camptothecin; Carcinoma, Small Cell; Cell Cycle; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; DNA Damage; Enzyme Inhibitors; Humans; Mutation; Osteosarcoma; Tetracycline; Topoisomerase I Inhibitors; Transcription, Genetic; Transcriptional Activation; Tumor Cells, Cultured; Tumor Suppressor Protein p53

The ability of the p53 protein to act as a sequence-specific transcriptional activator suggests that genes induced by p53 may encode critical mediators of p53 tumor suppression. Using a tetracycline-regulated p53 expression system and cDNA library subtraction procedure, we identified several p53-induced gene transcripts in human Saos-2 osteosarcoma cells that are novel on the basis of their size, regulation, and low abundance. Wild-type p53-dependent induction of these transcripts was observed in cells that are growth arrested by p53, as well as in cells that undergo apoptosis upon expression of an inducible wild-type p53 transgene. These results show that p53 activates the expression of numerous response genes and suggest that multiple effectors may play a role in mediating cellular functions of p53.

Topics: Apoptosis; Base Sequence; Binding Sites; Blotting, Northern; Blotting, Western; Cell Line; DNA, Neoplasm; Gene Expression Regulation, Neoplastic; Gene Library; Genes, p53; Humans; Kinetics; Luciferases; Molecular Sequence Data; Mutagenesis; Mutation; Nuclear Proteins; Osteosarcoma; Plasmids; RNA, Messenger; Temperature; Tetracycline; Transcription, Genetic; Tumor Cells, Cultured; Tumor Suppressor Protein p53

At the onset of the mineralization of bone, small membranous matrix vesicles are often observed. The information available on the production and release of these vesicles is limited. When treated with 10-20 nM of the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA), the human osteosarcoma cell line U-2 OS developed long cytoplasmic processes connecting adjacent cells. SEM and TEM show that TPA triggers a production and release of matrix vesicle-like membrane vesicles, mainly from the cellular processes. Tetracycline HCl was used to label intracellular bound calcium. The tetracycline HCl label was primarily localized to the end-feet of the cytoplasmic processes, indicating that these contain high concentrations of Ca2+, and to endoplasmic reticulum-like structures in the cell bodies. Together with our previous demonstration of the release of alkaline phosphatase-containing vesicles into the culture medium (Ringbom-Anderson T, Akerman KEO 1992 Calcif Tissue Int 50:533-540), the results presented here indicate that TPA induces a rapid induction of the primary steps of mineralization in U-2 OS osteosarcoma.

Topics: Actins; Calcification, Physiologic; Calcium; Cytoplasm; Endoplasmic Reticulum; Humans; Microscopy, Electron, Scanning Transmission; Microscopy, Fluorescence; Organelles; Osteoblasts; Osteosarcoma; Tetracycline; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured

Twelve patients with tumors and three with osteitis of the extremities were studied using 99mTc-labelled tetracycline in order to image the process with gamma camera. The diagnosis of all cases were verified histologically. The labelled tetracycline was found to give positive images for the most malignant tumors and active osteitis which together comprised about half of the studied cases.

Topics: Bone Neoplasms; Extremities; Humans; Osteitis; Osteosarcoma; Radionuclide Imaging; Soft Tissue Neoplasms; Technetium; Tetracycline; Tibia

A prospective study of 40 consecutive classic primary osteosarcomas employing tomography, angiography, radio-scanning, tetracycline labelling, macroscopic and microscopic study revealed that in 10 (25%) "skip" metastases were found. In 8 the "skips" were completely unsuspected prior to computation. It is evident that patients with "skips" are more prone to local recurrence and have a worse prognosis following amputation than those without such satellite lesions. Through-bone amputation entails a considered risk of leaving micro-foci of tumor; proximal disarticulation does not obviate the risk of cross joint "skips" when the lesion is in a sub-articular site. What affect such residual foci has upon the effectiveness of adjunct therapy and conversely whether such therapy will permit more conservative surgery by suppressing residual "skips" is an important but unanswered question.

Topics: Adolescent; Adult; Aged; Amputation, Surgical; Bone Neoplasms; Child; Child, Preschool; Female; Humans; Male; Middle Aged; Neoplasm Metastasis; Osteosarcoma; Prospective Studies; Tetracycline

Topics: Adenocarcinoma; Animals; Carcinoma, Bronchogenic; Carcinoma, Hepatocellular; Gallium; Glioblastoma; Hodgkin Disease; Humans; L-Lactate Dehydrogenase; Liver Neoplasms; Lung Neoplasms; Mediastinal Neoplasms; Methane; Mice; Muscular Diseases; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms, Experimental; Nitrosourea Compounds; Osteosarcoma; Rabbits; Radioisotopes; Radionuclide Imaging; Rats; Sarcoma; Sarcoma, Experimental; Technetium; Tetracycline; Transplantation, Homologous

Topics: Adolescent; Adult; Animals; Antigens, Neoplasm; Blood Proteins; Bone Neoplasms; Child; Child, Preschool; Epitopes; Female; Femoral Neoplasms; Fluoresceins; Humans; Immune Sera; Immunity, Active; Immunity, Maternally-Acquired; Immunization, Passive; Immunodiffusion; Immunoelectrophoresis; Immunotherapy; Infant; Leukocyte Count; Lung Neoplasms; Lymphocyte Activation; Lymphocytes; Male; Neoplasm Metastasis; Neoplasm Transplantation; Osteosarcoma; Rabbits; Skin Tests; Tetracycline; Tibia

Topics: Bone Neoplasms; Fluorescence; Humans; Neoplasms; Osteosarcoma; Sarcoma; Tetracycline

Topics: Bone and Bones; Bone Neoplasms; Fibrosarcoma; Humans; Neoplasms; Osteosarcoma; Tetracycline