cyclin-d1 and Pituitary-Neoplasms

In recent years, numerous studies have been performed to investigate the CCND1 G870A gene polymorphism impact on brain tumors susceptibility. Unfortunately, the results of previous studies were inconsistent. Therefore, we performed a meta-analysis to derive a more precise estimation of any association.. We conducted a search in PubMed, Embase and CNKI covering all published papers up to November, 2013. Odds ratios (ORs) and their 95% confidence intervals (95%CIs) were applied to assess associations.. A total of 6 publications including 9 case-control studies met the inclusion criteria. The pooled ORs for the total included studies showed significant association among comparison A vs G (OR= 1.246, 95%CI= 1.092-1.423, p= 0.001), homozygote comparison AA vs GG (OR= 1.566, 95%CI= 1.194-2.054, p= 0.001), heterozygote comparison AG vs GG (OR= 1.290, 95%CI= 0.934-1.782, p= 0.122), dominant model AA/GA vs GG (OR= 1.381, 95%CI= 1.048-1.821, p= 0.022) and recessive model AA vs GA/GG (OR= 1.323, 95%CI= 1.057- 1.657, p= 0.015) especially in glioma.. CCND1 G870A polymorphism may increase brain tumor risk, especially for gliomas. However, more primary large scale and well-designed studies are still required to evaluate the interaction of CCND1 G870A polymorphism with brain tumor risk.

Topics: Adenoma; Brain Neoplasms; Cyclin D1; Genetic Predisposition to Disease; Glioma; Humans; Meningioma; Neuroma, Acoustic; Odds Ratio; Pituitary Neoplasms; Polymorphism, Single Nucleotide

Pituitary cells are particularly sensitive to alterations of the cell cycle machinery. In fact, mutations affecting expression of proteins critical for cell cycle progression, including retinoblastoma protein, cyclins D1 and D3, p16(INK4A), and p27(kip1), are frequent in human pituitary adenomas. Similarly, both targeted disruption and overexpression of either cell cycle inhibitors or activators, respectively, lead to the development of pituitary adenomas in mice. Recent evidence has added the high mobility group A (HMGA) proteins as a new class of cell cycle regulators that play significant roles in the pathways that lead to pituitary tumor evolution in both humans and experimental animal models. Here, we first review the role of the cell cycle in pituitary tumorigenesis, as witnessed by human pathology and transgenic mice; and then, we focus on HMGA proteins and their cell cycle-related role in pituitary tumorigenesis.

Topics: Adenoma; Adult; Animals; Cell Cycle; Cell Cycle Proteins; Cyclin D1; Cyclin D3; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p27; Female; HMGA Proteins; Humans; Male; Mice; Mice, Transgenic; Pituitary Gland; Pituitary Neoplasms; Retinoblastoma Protein; Tumor Suppressor Proteins

Pituitary tumors constitute 10% of intracranial neoplasms and are mostly benign, monoclonal adenomas derived from single mutant cells. Pituitary oncogenes have been intensively studied and three of them, gsp, ccnd1, and PTTG are abundant in significant numbers of cases. gsp is present in approximately 40% of Caucasian patients with GH-secreting tumors and results from a mutated, constitutively active alpha subunit of Gs protein. Persistent activation of the cAMP-PKA-CREB pathway may lead to uncontrolled cell proliferation and GH secretion. ccnd1 is overexpressed cyclin D1, and cyclin D1 gene is amplified in some pituitary tumors. PTTG is expressed in most pituitary tumors. PTTG is localized to both the nucleus and cytoplasm and interacts with several protein partners. At least three tumorigenesis mechanisms are proposed for human PTTG. 1) PTTG and FGF form a positive feedback loop and stimulate tumor vascularity. 2) PTTG transactivates c-myc or other pro-proliferation genes. 3) PTTG overexpression causes aneuploidy. PTTG expression activates p53 and causes p53-dependent and -independent apoptosis. Due to lack of functional human pituitary cell cultures and appropriate animal models for pituitary tumors, many of the results reviewed here are obtained from heterologous systems.

Topics: Adenoma; Animals; Cell Cycle; Cell Division; Cloning, Molecular; Cyclin D1; Gene Expression Regulation, Neoplastic; GTP-Binding Protein alpha Subunits, Gs; Humans; Mice; Mice, Nude; Multigene Family; Neoplasm Proteins; Oncogenes; Organ Specificity; Pituitary Neoplasms; Rats; Securin; Signal Transduction; Species Specificity; Transcriptional Activation; Transfection

Topics: Adolescent; Adult; Aged; beta Catenin; Cell Cycle; Child; Child, Preschool; Craniopharyngioma; Cyclin D1; Female; Humans; In Situ Hybridization, Fluorescence; Ki-67 Antigen; Male; Middle Aged; Mutation; Pituitary Neoplasms; Prognosis; Proto-Oncogene Proteins B-raf; Receptor, trkA; Young Adult

Prolactinomas are the most frequent pituitary tumor subtype. Despite most of them respond to medical treatment, a proportion are resistant and become a challenge in clinical management. Wnt/β-Catenin pathway has been implicated in several cancers including pituitary tumors and other sellar region malignancies. Interestingly, Wnt/β-Catenin inhibition augments the cytotoxicity of the chemotherapeutic agent Temozolomide (TMZ) in different cancers. TMZ is now being implemented as rescue therapy for aggressive pituitary adenoma treatment. However, the molecular mechanisms associated with TMZ action in pituitary tumors remain unclear.. Our aims in the present study were to evaluate differential β-Catenin expression in human resistant prolactinomas and Wnt/β-Catenin signaling activation and involvement in Prolactin (PRL) secreting experimental models treated with TMZ.. We first evaluated by immunohistochemistry β-Catenin localization in human resistant prolactinomas in which we demonstrated reduced membrane β-Catenin in prolactinoma cells compared to normal pituitaries, independently of the Ki-67 proliferation indexes. In turn, in vivo 15 mg/kg of orally administered TMZ markedly reduced PRL production and increased prolactinoma cell apoptosis in mice bearing xenografted prolactinomas. Intratumoral β-Catenin strongly correlated with Prl and Cyclin D1, and importantly, TMZ downregulated both β-Catenin and Cyclin D1, supporting their significance in prolactinoma growth and as candidates of therapeutic targets. When tested in vitro, TMZ directly reduced MMQ cell viability, increased apoptosis and produced G2/M cell cycle arrest. Remarkably, β-Catenin activation and VEGF secretion were inhibited by TMZ in vitro.. We concluded that dopamine resistant prolactinomas undergo a β-Catenin relocalization in relation to normal pituitaries and that TMZ restrains experimental prolactinoma tumorigenicity by reducing PRL production and β-Catenin activation. Together, our findings contribute to the understanding of Wnt/β-Catenin implication in prolactinoma maintenance and TMZ therapy, opening the opportunity of new treatment strategies for aggressive and resistant pituitary tumors.

Topics: Animals; beta Catenin; Cyclin D1; Humans; Mice; Models, Theoretical; Pituitary Neoplasms; Prolactin; Prolactinoma; Temozolomide

As hyperprolactinemia is observed in patients with bromocriptine‑resistant prolactinoma, prolactin (PRL) has been implicated in the development of bromocriptine resistance. Since PRL primarily mediates cell survival and drug resistance via the Janus kinase‑2 (JAK2)/signal transducer and activator of transcription 5A (STAT5) signaling pathway, the STAT5 inhibitor, pimozide, may inhibit cell proliferation and reverse bromocriptine resistance in prolactinoma cells. In the present study, compared with bromocriptine or pimozide alone, the combination of pimozide and bromocriptine exerted enhanced reduction in cell growth and proliferation, and increased apoptosis and cell cycle arrest in bromocriptine‑resistant prolactinoma cells. A reduction in phospho‑STAT5, cyclin D1 and B‑cell lymphoma extra‑large (Bcl‑xL) expression levels were observed in cells treated with the combination of drugs. In addition, pimozide suppressed spheroid formation of human pituitary adenoma stem‑like cells, and reduced the protein expression of the cancer stem cell markers, CD133 and nestin. Pimozide did not exert any additional antitumor activity in STAT5‑knockdown primary culture cells of human bromocriptine‑resistant prolactinomas. Furthermore, Pimozide combined with bromocriptine treatment significantly reduced human prolactinoma xenograft growth. Western blot and immunohistochemical analyses also demonstrated significant inhibition of cell proliferation and stem cell marker proteins in vivo. Collectively, these data indicated that pimozide treatment reduced prolactinoma growth by targeting both proliferating cells and stem cells, at least in part, by inhibiting the STAT5/Bcl‑xL and STAT5/cyclin D1 signaling pathways.

Topics: Animals; bcl-X Protein; Bromocriptine; Cell Line, Tumor; Cyclin D1; Humans; Mice; Mice, Nude; Pimozide; Pituitary Neoplasms; Prolactinoma; Rats; Signal Transduction; STAT5 Transcription Factor; Xenograft Model Antitumor Assays

Prolactinomas are the most common type of functional pituitary adenoma. Although bromocriptine is the preferred first line treatment for prolactinoma, resistance frequently occurs, posing a prominent clinical challenge. Both the prolactin receptor (PRLR) and estrogen receptor α (ERα) serve critical roles in the development and progression of prolactinomas, and whether this interaction between PRLR and ERα contributes to bromocriptine resistance remains to be clarified. In the present study, increased levels of ERα and PRLR protein expression were detected in bromocriptine-resistant prolactinomas and MMQ cells. Prolactin (PRL) and estradiol (E2) were found to exert synergistic effects on prolactinoma cell proliferation. Furthermore, PRL induced the phosphorylation of ERα via the JAK2-PI3K/Akt-MEK/ERK pathway, while estrogen promoted PRLR upregulation via pERα. ERα inhibition abolished E2-induced PRLR upregulation and PRL-induced ERα phosphorylation, and fulvestrant, an ERα inhibitor, restored pituitary adenoma cell sensitivity to bromocriptine by activating JNK-MEK/ERK-p38 MAPK signaling and cyclin D1 downregulation. Collectively, these data suggest that the interaction between the estrogen/ERα and PRL/PRLR pathways may contribute to bromocriptine resistance, and therefore, that combination treatment with fulvestrant and bromocriptine (as opposed to either drug alone) may exert potent antitumor effects on bromocriptine-resistant prolactinomas.

Topics: Adolescent; Adult; Animals; Antineoplastic Combined Chemotherapy Protocols; Bromocriptine; Cell Line, Tumor; Cyclin D1; Drug Resistance, Neoplasm; Estradiol; Estrogen Receptor alpha; Female; Fulvestrant; Humans; Hypophysectomy; Male; MAP Kinase Signaling System; Middle Aged; Neoplasm Recurrence, Local; Pituitary Gland; Pituitary Neoplasms; Prolactin; Prolactinoma; Rats; Receptors, Prolactin; Young Adult

BACKGROUND As a kind of benign tumor, pituitary adenomas have attracted increasing attention from researchers. The plasmacytoma variant translocation 1 (PVT1) is a molecule in the lncRNA family protein that has been proven to play critical roles in many cancers; however, no study has explored the special biological roles of PVT1 in pituitary adenoma. MATERIAL AND METHODS The qRT-PCR assay was conducted to evaluate PVT1 expressions in various cell lines and tissues. Loss of function assays were carried out to detect the influence of silenced PVT1 on the proliferation, migration, and epithelial-mesenchymal transition (EMT) of pituitary adenoma cells. Western blotting was used to identify correlation between ß-catenin and PVT1. RESULTS The PVT1 expressions were significantly enhanced in tissues of pituitary adenoma and cancer cells. Cell migration and proliferation were inhibited when the PVT1 gene was knocked down. Knockdown of PVT1 repressed the migration and EMT of pituitary adenoma cells. The PVT1 downregulation obviously blocked Wnt/ß-catenin signaling pathway activity. PVT1 aggravated progression of pituitary adenoma through initiating the Wnt/ß-catenin signaling pathway. CONCLUSIONS PVT1 exerts an oncogenic role through activating Wnt/ß-catenin signaling in pituitary adenoma cells. The present results may provide a potential therapeutic target or approach for treating pituitary adenomas.

Topics: Adenoma; beta Catenin; Biomarkers, Tumor; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Pituitary Neoplasms; Proto-Oncogene Proteins c-myc; RNA, Long Noncoding; RNA, Small Interfering; Wnt Signaling Pathway

The pathogenesis of pituitary adenomas (PA) is complex. Ki-67, pituitary tumour transforming gene (PTTG), vascular endothelial growth factor (VEGF), cyclin D1, c-MYC and pituitary adenylate cyclase-activating peptide (PACAP) protein expression were analysed and correlated with tumour and patient characteristics.. 74 pituitary tumour samples (48 non-functional PA, 26 functional PAs); Immunohistochemical analysis of protein expression, retrospective analysis of MR images and in vitro analysis of octreotide treatment was carried out on GH3 cells.. PTTG expression was negatively associated with age and positively with PA size, regrowth and Ki-67 index. Cyclin D1 correlated with Ki-67 and tumour size. c-MYC negatively correlated with size of tumour and age; and correlated with PTTG expression. Somatostatin analogue treatment was associated with lower Ki-67, PTTG and Cyclin D1 expression while T2 hypointense PAs were associated with lower PTTG, cyclin D1, c-MYC and Ki-67. In vitro analyses confirmed the effect of somatostatin analogue treatment on Pttg and Cyclin D1 expression.. Interesting and novel observations on the differences in expression of tumour markers studied are reported. Correlation between Ki-67 expression, PTTG nuclear expression and recurrence/regrowth of PAs, emphasizes the role that Ki-67 and PTTG expression have as markers of increased proliferation. c-MYC and PTTG nuclear expression levels were correlated providing evidence that PTTG induces c-MYC expression in PAs and we propose that c-MYC might principally have a role in early pituitary tumorigenesis. Evidence is shown that the anti-proliferative effect of somatostatin analogue treatment in vivo occurs through regulation of the cell cycle.

Topics: Biomarkers; Cell Cycle; Cell Proliferation; Cyclin D1; Humans; Ki-67 Antigen; Pituitary Adenylate Cyclase-Activating Polypeptide; Pituitary Neoplasms; Retrospective Studies; Securin; Vascular Endothelial Growth Factor A

Considering that the role of ERβ in the growth of pituitary cells is not well known, the aim of this work was to determine the expression of ERβ in normal and tumoral cells and to investigate its implications in the proliferative control of this endocrine gland, by analyzing the participation of cyclin D1, Cdk4 and p21. Our results showed that the expression of ERβ decreased during pituitary tumoral development induced by chronic E2 stimulation. The 20 ± 1.6% of normal adenohypophyseal cells expressed ERβ, with this protein being reduced in the hyperplastic/adenomatous pituitary: at 20 days the ERβ+ population was 10.7 ± 2.2%, while after 40 and 60 days of treatment an almost complete loss in the ERβ expression was observed (40 d: 1 ± 0.6%; 60 d: 2 ± 0.6%). The ERα/β ratio increased starting from tumors at 40 days, mainly due to the loss of ERβ expression. The cell proliferation was analyzed in normal and hyperplastic pituitary and also in GH3β- and GH3β+ which contained different levels of ERβ expression, and therefore different ERα/β ratios. The over-expression of ERβ inhibited the GH3 cell proliferation and expression of cyclin D1 and ERα. Also, the ERβ activation by its agonist DPN changed the subcellular localization of p21, inducing an increase in the p21 nuclear expression, where it acts as a tumoral suppressor. These results show that ERβ exerts an inhibitory role on pituitary cell proliferation, and that this effect may be partially due to the modulation of some key regulators of the cell cycle, such as cyclin D1 and p21. These data contribute significantly to the understanding of the ER effects in the proliferative control of pituitary gland, specifically related to the ERβ function in the E2 actions on this endocrine gland.

Topics: Animals; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cells, Cultured; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Estradiol; Estrogen Receptor beta; Female; Gene Expression Regulation; NAD; Pituitary Gland, Anterior; Pituitary Neoplasms; Rats

In the pituitary the activation of cyclic adenosine 3'-5'-monophosphate (cAMP) dependent pathways generates proliferative signals in somatotrophs, whereas in pituitary cells of other lineages its effect remains uncertain. Moreover, the specific role of the two main cAMP effectors, protein kinase A (PKA) and exchange proteins directly activated by cAMP (Epac), has not been defined. Aim of this study was to investigate the effect of cAMP on pituitary adenomatous cells proliferation and to identify PKA and Epac differential involvement. We found that cAMP increased DNA synthesis and cyclin D1 expression in somatotropinomas, whereas it reduced both parameters in prolactinomas and nonfunctioning adenomas, these effects being replicated in corresponding cell lines. Moreover, the divergent cAMP effects were mimicked by Epac and PKA analogs, which activated Rap1 and CREB, respectively. In conclusion, we demonstrated that cAMP exerted opposite effects on different pituitary cell types proliferation, these effects being mediated by both Epac and PKA.

Topics: Adenoma; Animals; Cell Proliferation; Cyclic AMP; Cyclic AMP Response Element-Binding Protein; Cyclic AMP-Dependent Protein Kinases; Cyclin D1; Gene Expression Regulation, Neoplastic; Gonadotrophs; Guanine Nucleotide Exchange Factors; Humans; Lactotrophs; Pituitary Gland; Pituitary Neoplasms; Prolactinoma; Protein Subunits; rap1 GTP-Binding Proteins; Rats; Signal Transduction; Somatotrophs

Insulin-like growth factor (IGF1) and its receptor display potent proliferative and antiapoptotic activities and are considered key players in malignancy. The objective of the study was to explore the role of IGF1 and its downstream pathways in the proliferation of non-functioning pituitary tumor cells and to develop a targeted therapeutic approach for the treatment of these tumors. Cultures of human non-functioning pituitary adenomas and the non-secreting immortalized rat pituitary tumor cell line MtT/E were incubated with IGF1, IGF1 receptor inhibitor or both, and cell viability, proliferation and signaling were examined. Our results show that IGF1 elevated cell proliferation and enhanced cell cycle progression as well as the expression of cyclins D1 and D3. IGF1 also induced the phosphorylation of ERK, Akt and p70S6K. On the other hand, the selective IGF1R inhibitor NVP-AEW541 abrogated IGF1-induced cell proliferation as well as IGF1 receptor phosphorylation and downstream signaling.

Topics: Adenoma; Adult; Aged; Aged, 80 and over; Animals; Antineoplastic Agents; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin D3; Female; Humans; Insulin-Like Growth Factor I; Male; Middle Aged; Pituitary Neoplasms; Protein Processing, Post-Translational; Pyrimidines; Pyrroles; Rats

Smad nuclear interacting protein 1 (SNIP1) gene encodes a protein that contains a conservative C-terminal forkhead-associated domain and functions as a transcriptional coactivator to regulate cell proliferation and cancer progression. This study aimed to investigate the clinical and biological significance of SNIP1 expression in pituitary adenoma. We analyzed SNIP1 expressions in mouse fibroblast L929 cells and mouse pituitary adenoma AtT-20 cells by Western blotting. SNIP1 gene knockdown by small interfering RNA (siRNA) transfection was performed to evaluate SNIP1 function in pituitary adenoma cell lines. As expected, SNIP1 was found to be upregulated in pituitary adenoma cells. The mRNA and protein levels of SNIP1 were inhibited in AtT-20 cells transfected with siRNAs, which led to decreased proliferation, increased apoptosis, and cycle arrest of pituitary adenoma cells. Concomitantly, c-Myc and cyclin D1 protein levels were reduced. These findings may provide novel targets for the treatment of pituitary adenoma.

Topics: Adenoma; Animals; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclin D1; G1 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Intracellular Signaling Peptides and Proteins; Mice; Pituitary Neoplasms; Proto-Oncogene Proteins c-myc; RNA Interference; RNA-Binding Proteins; RNA, Messenger; RNA, Small Interfering; Up-Regulation

Deregulation of the Wnt pathway has been implicated in oncogenesis of numerous tissues including the pituitary gland. Immunohistochemical localization and quantification of β-catenin, Cyclin D1, c-MYC and Survivin expression in 47 pituitary adenomas (35 non-functioning, seven GH-secreting, three prolactinomas, two ACTH-secreting tumour) and six normal controls was undertaken in this study and correlation of protein expression to patient and tumour characteristics analysed. β-catenin was strictly membrane-bound with no difference observed between normal and tumour tissue. In contrast, Cyclin D1 and c-MYC localization was nuclear and significantly higher in tumour versus normal tissue (p < 0.05). c-MYC expression correlated negatively with age at diagnosis (p = 0.006, R = -0.395) while Cyclin D1 expression correlated positively with age (p = 0.036, R = 0.306) and was higher in males than in females (p = 0.036). c-MYC expression was significantly lower in patients with functional tumours requiring octreotide treatment and in patients with non-functioning tumours suffering from hypopituitarism. Survivin expression was extremely low in tumours and absent in normal controls. Involvement of the canonical Wnt pathway appears to be minimal, given the segregation of β-catenin to the membrane. Our data suggest that c-MYC may have an important role in early pituitary tumorigenesis while Cyclin D1 is likely to promote tumour growth at a later stage. We also report a novel gender difference in Cyclin D1 expression, the biological significance of which merits further analysis. The reported reduction of c-MYC in functional tumours subsequently treated with octreotide further supports a role of c-MYC in early tumorigenesis and not in recurrence. The decrease in c-MYC in patients with hypopituitarism provides the first in vivo evidence for hormonal regulation of c-MYC expression.

Topics: Adult; Aged; Antineoplastic Agents, Hormonal; beta Catenin; Biomarkers, Tumor; Combined Modality Therapy; Cyclin D1; Female; Humans; Hypopituitarism; Inhibitor of Apoptosis Proteins; Male; Middle Aged; Octreotide; Pituitary Neoplasms; Prolactinoma; Proto-Oncogene Proteins c-myc; Sex Factors; Survivin; Wnt Proteins; Wnt Signaling Pathway; Young Adult

The objective of this study was to investigate the effect of G870A gene polymorphism of CCND1 on the formation and behavioral features of prolactinomas. One hundred and thirteen patients with prolactinoma and 108 age and gender matched control were included in the study. The patients were divided into two groups as noninvasive and invasive tumors. CCND1 G870A gene polymorphism was compared in patients/control and invasive/noninvasive groups. A and G allele frequencies were found as 41.7% and 58.3% in the controls, and 61.1% and 38.9% in the patients (p<0.01). Rates of G/G, G/A and A/A genotypes were found as 11.8%, 55.9% and 32.4% in the noninvasive group, and 15.6%, 44.4% and 40.0% in the invasive group, respectively. Differences between patient and control groups were significant but were not between invasive and noninvasive groups in terms of the allele frequencies and genotype distribution. Mean tumor size and serum levels of prolactin at the time of diagnosis and change in these values after the treatment were not found statistically significant in genotype subgroups. CCND1 G870A gene polymorphism may be an important factor in the early stages of the tumor formation. However, it did not affect the features of the tumor.

Topics: Adolescent; Adult; Aged; Base Sequence; Case-Control Studies; Cross-Sectional Studies; Cyclin D1; DNA Primers; Female; Gene Frequency; Humans; Magnetic Resonance Imaging; Male; Middle Aged; Neoplasm Invasiveness; Pituitary Neoplasms; Polymorphism, Single Nucleotide; Prolactinoma; Proto-Oncogenes; Retrospective Studies; Young Adult

Pituitary tumors are a common form of endocrine neoplasia. However few studies assessed the expression of the principal cyclin regulating checkpoint exit, cyclin D1. Cyclin D1 expression in pituitary tumors and its possible relation to MIB-1 and p27/Kip1 labeling indices (LIs) was explored.. We studied a total of 199 pituitaries, including normal pituitaries (n=7), pituitary adenomas (n=187), and pituitary carcinoma (n=5). All tissues were tested as cores of archived tissue microarrays that were immunostained for cyclin D1, MIB-1 and p27 using a standard technique. Tissue cores were subjected to automated analysis to evaluate the staining LIs.. No cyclin D1 positive cells in the normal anterior pituitary gland was found. Sparse nuclear staining was noted in pituitary tumors. Higher expression of cyclin D1 was noted in pituitary carcinomas compared to adenomas (p<0.001), in non-functioning adenomas compared to functioning ones (p<0.001) in macroadenomas versus microadenomas (p=0.017) and in recurrent non recurrent adenomas (p<0.001). Cyclin D1 LI and MIB-1 LI were related among adenomas (p<0.001) and carcinomas (p=0.041). p27 LI was neither related to pituitary adenoma recurrence nor invasion.. Expression of cyclin D1 in pituitary tumors is related to cell proliferation, recurrence, and metastatic potential. Nuclear cyclin D1 expression is a good marker of aggressive behavior in pituitary tumors.

Topics: Adenoma; Adult; Carcinoma; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Female; Gene Expression; Humans; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Pituitary Neoplasms; ROC Curve; Tissue Array Analysis; Ubiquitin-Protein Ligases

The thyroid hormone, T3, plays important roles in metabolism, growth, and differentiation. Germline mutations in thyroid hormone receptor beta (TRbeta) have been identified in many individuals with resistance to thyroid hormone, a syndrome of reduced sensitivity to T3. A close association of somatic mutations of TRbeta with several human cancers has become increasingly apparent, but how TRbeta mutants could be involved in the carcinogenesis in vivo has not been addressed. The creation of a mouse model (TRbeta(PV/PV) mouse) that harbors a knockin mutation of TRbeta (denoted TRbetaPV) has facilitated the study of the molecular actions of TRbeta mutants in vivo. The striking phenotype of thyroid cancer and the development of pituitary tumors exhibited by TRbeta(PV/PV) mice have uncovered novel functions of a TRbeta mutant in tumorigenesis. It led to the important findings that the oncogenic action of TRbetaPV is mediated by both genomic and nongenomic actions to alter gene expression and signaling pathways activity.

Topics: Animals; Cyclin D1; Gene Expression Regulation, Neoplastic; Mice; Mice, Mutant Strains; Mutation; Pituitary Neoplasms; Signal Transduction; Thyroid Hormone Receptors beta; Thyroid Neoplasms

Raf/MEK/ERK and phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) cascades are key signalling pathways interacting with each other to regulate cell growth and tumourigenesis. We have previously shown B-Raf and Akt overexpression and/or overactivation in pituitary adenomas. The aim of this study is to assess the expression of their downstream components (MEK1/2, ERK1/2, mTOR, TSC2, p70S6K) and effectors (c-MYC and CYCLIN D1). We studied tissue from 16 non-functioning pituitary adenomas (NFPAs), six GH-omas, six prolactinomas and six ACTH-omas, all collected at transsphenoidal surgery; 16 normal autopsy pituitaries were used as controls. The expression of phospho and total protein was assessed with western immunoblotting, and the mRNA expression with quantitative RT-PCR. The expression of pSer217/221 MEK1/2 and pThr183 ERK1/2 (but not total MEK1/2 or ERK1/2) was significantly higher in all tumour subtypes in comparison to normal pituitaries. There was no difference in the expression of phosphorylated/total mTOR, TSC2 or p70S6K between pituitary adenomas and controls. Neither c-MYC phosphorylation at Ser 62 nor total c-MYC was changed in the tumours. However, c-MYC phosphorylation at Thr58/Ser62 (a response target for Akt) was decreased in all tumour types. CYCLIN D1 expression was higher only in NFPAs. The mRNA expression of MEK1, MEK2, ERK1, ERK2, c-MYC and CCND1 was similar in all groups. Our data indicate that in pituitary adenomas both the Raf/MEK/ERK and PI3K/Akt/mTOR pathways are upregulated in their initial cascade, implicating a pro-proliferative signal derangement upstream to their point of convergence. However, we speculate that other processes, such as senescence, attenuate the changes downstream in these benign tumours.

Topics: Adult; Aged; Blotting, Western; Case-Control Studies; Cell Proliferation; Cyclin D1; Female; Humans; Immunoenzyme Techniques; Intracellular Signaling Peptides and Proteins; Male; MAP Kinase Kinase Kinases; Middle Aged; Mitogen-Activated Protein Kinase 3; Phosphatidylinositol 3-Kinases; Phosphorylation; Pituitary Neoplasms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-myc; raf Kinases; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; TOR Serine-Threonine Kinases; Young Adult

Curcumin (diferuloylmethane) is the active ingredient of the spice plant Curcuma longa and has been shown to act anti-tumorigenic in different types of tumours. Therefore, we have studied its effect in pituitary tumour cell lines and adenomas. Proliferation of lactosomatotroph GH3 and somatotroph MtT/S rat pituitary cells as well as of corticotroph AtT20 mouse pituitary cells was inhibited by curcumin in monolayer cell culture and in colony formation assay in soft agar. Fluorescence-activated cell sorting (FACS) analysis demonstrated curcumin-induced cell cycle arrest at G2/M. Analysis of cell cycle proteins by immunoblotting showed reduction in cyclin D(1), cyclin-dependent kinase 4 and no change in p27(kip). FACS analysis with Annexin V-FITC/7-aminoactinomycin D staining demonstrated curcumin-induced early apoptosis after 3, 6, 12 and 24 h treatment and nearly no necrosis. Induction of DNA fragmentation, reduction of Bcl-2 and enhancement of cleaved caspase-3 further confirmed induction of apoptosis by curcumin. Growth of GH3 tumours in athymic nude mice was suppressed by curcumin in vivo. In endocrine pituitary tumour cell lines, GH, ACTH and prolactin production were inhibited by curcumin. Studies in 25 human pituitary adenoma cell cultures have confirmed the anti-tumorigenic and hormone-suppressive effects of curcumin. Altogether, the results described in this report suggest this natural compound as a good candidate for therapeutic use on pituitary tumours.

Topics: Animals; Antineoplastic Agents; Apoptosis; Blotting, Western; Caspase 3; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Curcumin; Cyclin D1; Cyclin-Dependent Kinase 4; Flow Cytometry; Humans; Male; Mice; Mice, Nude; Pituitary Hormones; Pituitary Neoplasms; Rats

The etiology of sporadic pituitary tumors is currently unknown. The Wnt pathways have been implicated in the pathogenesis of a variety of human tumors, but the role of these pathways in pituitary tumors is unclear. Microarray analysis using the Affymetrix HG U133 plus 2.0 GeneChips identified four secreted frizzled-related protein (sFRP) family members of Wnt pathway inhibitors that were differentially expressed in both nonfunctioning and clinically functioning pituitary tumors (n = 20) compared with normal pituitary controls (n = 3). Reduced tumor expression of Wnt inhibitory factor-1 (WIF1), sFRP2, and sFRP4 mRNA was confirmed by real-time quantitative RT-PCR (P <0.001 and P = 0.002 and 0.013, respectively) in all pituitary subtypes. Hypermethylation of the WIF1 promoter was present in 88% of the pituitary tumors (n = 41). Seventy-six percent of pituitary tumors demonstrated absent or weak cytoplasmic WIF1 staining by immunohistochemistry (n = 41), although preserved staining was seen in some functioning tumors, with strong staining in 92% of normal pituitary controls (n = 13). The Wnt pathway target gene cyclin D1 was found to be up-regulated specifically in the nonfunctioning pituitary tumors compared with controls at both mRNA and protein level, supportive of activation of the Wnt-beta-catenin pathway. Nuclear accumulation of beta-catenin, however, was not observed in any pituitary tumors (n = 70). By transfecting GH3 cells with WIF1, decreased cell proliferation and colony formation was observed compared with empty vector controls. In conclusion, our data suggest that WIF1 may be a tumor suppressor, specifically in nonfunctioning pituitary tumors, and that the Wnt pathways are important in pituitary tumorigenesis.

Topics: Adaptor Proteins, Signal Transducing; Adult; Aged; Aged, 80 and over; Animals; beta Catenin; Case-Control Studies; Cell Line; Cell Proliferation; Cyclin D1; Down-Regulation; Extracellular Matrix Proteins; Female; Gene Expression Regulation, Neoplastic; Glycoproteins; Humans; Intercellular Signaling Peptides and Proteins; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Middle Aged; Pituitary Neoplasms; Proto-Oncogene Proteins; Rats; Repressor Proteins; Somatotrophs; Transfection

The cyclin D1 gene (CCND1) is a proto-oncogene playing a critical role in the transition through the G1 to the S phase of the cell cycle and is overexpressed in many tumors. G870A polymorphism at the exon4/intron4 splicing region of the CCND1 gene may play a role in pituitary tumorigenesis and invasiveness. The objective of this study was to examine CCND1 polymorphism in patients with different types of sporadic pituitary adenomas.. One hundred thirty patients (38 male, 92 female, mean age: 45.37+/-13.55 SD years) with sporadic pituitary adenomas (PA group) and 129 healthy controls (HC group) were included in the study. The CCND1 G870A polymorphism in PA and HC were genotyped by PCR-RFLP using peripheral blood samples. CCND1 expression was also evaluated with an immunohistochemical method in tumor tissues of 39 patients of the PA group.. The genotype distribution in the PA [AA: 30 (23.1%), AG: 90 (69.2%), GG: 10 (7.7%)] was statistically different from the HC group [AA: 36 (27.9%), AG: 64 (49.6%), GG: 29 (22.5%), p=0.001]. Patients carrying the AG genotype were more frequent compared with the control group. Tumor type, volume, and invasion were not related to the genotype. Immunohistochemically, 21 of the 39 tumors showed nuclear positivity for CCND1, varying between 1 and 40% of tumor cells. Positive staining was observed more intense in patients carrying the AG genotype.. CCND1 polymorphism may be an early event in tumorigenesis, but it is not a reliable prognostic criterion.

Topics: Case-Control Studies; Cyclin D1; Female; Gene Frequency; Genotype; Humans; Immunohistochemistry; Male; Middle Aged; Pituitary Neoplasms; Proto-Oncogene Mas

Topics: Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; DNA, Neoplasm; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Proteins; Oncogene Proteins; Pituitary Neoplasms; Securin; Tumor Suppressor Proteins; World Health Organization

Thyroid-stimulating hormone (TSH)-secreting tumors (TSH-omas) are pituitary tumors that constitutively secrete TSH. The molecular genetics underlying this abnormality are not known. We discovered that a knock-in mouse harboring a mutated thyroid hormone receptor (TR) beta (PV; TRbeta(PV/PV) mouse) spontaneously developed TSH-omas. TRbeta(PV/PV) mice lost the negative feedback regulation with highly elevated TSH levels associated with increased thyroid hormone levels (3,3',5-triiodo-l-thyronine [T3]). Remarkably, we found that mice deficient in all TRs (TRalpha1(-/-) TRbeta(-/-)) had similarly increased T3 and TSH levels, but no discernible TSH-omas, indicating that the dysregulation of the pituitary-thyroid axis alone is not sufficient to induce TSH-omas. Comparison of gene expression profiles by cDNA microarrays identified overexpression of cyclin D1 mRNA in TRbeta(PV/PV) but not in TRalpha1(-/-) TRbeta(-/-) mice. Overexpression of cyclin D1 protein led to activation of the cyclin D1/cyclin-dependent kinase/retinoblastoma protein/E2F pathway only in TRbeta(PV/PV) mice. The liganded TRbeta repressed cyclin D1 expression via tethering to the cyclin D1 promoter through binding to the cyclic AMP response element-binding protein. That repression effect was lost in mutant PV, thereby resulting in constitutive activation of cyclin D1 in TRbeta(PV/PV) mice. The present study revealed a novel molecular mechanism by which an unliganded TRbeta mutant acts to contribute to pituitary tumorigenesis in vivo and provided mechanistic insights into the understanding of pathogenesis of TSH-omas in patients.

Topics: Animals; Blotting, Western; Cell Cycle Proteins; Cell Proliferation; Cyclic AMP; Cyclin D1; Cyclin-Dependent Kinases; DNA-Binding Proteins; DNA, Complementary; E2F Transcription Factors; Gene Deletion; Gene Expression Regulation; Glutathione Transferase; Immunoprecipitation; Ligands; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutation; Nucleic Acid Hybridization; Oligonucleotide Array Sequence Analysis; Organ Size; Pituitary Gland; Pituitary Hormones; Pituitary Neoplasms; Plasmids; Promoter Regions, Genetic; Protein Binding; Receptors, Thyroid Hormone; Retinoblastoma Protein; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Messenger; Thyroid Hormone Receptors beta; Time Factors; Transcription Factors; Transcriptional Activation; Transfection; Triiodothyronine

Alterations in cAMP signaling have been identified as a cause of endocrine neoplasia. In particular, activating mutations of the G(s)alpha gene and protein kinase A (PKA) overactivity due to low expression of PKA regulatory subunit 1A (R1A) have been implicated in somatotroph proliferation.. The objective of this study was to evaluate the effects of cAMP-PKA cascade activation in nonfunctioning pituitary adenomas (NFPA).. By immunohistochemistry, R1A, R2A, and R2B expression was evaluated in cells obtained from eight surgically removed NFPA positive for gonadotropins. Cyclin D1 expression and ERK1/2 activity were analyzed under basal conditions and after cAMP-PKA cascade activation.. Immunohistochemistry studies demonstrated a low R1/R2 ratio in all NFPA. Additional unbalance of R1/R2 ratio by 8-chloroadenosine cAMP (8-Cl-cAMP) and direct adenylyl cyclase stimulation by forskolin did not increase cyclin D1 expression or ERK1/2 activity in five NFPA (group 1), but even caused 74 +/- 15% and 85 +/- 13% inhibitions of cyclin D1 and ERK1/2 activity, respectively, in the remaining NFPA (group 2). Moreover, in group 2, PKA blockade by the specific inhibitor PKI increased cyclin D1 expression (96 +/- 25% over basal) and ERK1/2 activity (116 +/- 28% over basal).. These data show that in contrast with what was previously observed in transformed somatotrophs, activation of the cAMP-PKA pathway did not generate proliferative signals in tumoral cells of the gonadotroph lineage, and in a subset of tumors even exerted a tonic inhibitory effect, thus confirming a different role for the cAMP-mediated pathway in promoting proliferation in the pituitary.

Topics: Adenoma; Base Sequence; Biomarkers, Tumor; Cell Proliferation; Cyclic AMP; Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; Cyclic AMP-Dependent Protein Kinases; Cyclin D1; Enzyme Activation; Humans; In Vitro Techniques; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Pituitary Neoplasms; Proteins; Receptors, Cell Surface

The two regulatory subunits (R1 and R2) of protein kinase A (PKA) are differentially expressed in cancer cell lines and exert diverse roles in growth control. Recently, mutations of the PKA regulatory subunit 1A gene (PRKAR1A) have been identified in patients with Carney complex. The aim of this study was to evaluate the expression of the PKA regulatory subunits R1A, R2A, and R2B in a series of 30 pituitary adenomas and the effects of subunit activation on cell proliferation. In these tumors, neither mutation of PRKAR1A nor loss of heterozygosity was identified. By real-time PCR, mRNA of the three subunits was detected in all of the tumors, R1A being the most represented in the majority of samples. By contrast, immunohistochemistry documented low or absent R1A levels in all tumors, whereas R2A and R2B were highly expressed, thus resulting in an unbalanced R1/R2 ratio. The low levels of R1A were, at least in part, due to proteasome-mediated degradation. The effect of the R1/R2 ratio on proliferation was assessed in GH3 cells, which showed a similar unbalanced pattern of R subunits expression, and in growth hormone-secreting adenomas. The R2-selective cAMP analog 8-Cl cAMP and R1A RNA silencing, stimulated cell proliferation and increased Cyclin D1 expression, respectively, in human and rat adenomatous somatotrophs. These data show that a low R1/R2 ratio promoted proliferation of transformed somatotrophs and are consistent with the Carney complex model in which R1A inactivating mutations further unbalance this ratio in favor of R2 subunits. These results suggest that low expression of R1A protein may favor cAMP-dependent proliferation of transformed somatotrophs.

Topics: Adenoma; Cell Growth Processes; Cyclic AMP; Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; Cyclic AMP-Dependent Protein Kinases; Cyclin D1; Enzyme Activation; Exons; Humans; Immunohistochemistry; Introns; Lysosomes; Pituitary Neoplasms; Proteasome Inhibitors; Protein Subunits; Proteins; Reverse Transcriptase Polymerase Chain Reaction; Tumor Cells, Cultured

The G protein-coupled receptor (GPCR) activation has been demonstrated to affect the ERK1/2 cascade in different cell lines. We investigated the effects of hypothalamic neuropeptides acting via GPCR on this pathway in GH-secreting (GH-oma) and nonsecreting (NFPA) pituitary adenomas. GHRH increased ERK1/2 activity (236 +/- 80%) in both gsp- and gsp+ GH-omas, this effect being almost completely abolished by protein kinase C (PKC) blockade. Both GnRH and pituitary adenylate-activating peptide caused a similar PKC-dependent activation of ERK1/2 in most NFPA. Increasing cAMP by forskolin caused a protein kinase A-dependent increase of ERK activity (287 +/- 37%) in GH-omas and had no effect in NFPA. ERK cascade blockade in GH-omas did not affect basal and GHRH-stimulated GH release, whereas it totally prevented the 3-fold increase in cyclin D1 protein expression induced by GHRH. In conclusion, this study demonstrated that in pituitary adenomas the activation of GPCR by neurohormones caused a PKC-dependent activation of ERK1/2 cascade that, at least in GH-omas, resulted to be involved in cyclin D1 induction by GHRH. Moreover, a stimulatory effect of the protein kinase A-dependent pathway on ERK1/2 cascade occurred selectively in GH-omas, probably contributing to the mitogenic potential of the cAMP pathway in this cell type.

Topics: Adenoma; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclin D1; Enzyme Inhibitors; Growth Hormone; Growth Hormone-Releasing Hormone; Heterotrimeric GTP-Binding Proteins; Humans; Hypothalamus; MAP Kinase Kinase 1; MAP Kinase Kinase 2; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Neuropeptides; Phosphorylation; Pituitary Neoplasms; Protein Kinase C; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Tumor Cells, Cultured

Components of the pRb/p16/cyclin D1/CDK4 pathway are frequent targets in numerous tumour types, including those of pituitary origin. However, previous studies of pituitary tumours have examined individual components of this pathway. Therefore, to determine their overall contribution we have simultaneously examined the immunohistochemical status of pRb, p16 and cyclin D1 and analysed the CDK4 gene for a characterized activating mutation. Of the total pituitary tumour cohort (29 clinically non-functioning adenomas and 16 somatotrophinomas) abnormal expression of either pRb, p16 or cyclin D1 was observed in 36 of 45 (80%) tumours and was significantly (P = 0.005) associated with non-functioning tumours (27/29; 93%) compared with somatotrophinomas (9/16, 56%). Loss of either pRb or p16 expression was mutually exclusive in 23 of 45 (51%) tumours, whilst concomitant loss of pRb and p16 expression was observed in five tumours. Cyclin D1 overexpression was observed in 22 of 45 (49%) tumours, however, there was no significant association between overexpression of cyclin D1 and the expression status of either pRb or p16. In addition, no activating mutations within codon 24 of the CDK4 gene were detected. This study provides evidence for the first time that components of the pRb/p16/cyclin D1/CDK4 pathway, either alone or in combination, are frequently deregulated in human pituitary tumours, suggesting that this pathway may be a useful target in drug or gene therapeutic approaches.

Topics: Adenoma; Codon; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinases; DNA Primers; G1 Phase; Humans; Immunohistochemistry; Mutation; Pituitary Neoplasms; Proto-Oncogene Proteins; Retinoblastoma Protein; S Phase

The oncogenes cyclin D1 and D3 are overexpressed in many tumors. Topoisomerase II alpha is found in proliferating cells. The immunohistological expression of cyclin D1, cyclin D3, and Topoisomerase II alpha was studied in a collection of 60 clinically inactive surgically removed pituitary adenomas of the follicle-stimulating hormone/luteinizing hormone (FSH/LH) cell complex (20 null cell adenomas, 20 oncocytomas, and 20 FSH/LH cell adenomas) for correlation with other proliferation markers (Ki-67, PCNA) and with clinical data. Whereas cyclin D1 was positive only in one invasive null cell adenoma (1.7%) with some p53-positive nuclei, cyclin D3 was overexpressed in the nuclei of 41 tumors (68%). Topoisomerase II alpha was demonstrated in the nuclei of 42 adenomas (70%) with no significant differences discernible between the three adenoma subtypes. There was no significant correlation to the time of development of tumor symptoms, but a correlation of Topoisomerase II alpha with cyclin D3 and the proliferation marker Ki-67 (Mib1). From these data we conclude that cyclin D3 and Topoisomerase II alpha appear to be additional markers for proliferation which can be used for prognosis index in surgical pathology of the pituitary.

Topics: Adenoma; Adolescent; Adult; Aged; Aged, 80 and over; Antigens, Neoplasm; Biomarkers, Tumor; Cyclin D1; Cyclin D3; Cyclins; DNA Topoisomerases, Type II; DNA-Binding Proteins; Humans; Immunohistochemistry; Ki-67 Antigen; Middle Aged; Pituitary Neoplasms; Proliferating Cell Nuclear Antigen; Tumor Suppressor Protein p53

The cyclin D1 (CCND1) gene contains a frequent A/G polymorphism within the splice donor region of exon 4/intron 4. CCND1 genotype is associated with clinical outcome in a number of malignancies although prognostic significance varies with tumour type. We examined CCND1 allele frequencies and genotype distribution in 294 patients with sporadic pituitary adenomas of various histologies. CCND1 allele frequencies and distribution of genotypes were similar in the 294 cases compared with previously reported control populations. Analysis according to tumour subtype showed no statistical difference in allele frequencies compared with controls. However, CCND1 genotype distribution in the somatotrophinomas showed a significant difference compared with normal controls (P = 0.008). We next examined CCND1 allele frequencies and genotype distribution across the tumour grades. Within the total tumour cohort the CCND1 allele frequencies showed a significant inverse relationship across the tumour grades (P = 0.005). The CCND1 A allele progressively increased from grade 1 (0.37) through to grade 4 (0.62) tumours, whilst the CCND1 G allele frequency progressively decreased from grade 1 (0.63) through to grade 4 (0.38) tumours. Trend analysis of CCND1 genotypes showed a significant progressive increase in AA frequency from grade 1 (15%) through to grade 4 (46%) tumours (P = 0.005). The CCND1 GG genotype progressively decreased from grade 1 (41%) through to grade 4 (23%) tumours (P = 0.204). No statistical significance was observed between CCND1 AG genotype and tumour grades. While the functional significance of the observed segregation of the CCND1 A/G polymorphism and tumour grade is unclear, our data suggest that CCND1 allele frequencies and genotype distributions show significant differences between tumour grades in sporadic pituitary adenomas. Since CCND1 genotype may be determined by analysis of peripheral blood samples it may provide a useful predictive marker for those tumours likely to show invasive behaviour. This may be clinically useful in indicating which tumours should receive adjunctive treatment (e.g. radiotherapy) immediately after surgical resection.

Topics: Adenoma; Adrenocorticotropic Hormone; Codon; Cohort Studies; Cyclin D1; DNA Primers; Exons; Genotype; Growth Hormone; Humans; Introns; Neoplasm Staging; Pituitary Neoplasms; Polymerase Chain Reaction; Polymorphism, Genetic; Prolactinoma; Recurrence; Retrospective Studies; Sequence Analysis, DNA

This study was conducted to determine whether comparative genomic hybridization (CGH) is a more sensitive method for detecting genetic aberrations than other tests currently in use.. The authors used CGH to examine 40 primary and 13 recurrent adenomas obtained from 52 patients for loss and gain of genetic material. Copy number aberrations (CNAs) were detected in 25 (48%) of the 52 patients studied. The chromosomes affected were, in order of decreasing frequency, 11, 7, X, 1, 8, 13, 5, 14, 2, 6, 9, 10, 12, 3, 18, 21, 4, 16, 15, 19, 22, and Y. Endocrinologically active adenomas were more likely to contain (p = 0.009) and had a greater number (p = 0.003) of CNAs. Of 26 adenomas with CNAs, 18 showed multiple aberrations involving entire chromosomes or chromosome arms. The most frequent CNA involving a chromosome subregion, which was present in four (8%) of 53 adenomas, was the loss of all chromosome 11 material except for a preserved common segment containing 11q13. Immunoperoxidase staining did not detect cyclin D1 expression in those four cases, making cyclin D1 an unlikely target of this rearrangement.. These findings indicate that genetic abnormalities are present in pituitary adenomas at a higher rate than previously reported, are associated with endocrinological activity, and often involve several chromosomes. Rearrangement at 11q13 may inactivate a tumor suppressor gene or activate an oncogene that is important in the initiation or progression of sporadic pituitary adenomas.

Topics: Adenoma; Adolescent; Adult; Aged; Chromosome Aberrations; Chromosomes, Human, Pair 11; Cyclin D1; Female; Gene Rearrangement; Humans; Immunoenzyme Techniques; Male; Middle Aged; Nucleic Acid Hybridization; Pituitary Neoplasms; Sensitivity and Specificity

Cyclin D1 plays an important role in the regulation of cell progression through G1 of the cell cycle and has been demonstrated to have oncogenic properties. Using RFLP-PCR, an A/G polymorphism within the cyclin D1 (CCND1) gene was analyzed in 151 sporadic human pituitary tumors, of which 60 were informative at this locus. Further analysis showed that in 15 of 60 (25%) tumors, there was evidence of allelic imbalance, which is indicative of gene amplification. Allelic imbalance was observed more frequently in invasive tumors (11 of 29 tumors; 38%) than in their noninvasive counterparts (4 of 31 tumors; 13%; P = 0.02). Forty-six of the tumors informative for the polymorphism were available for immunohistochemical analysis. Cyclin D1 expression (nuclear and/or cytoplasmic) was detected in 25 of 46 (54%) tumors. Of these cases, expression of nuclear cyclin D1 was detected in 9 of 46 (20%) tumors, whereas 16 of 46 (35%) tumors showed cyclin D1 staining exclusively confined to the cytoplasm. Neither nuclear staining nor cytoplasmic staining was observed in any of the normal pituitaries or in the negative control. Expression of cyclin D1 was observed in significantly more nonfunctional tumors (18 of 27 tumors; 67%) than in somatotrophinomas (7 of 19 tumors; 37%; P = 0.046). Nuclear cyclin D1 expression was observed more frequently in nonfunctional tumors (8 of 27 tumors; 30%) than in somatotrophinomas (1 of 19 tumors; 5%; P = 0.04). There was no correlation between cyclin D1 expression and tumor grade or between allelic imbalance of CCND1 and cyclin D1 expression. We conclude that amplification of CCND1 occurs in pituitary tumors and that the overexpression of cyclin D1 may be an early event in tumorigenesis. Cyclin D1 overexpression occurring in the absence of CCND1 allelic imbalance suggests that additional mechanisms responsible for deregulated cyclin D1 expression are involved in human pituitary tumorigenesis.

Topics: Adenoma; Alleles; Cell Cycle; Cyclin D1; Gene Amplification; Growth Hormone; Humans; Immunohistochemistry; Leukocytes; Pituitary Neoplasms; Polymerase Chain Reaction; Polymorphism, Genetic; Polymorphism, Restriction Fragment Length; Subcellular Fractions

A rat pituitary tumor sub-line MtT/E-2 was established from a rat pituitary tumor cell line MtT/E. Its growth was found to depend on the presence of estradiol (E2) in culture media at 10(-13)-10(-9) M, whereas the original cell line MtT/E proliferated autonomously. The recently discovered PTTG (pituitary tumor transforming gene) is highly expressed in this cell line, although not regulated by E2. On the other hand, E2 induced c-myc and cyclin D1 proteins in MtT/E-2, which contains a lot (220+/-18 fmol/mg protein) of estrogen receptor demonstrated by RT-PCR analysis to be predominantly alpha type. MtT/E-2 secretes growth hormone which is, interestingly, regulated by retinoic acid and dexamethasone rather than thyroid hormones.

Topics: Animals; Cell Division; Culture Media; Cyclin D1; Dexamethasone; Estradiol; Gene Expression; Genes, myc; Growth Hormone; Neoplasm Proteins; Neoplasm Transplantation; Oncogene Proteins; Pituitary Neoplasms; Rats; Receptors, Estrogen; Reverse Transcriptase Polymerase Chain Reaction; Securin; Tretinoin; Tumor Cells, Cultured

The thyroid hormone, 3,3', 5-triiodo-L-thyronine (T3), is essential for growth and regulation of metabolic functions. The biological activities of T3 are mediated by its interaction with the thyroid hormone nuclear receptors (TRs). The mechanism by which TRs mediate cell growth is unknown. We found that T3 stimulated cell growth in GC cells by shortening the doubling time approximately 3-fold. Flow cytometric analysis indicated that the growth stimulatory effect was mainly due to shortening of G1 phase accompanied by increases in S and G2/M phases of the cell cycle. These changes correlated with T3-induced increases in messenger RNA and protein levels of two key regulators of G1 progression, cyclins D1 and E, as well as cdk2. Furthermore, the kinase activities associated with cyclin D1 and E were activated up to 4-fold by T3, which led to increased phosphorylation of the retinoblastoma protein (Rb), the driving force in G1 to S cell cycle progression. These results show for the first time that the growth promoting effect of T3 in GC cells is mediated, at least in part, by increases in cyclin/cdk activities and the phosphorylation state of Rb. The functional link of T3 to Rb has important implications for the understanding of the biology of normal and cancer cells.

Topics: Animals; CDC2-CDC28 Kinases; Cell Division; Cyclin D1; Cyclin E; Cyclin G; Cyclin G1; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinases; Cyclins; Flow Cytometry; G1 Phase; G2 Phase; Mitosis; Phosphorylation; Pituitary Neoplasms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Rats; Retinoblastoma Protein; RNA, Messenger; S Phase; Triiodothyronine; Tumor Cells, Cultured

The D-type cyclins (D1, D2, and D3) are involved in progression through the G1 phase of the cell cycle and are induced as part of the delayed early response to growth factor stimulation. To better understand the role of D-type cyclins in pituitary cell function and the regulatory role of growth factors in the cell cycle, we analyzed the expression and regulation of D-type cyclins in normal and neoplastic rat pituitary cells. Immunocytochemical and RT-PCR analyses showed expression of all three D-type cyclins in the normal pituitary, with higher percentages of positive cells by immunocytochemistry in the nuclei of normal pituitaries (D1, 20-30%; D2, 50-60%; D3, 70-80%), compared with GH3 cells. In the normal pituitary, there were significantly higher levels of cyclins D2 and D3 in PRL, GH, LH, and TSH cells, compared with ACTH cells. Cyclin D1 protein was not detected in GH3 cells, while D2 was present in less than 1 percent and D3 in 10-15 percent of GH3 cells. There were low levels of cyclin D1 and D2 messenger RNA expression in GH3 cells, by RT-PCR. When dissociated rat pituitary cells were cultured in the presence of basic fibroblast growth factor (5.6 nM) for 3 days, cyclin D2 was up-regulated 2-fold in normal PRL cells (control, 33 +/- 1%; treated, 68 +/- 2%). Similarly, bFGF treatment stimulated cyclin D3 expression 3-fold in GH3 cells (control, 15 +/- 1%; treated, 44 +/- 1%). Treatment of GH3 cells with 5-aza-2'-deoxycytidine, which induces gene demethylation, produced marked increases in cyclin D2 and D3 expression. Transfection of mouse cyclin D1 complementary DNA, driven by a cytomegalovirus promoter into GH3 cells, led to ectopic cyclin D1 expression; and there was a slight stimulation of cell proliferation and increased apoptosis in GH3 cells. These results indicate that there is a differential expression of various D-type cyclins in different types of normal pituitary cells and between normal pituitary and GH3 cells. Growth factors, such as bFGF and demethylation increased D-type cyclin expression, whereas ectopic overexpression of cyclin D1 stimulates cell proliferation and increases apoptosis in GH3 pituitary tumor cells.

Topics: Animals; Apoptosis; Azacitidine; Cell Cycle; Cyclin D1; Cyclin D2; Cyclin D3; Cyclins; Decitabine; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Immunohistochemistry; Mice; Pituitary Gland; Pituitary Neoplasms; Polymerase Chain Reaction; Rats; RNA-Directed DNA Polymerase; RNA, Messenger; Transfection; Tumor Cells, Cultured