transforming-growth-factor-beta and Pituitary-Neoplasms

The pituitary gland is an organ that functionally connects the hypothalamus with the peripheral organs. The pituitary gland is an important regulator of body homeostasis during development, stress, and other processes. Pituitary adenomas are a group of tumors arising from the pituitary gland: they may be subdivided in functional or non-functional, depending on their hormonal activity. Some trophic and neurotrophic factors seem to play a key role in the development and maintenance of the pituitary function and in the regulation of hypothalamo-pituitary-adrenocortical axis activity. Several lines of evidence suggest that trophic and neurotrophic factors may be involved in pituitary function, thus suggesting a possible role of the trophic and neurotrophic factors in the normal development of pituitary gland and in the progression of pituitary adenomas. Additional studies might be necessary to better explain the biological role of these molecules in the development and progression of this type of tumor. In this review, in light of the available literature, data on the following neurotrophic factors are discussed: ciliary neurotrophic factor (CNTF), transforming growth factors β (TGF‑β), glial cell line-derived neurotrophic factor (GDNF), nerve growth factor (NGF), vascular endothelial growth factor (VEGF), vascular endothelial growth inhibitor (VEGI), fibroblast growth factors (FGFs) and epidermal growth factor (EGF) which influence the proliferation and growth of pituitary adenomas.

Topics: Adenoma; Animals; Ciliary Neurotrophic Factor; Disease Progression; Epidermal Growth Factor; Fibroblast Growth Factors; Glial Cell Line-Derived Neurotrophic Factor; Humans; Nerve Growth Factor; Nerve Growth Factors; Pituitary Gland; Pituitary Neoplasms; Transforming Growth Factor beta; Tumor Necrosis Factor Ligand Superfamily Member 15; Vascular Endothelial Growth Factor A

Pituitary adenomas are common monoclonal neoplasms accounting for approximately one-fifth of primary intracranial tumors. Prolactin-secreting pituitary adenomas (prolactinomas) are the most common form of pituitary tumors in humans. They are associated with excessive release of the hormone prolactin and increased tumor growth, giving rise to severe endocrine disorders and serious clinical concerns for the patients. Recent studies indicated that the activin/TGF-beta family of growth factors plays a prominent role in regulating pituitary tumor growth and prolactin secretion from anterior pituitary lactotrope cells. Furthermore, these studies highlighted the tumor suppressor menin and the protein Smads as central regulators of these biological processes in the pituitary. Alterations in the activin/TGF-beta downstream signaling pathways are critical steps towards tumor formation and progression. This chapter will review the role and intracellular molecular mechanisms of action by which activin, TGF-beta, Smads and menin act in concert to prevent pituitary tumor cell growth and control hormonal synthesis by the anterior pituitary.

Topics: Activins; Adenoma; Gene Expression Regulation, Neoplastic; Humans; Pituitary Neoplasms; Prolactin; Proto-Oncogene Proteins; Signal Transduction; Smad Proteins; Transforming Growth Factor beta

Prolactin-secreting adenomas (prolactinomas) are the most prevalent form of pituitary tumors in humans. Our knowledge of the formation of these tumors is limited. Experimental work in animal has uncovered that estradiol exposure leads to prolactinoma formation via orchestrated events involving dopamine D2 receptors, transforming growth factor-beta(TGF-beta) isoforms and their receptors, as well as factors secondary to TGF-beta action. Additionally, these studies determined that TGF-beta and b-FGF interact to facilitate the communication between lactotropes and folliculo-stellate cells that is necessary for the mitogenic action of estradiol. The downstream signaling that governs lactotropic cell proliferation involves activation of the MAP kinase p44/42-dependent pathway.

Topics: Animals; Cell Communication; Cell Cycle; Estrogens; Gene Expression; Genomic Instability; Humans; MAP Kinase Signaling System; Models, Animal; Models, Biological; Pituitary Neoplasms; Prolactinoma; Protein Isoforms; Rats; Transforming Growth Factor beta

Pituitary adenomas are neuroendocrine tumors that produce different endocrine and metabolic alterations, including hyperprolactinemia, acromegaly and Cushing's disease. These different clinical features of pituitary tumors are the result of the overproduction of hormones produced by the different pituitary cell types. Recent advances in the understanding of the signaling pathways that control hormone production in pituitary cells provide a source of potential therapeutic targets. In ACTH-secreting cells, the mechanisms that control hormone biosynthesis have been clarified to a great extent, indicating a number of protein kinases and ligand-activated nuclear receptors as targets for experimental drugs. ACTH production requires the activation of signal transduction through the PKA, the MAPK and the CamK pathways. These pathways activate nuclear receptors, including Nur and PPAR gamma. The inhibition of these kinases and nuclear receptors has been shown to produce therapeutic effects in mouse models of Cushing's syndrome. On the other hand, the signaling pathways that control prolactin and growth hormone production also have potential targets. It has been recently shown that SMAD proteins activated by growth factors of the TGF beta and BMP family interact with estrogen receptors to stimulate the proliferation of prolactin and growth hormone-secreting cells. Cytokines that bind to the membrane protein gp130 also stimulate the proliferation of these cells. The inhibition of both of these pathways results in the decrease of tumor growth in animal models of prolactinoma. Therefore, the study of signaling pathways that control hormone production and proliferation is a good source of candidate targets in pituitary tumors.

Topics: Adrenocorticotropic Hormone; Animals; Cytokines; Drug Delivery Systems; Gene Expression Regulation; Humans; Neurosecretory Systems; Pituitary Neoplasms; Prolactin; Receptors, Cytokine; Signal Transduction; Transforming Growth Factor beta

Prolactin-secreting adenomas (prolactinomas) are the most prevalent form of pituitary tumors in humans, and increased tumor growth under estrogenic influence in female patients is often of clinical concern. Extensive experimental work has uncovered the roles of estrogen receptors and various growth-regulatory peptides in estradiol action on lactotropes. However, it is only recently that we are beginning to gain insight into how these growth factors interact to regulate estradiol action on lactotrope cell proliferation. Recent studies have identified the regulatory role of TGF-beta-related peptides in estradiol action on lactotropes. Additionally, these studies determined that TGF-beta and FGF interact to facilitate the communication between lactotropes and folliculostellate cells that is necessary for the mitogenic action of estradiol. This review addresses the role of estradiol in prolactinoma formation and summarizes data that support a novel concept: Alterations in the expression and action of TGF-beta isoforms are crucial steps in estradiol-induced tumorigenesis.

Topics: Animals; Cell Communication; Estradiol; Estrogens; Gene Expression; Humans; Hyperprolactinemia; Pituitary Neoplasms; Prolactinoma; Receptors, Estrogen; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

The effect of the TGF-

Topics: Animals; Cell Line; Chromans; Male; Matrix Metalloproteinase 9; Pituitary Neoplasms; Rats; Rats, Sprague-Dawley; Thiazolidinediones; Transforming Growth Factor beta; Transforming Growth Factor beta1; Troglitazone

Nonfunctioning pituitary adenoma is a common intracranial tumor. Though benign in the majority of cases, complications can be excruciating to the affected individual, and recurrences after tumor removal may happen with more aggressive clinical features. T regulatory (Treg) cells are generally considered a tumor-promoting immune cell type in malignant cancers with currently unclear roles in pituitary adenoma patients. Therefore, we investigated the frequency and functional characteristics of Treg cells in nonfunctioning pituitary adenoma patients before and after tumor removal. Compared to healthy controls, untreated patients with nonfunctioning pituitary adenomas presented an overrepresentation of highly functional circulating FOXP3

Topics: Adenoma; Adult; Cell Proliferation; Female; Forkhead Transcription Factors; Hepatitis A Virus Cellular Receptor 2; Humans; Immune Tolerance; Interleukin-10; Lymphocyte Activation; Male; Middle Aged; Pituitary Neoplasms; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

In pituitary adenomas, early recurrences and resistance to conventional pharmacotherapies are common, but the mechanisms involved are still not understood. The high expression of epidermal growth factor receptor 2 (HER2)/extracellular signal-regulated kinase (ERK1/2) signal observed in human pituitary adenomas, together with the low levels of the antimitogenic transforming growth factor beta receptor 2 (TBR2), encouraged us to evaluate the effect of the specific HER2 inhibition with trastuzumab on experimental pituitary tumor cell growth and its effect on the antiproliferative response to TGFB1. Trastuzumab decreased the pituitary tumor growth as well as the expression of ERK1/2 and the cell cycle regulators CCND1 and CDK4. The HER2/ERK1/2 pathway is an attractive therapeutic target, but its intricate relations with other signaling modulators still need to be unraveled. Thus, we investigated possible cross-talk with TGFB signaling, which has not yet been studied in pituitary tumors. In tumoral GH3 cells, co-incubation with trastuzumab and TGFB1 significantly decreased cell proliferation, an effect accompanied by a reduction in ERK1/2 phosphorylation, an increase of SMAD2/3 activation. In addition, through immunoprecipitation assays, a diminution of SMAD2/3-ERK1/2 and an increase SMAD2/3-TGFBR1 interactions were observed when cells were co-incubated with trastuzumab and TGFB1. These findings indicate that blocking HER2 by trastuzumab inhibited pituitary tumor growth and modulated HER2/ERK1/2 signaling and consequently the anti-mitogenic TGFB1/TBRs/SMADs cascade. The imbalance between HER2 and TGFBRs expression observed in human adenomas and the response to trastuzumab on experimental tumor growth may make the HER2/ERK1/2 pathway an attractive target for future pituitary adenoma therapy.

Topics: Adenoma; Adult; Cell Cycle; Cell Proliferation; Female; Humans; Male; Middle Aged; Phosphorylation; Pituitary Neoplasms; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Trastuzumab; Young Adult

The calcification of adamantinomatous craniopharyngioma (ACP) often creates difficulties for surgical therapy. Nevertheless, the mechanism of ACP calcification is unclear. Our previous studies demonstrated that osteoblastic factors might play important roles in ACP calcification.. We examined the effects of recombinant human Bmp2 on ACP cell differentiation by testing osteoblastic proteins and calcium deposition.. The expression of osteoblastic factors including osteopontin (OPN), Runx2, and osterix in Bmp2-treated ACP cells was examined by western blot and/or real time PCR. ALP activity and calcium deposition after Bmp2 induction were also tested.. Bmp2 significantly amplified the expression of Runx2, Osterix and OPN, as well as ALP activity. Both of these effects could be repressed by noggin treatment. Bmp2 also significantly induced the calcification of ACP, and noggin inhibited this calcium deposition.. Our study demonstrated for the first time that ACP cells could differentiate into an osteoblastic lineage via induction by Bmp2. The mechanism of ACP calcification likely involves osteoblastic differentiation modulated by Bmp2. Further studies targeting Bmp2 cascades could result in novel therapeutic interventions for recurrent ACP.

Topics: Adolescent; Adult; Alkaline Phosphatase; Bone Morphogenetic Protein 2; Calcinosis; Cell Differentiation; Core Binding Factor Alpha 1 Subunit; Craniopharyngioma; Female; Gene Expression Regulation, Neoplastic; Humans; Male; Osteoblasts; Osteopontin; Pituitary Neoplasms; Recombinant Proteins; Sp7 Transcription Factor; Transcription Factors; Transforming Growth Factor beta; Tumor Cells, Cultured

Sporadic tumors of the pituitary, parathyroids and adrenal cortex are unique, as their benign forms are very common, but malignant forms are exceptionally rare. Hereditary forms of these tumors occur in multiple endocrine neoplasia syndrome type 1 (MEN1). We hypothesize that the pathogenic link among the sporadic tumors of these organs of different germ layers might be represented by common molecular pathways involving the MEN1 gene and microRNAs (miR). miR-24 might be a microRNA linking the three tumor entities, but other candidates such as miR-142-3p and microRNAs forming the DLK1-MEG3 miRNA cluster might also be of importance.

Topics: Adrenal Cortex Neoplasms; Calcium-Binding Proteins; Cell Transformation, Neoplastic; Computational Biology; Gene Expression Profiling; Humans; Intercellular Signaling Peptides and Proteins; Membrane Proteins; MicroRNAs; Models, Theoretical; Multigene Family; Multiple Endocrine Neoplasia Type 1; Mutation; Parathyroid Neoplasms; Pituitary Gland; Pituitary Neoplasms; Proto-Oncogene Proteins; RNA, Long Noncoding; Smad3 Protein; Transforming Growth Factor beta

MicroRNAs (miRs) are small, 16-29 nucleotide long, non-coding RNA molecules which regulate the stability or translational efficiency of targeted mRNAs via RNA interference. MiRs participate in the control of cell proliferation, cell differentiation, signal transduction, cell death, and they play a role in carcinogenesis. The aims of our study were to analyse the expression profile of miRs in sporadic clinically non-functioning pituitary adenomas (NFPA) and in normal pituitary tissues, and to identify biological pathways altered in these pituitary tumors. MiR expression profiles of 12 pituitary tissue specimens (8 NFPA and 4 normal pituitary tissues) were determined using miR array based on quantitative real-time PCR with 678 different primers. Five overexpressed miRs and mRNA expression of Smads (Smad1-9), MEG and DLK1 genes were evaluated with individual Taqman assays in 10 NFPA and 10 normal pituitary tissues. Pathway analysis was performed by the DIANA-mirPath tool. Complex bioinformatical analysis by multiple algorithms and association studies between miRs, Smad3 and tumor size was performed. Of the 457 miRs expressed in both NFPA and normal tissues, 162 were significantly under- or overexpressed in NFPA compared to normal pituitary tissues Expression of Smad3, Smad6, Smad9, MEG and DLK1 was significantly lower in NFPA than in normal tissues. Pathway analysis together with in silico target prediction analysis indicated possible downregulation of the TGFβ signaling pathway in NFPA by a specific subset of miRs. Five miRs predicted to target Smad3 (miR-135a, miR-140-5p, miR-582-3p, miR-582-5p and miR-938) were overexpressed. Correlation was observed between the expression of seven overexpressed miRs and tumor size. Downregulation of the TGFβ signaling through Smad3 via miRs may have a possible role in the complex regulation of signaling pathways involved in the tumorigenesis process of NFPA.

Topics: Adenoma; Adult; Aged; Down-Regulation; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Male; MicroRNAs; Middle Aged; Oligonucleotide Array Sequence Analysis; Pituitary Neoplasms; Signal Transduction; Transforming Growth Factor beta; Validation Studies as Topic

Tenascins represent a family of extracellular matrix glycoproteins with distinctive expression patterns. Here we have analyzed the most recently described member, tenascin-W, in breast cancer. Mammary tumors isolated from transgenic mice expressing hormone-induced oncogenes reveal tenascin-W in the stroma around lesions with a high likelihood of metastasis. The presence of tenascin-W was correlated with the expression of its putative receptor, alpha8 integrin. HC11 cells derived from normal mammary epithelium do not express alpha8 integrin and fail to cross tenascin-W-coated filters. However, 4T1 mammary carcinoma cells do express alpha8 integrin and their migration is stimulated by tenascin-W. The expression of tenascin-W is induced by BMP-2 but not by TGF-beta1, though the latter is a potent inducer of tenascin-C. The expression of tenascin-W is dependent on p38MAPK and JNK signaling pathways. Since preinflammatory cytokines also act through p38MAPK and JNK signaling pathways, the possible role of TNF-alpha in tenascin-W expression was also examined. TNF-alpha induced the expression of both tenascin-W and tenascin-C, and this induction was p38MAPK- and cyclooxygenase-dependent. Our results show that tenascin-W may be a useful diagnostic marker for breast malignancies, and that the induction of tenascin-W in the tumor stroma may contribute to the invasive behavior of tumor cells.

Topics: Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Breast Neoplasms; Cell Line, Tumor; Female; Gene Expression Regulation, Neoplastic; Humans; Integrin alpha Chains; Oligonucleotide Array Sequence Analysis; p38 Mitogen-Activated Protein Kinases; Pituitary Neoplasms; RNA, Neoplasm; Tenascin; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

To study the effect of interleukin-11(IL-11), ciliary neurotropic factor (CNTF) and transforming growth factor-beta (TGF-beta) on the hGH gene promoter activity in rat pituitary GH(3) cells and the interaction with pituitary-specific transcription factor Pit-1, firstly the stable transformed GH(3) cell line which contained hGH gene promoter 484-30 bp and luciferase reporter gene was established, then the concentration of GH in the medium and lysate of GH(3) cells and luciferase activities in GH(3) cells were measured, after treating these cells with the above cytokines, the effects of cytokines on secretion and synthesis of GH, and the promoter activity of the hGH gene were observed. The results of our experiments showed that IL-11(20 nmol/L), CNTF(10 nmol/L) and TGF-beta(5 nmol/L) regulated secretion and synthesis of GH, and the luciferase expression in stable-transformed GH(3) cells. IL-11 and CNTF had a stimulatory effect, whereas TGF-beta had an inhibitory one. Neither overexpression of Pit-1 nor inhibition of Pit-1 expression could affect the regulatory role of these cytokines. In conclusion, IL-11, CNTF and TGF-beta regulated the GH production in pituitary GH(3) cell line by regulating the hGH gene promoter activity, while Pit-1 might not be involved in the roles.

Topics: Animals; Ciliary Neurotrophic Factor; Cytokines; DNA-Binding Proteins; Gene Expression; Growth Hormone; Human Growth Hormone; Humans; Interleukin-11; Luciferases; Pituitary Neoplasms; Plasmids; Promoter Regions, Genetic; Rats; Recombinant Fusion Proteins; Transcription Factor Pit-1; Transcription Factors; Transforming Growth Factor beta; Tumor Cells, Cultured

Galectin-3 (Gal-3), a beta-galactoside-binding protein, has been implicated in a variety of biological functions, including cell proliferation and differentiation, tumor cell adhesion, angiogenesis, apoptosis, tumor progression, and metastasis. We investigated the role of Gal-3 in the development and progression of pituitary tumors. Immunohistochemical and Western blot analysis of normal and neoplastic human pituitaries showed that only lactotroph (PRL) and corticotroph (ACTH) hormone-producing cells and tumors expressed Gal-3. Gal-3 was present in 24 of 38 (63.2%) PRL adenomas, 5 of 6 (83.3%) PRL carcinomas, 19 of 41 (46.3) ACTH adenomas, and 7 of 8 (87.5%) ACTH carcinomas, but not in 112 other pituitary adenomas and carcinomas. Pituitary folliculo-stellate cells, which have macrophage-type functions in the anterior pituitary, also expressed Gal-3. Hyperplastic and neoplastic pituitaries from p27(Kip1) (p27)-null mice, which produce mainly ACTH, showed increased Gal-3 expression levels compared with control mice. Treatment with transforming growth factor beta1, which regulates pituitary cell proliferation, reduced Gal-3 as well as p27 expression levels in cultured HP75 pituitary cells and Gal-3 in cultured pituitary cells from p27-null mice, suggesting that p27 is not necessary for the inhibitory effects of transforming growth factor beta1 on the cell cycle in the pituitary. The role of Gal-3 in pituitary cell function was examined by RNA interference experiments. Inhibition of Gal-3 gene expression by RNA interference decreased HP75 cell proliferation and increased apoptosis. These results indicate that Gal-3 has an important role in pituitary cell proliferation and tumor progression.

Topics: Adenoma; Animals; Blotting, Western; Carcinoma; Cell Cycle Proteins; Cell Division; Cyclin-Dependent Kinase Inhibitor p27; Disease Progression; DNA, Antisense; Galectin 3; Humans; Immunohistochemistry; In Situ Hybridization; Mice; Pituitary Neoplasms; RNA Interference; RNA, Small Interfering; Transfection; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Cells, Cultured; Tumor Suppressor Proteins

Recent studies have shown that 7B2 and the neuroendocrine- specific proconvertase PC2 have important roles in pituitary cell proliferation and hormone secretion. Studies from our laboratory have also shown that TGFb1 regulates anterior pituitary cell proliferation and hormone secretion. To study the regulation of 7B2 in human pituitary tumors, we used a cell line derived from a human pituitary adenoma (HP75) that has been shown to express 7B2, PC1, PC2, and TGFbeta receptors to analyze the effects of TGFbeta1 and the histone deacetylase inhibitor (HDACI) sodium butyrate (NaB) treatment on 7B2 mRNA expression along with the neuroendocrine-specific proconvertases 1/3 (PC1) and PC2 mRNA and protein expression. RNA was quantified by real-time PCR and proteins were detected by immunohistochemistry and Western blotting. Treatment of cells with 1 mM NaB or 1 nM TGF 1 for 4 d decreased cell proliferation with a concomitant increase in the cell cycle protein p21. Real-time PCR analysis showed a significant increase in 7B2 mRNA after NaB and TGFbeta1 treatment. PC2 mRNA was down regulated by NaB while PC1 mRNA was unchanged. TGFbeta1 stimulated PC1, but not PC2 mRNA levels. Changes in PC1 and PC2 protein were similar to changes in the mRNAs, but the differences were not significant. These results indicated that NaB and TGFbeta1 inhibit pituitary cell proliferation and regulate the expression of 7B2, PC1, and PC2 in a cell culture model of pituitary tumors. Our results also indicate that inhibition of pituitary cell proliferation is associated with increased expression of 7B2 mRNA.

Topics: Blotting, Western; Butyrates; Cell Line, Tumor; Enzyme Inhibitors; Humans; Immunohistochemistry; Nerve Tissue Proteins; Neuroendocrine Secretory Protein 7B2; Pituitary Gland; Pituitary Hormones; Pituitary Neoplasms; Proliferating Cell Nuclear Antigen; Proprotein Convertase 1; Proprotein Convertase 2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta

The sst2 somatostatin receptor is an inhibitory G protein-coupled receptor, which exhibits anti-tumor properties. Expression of sst2 is lost in most human pancreatic cancers. We have cloned 2090 base pairs corresponding to the genomic DNA region upstream of the mouse sst2 (msst2) translation initiation codon (ATG). Deletion reporter analyses in mouse pituitary AtT-20 and human pancreatic cancer PANC-1, BxPC-3, and Capan-1 cells identify a region from nucleotide -260 to the ATG codon (325 base pairs) showing maximal activity, and a region between nucleotides -2025 and -260 likely to comprise silencer or transcriptional suppressor elements. In PANC-1 and AtT-20 cells, transforming growth factor (TGF)-beta up-regulates msst2 transcription. Transactivation is mediated by Smad4 and Smad3. The cis-acting region responsible for such regulation is comprised between nucleotides -1115 and -972 and includes Sp1 and CAGA-box sequences. Expression of Smad4 in Smad4-deficient Capan-1 and BxPC-3 cells restores TGF-beta-dependent and -independent msst2 transactivation. Expression of Smad4 in BxPC-3 cells reestablishes both endogenous sst2 expression and somatostatin-mediated inhibition of cell growth. These findings demonstrate that msst2 is a new target gene for TGF-beta transcription regulation and underlie the possibility that loss of Smad4 contributes to the lack of sst2 expression in human pancreatic cancer, which in turn may contribute to a stimulation of tumor growth.

Topics: 5' Untranslated Regions; Animals; Base Sequence; Cloning, Molecular; Codon; DNA-Binding Proteins; Gene Expression Regulation, Neoplastic; Genes, Reporter; Genomic Library; Humans; Luciferases; Mice; Molecular Sequence Data; Pancreatic Neoplasms; Pituitary Neoplasms; Promoter Regions, Genetic; Receptors, Somatostatin; Reverse Transcriptase Polymerase Chain Reaction; Sequence Deletion; Smad4 Protein; Trans-Activators; Transcription, Genetic; Transcriptional Activation; Transforming Growth Factor beta; Tumor Cells, Cultured

Male SD rats transplanted with an extra pituitary in the renal capsule and treated chronically with 17-beta-estradiol (E2) were used in the studies on the pathogenesis and the underlying mechanisms of pituitary prolactin-secreting tumor (prolactinoma). The results indicated that after long treatment with E2, prolactin-secreting tumor was generated in both eutopic and ectopic pituitary, accompanied by hyperprolactinemia and overexpression of PRL gene. No apparent difference was observed in histology and ultrastructure between them. Further investigations showed that some growth factors might be involved in the tumorigenesis of prolactinoma in vivo and in vitro. A point mutation was found in the proximal promoter of PRL gene only in eutopic prolactinoma by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP), suggesting that the base change in the promoter may be related to the overexpression of PRL gene. This hypothesis has been confirmed by the increased activity of luciferase reporter fused to the mutant promoter in vitro. The coincidence of overexpression of PRL, TGF alpha and TGF beta 1 gene, and the point mutation detected in eutopic pituitary prolactinoma suggested that neuro-endocrine-immune interactions in vivo might be associated with pituitary prolactinoma formation. The mechanisms mediating tumorigenesis of eutopic and ectopic prolactinoma, respectively, may be different.

Topics: Animals; Endothelial Growth Factors; Estradiol; Male; Pituitary Neoplasms; Point Mutation; Prolactin; Prolactinoma; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Pituitary adenylate cyclase-activating polypeptide (PACAP) was originally isolated from hypothalamic tissues based on its ability to stimulate cAMP production in cultured anterior pituitary cells. Recent studies have suggested a functional role for PACAP in the apoptosis of brain cells. However, the role of PACAP in regulating apoptosis in human pituitary adenomas has not previously been examined. Analysis of the cultured human pituitary adenoma cell line HP75, which expresses all three major PACAP receptors, showed that both PACAP-38 and PACAP-27 inhibited TGF-beta1-induced apoptosis. Treatment with the PACAP receptor antagonists PACAP 6-38 (PACAP type I receptor antagonist) and (p-chloro-D-Phe(6), Leu(17))-VIP (PACAP type II receptor antagonist) blocked the effects of PACAP-38 on the inhibition of transforming growth factor-beta1 (TGF-beta1)-induced apoptosis, confirming the specificity of the role of PACAP. Treatment with forskolin but not phorbol 12-myristate 13-acetate (PMA) also inhibited TGF-beta1-induced apoptosis. TGF-beta1 treatment was associated with an increase in mitogen-activated protein kinase (MAP kinase) when analyzed by Western blotting, but PACAP inhibition of TGF-beta1-induced apoptosis was not associated with activation of MAP kinase. Immunocytochemical analysis of the cell cycle cyclin-dependent kinase inhibitor p27 showed that treatment with TGF-beta1, forskolin, PMA, and PACAP increased p27 expression in cultured HP75 cells. These results indicate that PACAP is a highly specific inhibitor of TGF-beta1-induced apoptosis in the HP75 human pituitary adenoma cell line and that PACAP, TGF-beta1, forskolin, and PMA all stimulate expression of the TGF-beta-regulated cell cycle protein p27 in the HP75 human pituitary adenoma cell line. The HP75 cell line can be used as a model to study the regulation of apoptosis in human pituitary cells.

Topics: Adenoma; Apoptosis; Cell Cycle; Cell Cycle Proteins; Cyclin-Dependent Kinase Inhibitor p27; Humans; Immunohistochemistry; Microtubule-Associated Proteins; Mitogen-Activated Protein Kinases; Neuropeptides; Nucleic Acid Hybridization; Pituitary Adenylate Cyclase-Activating Polypeptide; Pituitary Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Suppressor Proteins

The Sprague-Dawley (SD) rat bearing a heteroplasted pituitary underneath renal capsule was used to observe differential expression of prolactin (PRL), transforming growth factor alpha (TGF alpha) and transforming growth factor beta 1 (TGF beta 1) genes during the formation of pituitary prolactin-secreting tumor (prolactinoma) induced by 17 beta-estradiol (E2). Our results indicated that in both eutopic and ectopic pituitaries disconnected from hypothalamus formed simultaneously PRL-secreting tumors after the rats treated with E2 for 120 days in vivo, which was accompanied by overexpression of PRL gene (P < 0.05-0.01). The PRL mRNA level was higher in eutopic prolactinoma than that in ectopic prolactinoma (P < 0.05). Overexpression of TGF alpha and TGF beta 1 genes were also detected in eutopic prolactinoma. However, the expression of TGF alpha and TGF beta 1 genes in ectopic prolactinoma was similar to that in normal pituitary. It is suggested that TGF alpha and TGF beta 1 may be involved in prolactinoma tumorigenesis of eutopic pituitary. However, the mechanism mediating eutopic and ectopic prolactin-secreting tumor formation seems different.

Topics: Animals; Estradiol; Gene Expression; Male; Neoplasm Transplantation; Pituitary Neoplasms; Prolactin; Prolactinoma; Rats; Rats, Sprague-Dawley; RNA; Subrenal Capsule Assay; Transforming Growth Factor alpha; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor (TGF)-beta has been implicated in the regulation of normal and neoplastic anterior pituitary cell function. TGF-beta regulates the expression of various proteins, including p27Kip1 (p27), a cell cycle inhibitory protein. We examined TGF-beta, TGF-beta type II receptor (TGF-beta-RII), and p27 expression in normal pituitaries, pituitary adenomas, and carcinomas to analyze the possible roles of these proteins in pituitary tumorigenesis. Normal pituitary, pituitary adenomas, and pituitary carcinomas all expressed TGF-beta and TGF-beta-RII immunoreactivity. Reverse transcription polymerase chain reaction analysis showed TGF-beta 1, -beta 2, and -beta 3 isoforms and TGF-beta-RII in normal pituitaries and pituitary adenomas. Pituitary adenomas cells cultured for 7 days in defined media showed a biphasic response to TGF-beta with significant inhibition of follicle-stimulating hormone secretion at higher concentrations (10(-9) mol/L) and stimulation of follicle-stimulating hormone secretion at lower concentrations (10(-13) mol/L) of TGF-beta 1 in gonadotroph adenomas. Immunohistochemical analysis for p27 protein expression showed the highest levels in nontumorous pituitaries with decreased immunoreactivity in adenomas and carcinomas. When nontumorous pituitaries and various adenomas were analyzed for p27 and specific hormone production, growth hormone, luteinizing hormone, and thyroid-stimulating hormone cells and tumors had the highest percentages of cells expressing p27, whereas adrenocorticotrophic hormone cells and tumors had the lowest percentages. Immunoblotting analysis showed that adrenocorticotrophic hormone adenomas also had the lowest levels of p27 protein. Semiquantitative reverse transcription polymerase chain reaction and Northern hybridization analysis did not show significant differences in p27 mRNA expression in the various types of adenomas or in nontumorous pituitaries. In situ hybridization for p27 mRNA showed similar distributions of the gene product in nontumorous pituitaries, pituitary adenomas, and carcinomas. These results indicate that TGF-beta and TGF-beta-RII are widely expressed in nontumorous pituitaries and in pituitary neoplasms and that TGF-beta 1 regulates pituitary hormone secretion. The levels of the TGF-beta-regulated protein p27 decreases in the progression of normal to neoplastic pituitaries. In contrast, the mRNA levels of p27 remained relatively constant in nontumorous pituitaries, pituitary adeno

Topics: Adenoma; Carcinoma; Cell Cycle Proteins; Cyclin-Dependent Kinase Inhibitor p27; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Humans; Immunohistochemistry; In Situ Hybridization; Microtubule-Associated Proteins; Pituitary Gland; Pituitary Neoplasms; Receptors, Transforming Growth Factor beta; RNA, Messenger; Transforming Growth Factor beta; Tumor Suppressor Proteins

Activin, a member of the transforming growth factor-beta (TGF beta) cytokine family, acts as a pituitary cell mitogen via a novel family of receptor-linked serine/threonine (Ser/Thr) kinases. Pituitary tumors synthesize activin subunits, and the autocrine action of these growth factors may modulate tumor proliferation. We, therefore, investigated the expression of activin/TGF beta type I receptor messenger ribonucleic acids (mRNAs), designated ALK1 through ALK5 (ALK = activin receptor-like kinase), and type II receptor mRNAs using RT-PCR in 34 human pituitary adenomas of all phenotypes and normal pituitary tissue. ALK2 and ALK5, specific mediators of activin and TGF beta signals, respectively, were found to be expressed only in tumor and not in normal pituitary cells, and ALK2 expression was found only in tumors of a mammosomatotroph cell lineage. ALK1, ALK3, and ALK4 mRNAs were found in both normal and neoplastic pituitary cells. The alternatively spliced cytoplasmic domain of ALK4 consists of 11 kinase subdomains, that are critical for modulating receptor function and intracellular signaling. Truncated forms of the ALK4 cytoplasmic domain lacking these subdomains may attenuate activin signal transduction and affect both tumor phenotype and proliferation via the formation of inactive type I/type II complexes. Three truncated ALK4 receptor mRNAs generated by alternate splicing of the cytoplasmic Ser/Thr kinase domain were found to be tumor specific. One of these truncated receptor mRNAs, ALK4-5, is a novel splice variant that has not been previously described. Expression of the ActRII and T beta RII type II receptor mRNAs, which specifically bind activin and TGF beta, respectively, was highly prevalent among all tumor subtypes and normal pituitary tissue. However, ActRIIB, an activin-specific type II receptor that displays a 3- to 4-fold higher affinity for ligand than ActRII, was expressed in 94% of tumors, but was not prevalent in normal tissue. These data are the first to demonstrate tumor-specific expression of Ser/Thr kinase receptors mRNAs and their splice variants in human pituitary adenomas.

Topics: Activin Receptors; Activins; Adenoma; Adolescent; Adult; Alternative Splicing; Amino Acid Sequence; Base Sequence; Gene Expression; Humans; Inhibins; Middle Aged; Molecular Sequence Data; Pituitary Neoplasms; Polymerase Chain Reaction; Receptors, Growth Factor; Receptors, Transforming Growth Factor beta; RNA, Messenger; Transforming Growth Factor beta

Transforming growth factor-beta (TGF beta) is a member of a family of growth factors that regulates differentiation and cellular proliferation in a wide variety of tissues, including the anterior pituitary gland. TGF beta regulates the expression of various proteins, including p27Kipl (p27), a cell cycle inhibitory protein. The cell types in normal rat anterior pituitary producing TGF beta1, one of the principal isoforms of TGF beta, and p27 were examined by in situ methods. The regulation of p27 messenger RNA (mRNA) and protein by TGF beta1 was also examined in cultured anterior pituitary cells. In situ hybridization, in situ reverse transcriptase PCR, and immunocytochemical staining for pituitary hormones showed that PRL, TSH, and gonadotroph cells all had a higher percentage of cells expressing TGF beta1 mRNA and p27 protein than did GH and ACTH cells. After treatment with 10(-9) M TGF beta1 in vitro for 3 days, there were significant decreases in p27 mRNA and protein levels (P < 0.05) in normal pituitary cells. The GH3 and GHRH-CL1 cell lines, which secrete PRL and GH, had undetectable p27 protein by immunocytochemical staining and immunoblotting, although the GH3 cell line had p27 mRNA detected by reverse transcriptase PCR. Analysis of [3H]thymidine uptake in cultured dissociated pituitary cells by double staining for hormones showed that only PRL cells had significant proliferative activity during a 3-day cell culture period. There was a biphasic effect of TGF beta1 on PRL cell proliferation, with marked inhibition by 10(-9) M and a slight stimulation by 10(-13) M. These results indicate that there is a differential distribution of both TGF beta1 and p27 in various anterior pituitary cell types and that TGF beta1 directly down-regulates p27 in cultured anterior pituitary cells.

Topics: Animals; Base Sequence; Cell Cycle Proteins; Cell Division; Cell Line; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p27; DNA Primers; DNA Probes; Female; Gene Expression Regulation; In Situ Hybridization; Microtubule-Associated Proteins; Molecular Sequence Data; Pituitary Gland, Anterior; Pituitary Neoplasms; Polymerase Chain Reaction; Rats; Rats, Inbred WF; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Suppressor Proteins

There is increasing evidence for the role of cytokines in pituitary differentiated function and tumorigenesis, but the spectrum of cytokines found in the pituitary is unknown. Therefore profiles of cytokine expression were determined in different human anterior pituitary adenoma sub-types.. The reverse transcriptase-linked polymerase chain reaction (PCR) was used to identify the presence of cytokine mRNA within human pituitary adenomas.. Seventeen pituitary adenoma biopsies removed at transsphenoidal surgery were examined: 4 somatotrophinomas, 7 non-functional adenomas, 4 prolactinomas, one case of Cushing's disease and one case of Nelson's syndrome.. RNA was extracted from each adenoma biopsy and reverse transcribed into cDNA. This was specifically amplified in a PCR using oligonucleotide primers complementary to each cytokine. The cytokines investigated were interleukin (IL)-I alpha, IL-I beta, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, tumour necrosis factor (TNF)-alpha, TNF-beta and transforming growth factor (TGF)-beta 1, beta 2 and beta 3. The products of each PCR were visualized using agarose gel electrophoresis.. All 17 adenomas expressed IL-8 transcripts, but no expression of IL-2, IL-5 or IL-7 was found. IL-6 was expressed in all 4 somatotrophinomas, 3 of 7 non-functional tumours, 2 of 4 prolactinomas and in the single case of Nelson's syndrome. At least one of the 3 isoforms of TGF-beta was found in all but 2 tumours; one prolactinoma and one non-functional adenoma. IL-1 alpha, IL-beta, IL-4, TNF-alpha and TNF-beta were expressed sporadically by individual adenomas.. These data suggest that whilst IL-8 may be important, the local expression of the cytokines IL-2, IL-5 and IL-7 is not important in human anterior pituitary tumorigenesis.

Topics: Adenoma; Adult; Aged; Base Sequence; Cushing Syndrome; Cytokines; DNA Primers; Female; Growth Hormone; Humans; Interleukin-6; Interleukin-8; Male; Middle Aged; Molecular Sequence Data; Nelson Syndrome; Pituitary Gland, Anterior; Pituitary Neoplasms; Polymerase Chain Reaction; Prolactinoma; RNA, Messenger; Transforming Growth Factor beta

We have previously shown that transforming growth factor beta 1 (TGF-beta 1) receptor and TGF-beta type II receptor (T beta R-II) are produced in lactotropes, and that TGF-beta 1 inhibits the growth of these anterior pituitary cells by an autocrine mechanism. To study the changes of the expression and function of this growth factor during tumorigenesis, we have measured the levels of TGF-beta 1 and T beta R-II mRNAs and proteins in the normal and tumor anterior pituitary cells in vivo and in vitro and have compared the cell growth responses to TGF-beta 1 in normal and tumor pituitary cells in vitro. Treatment with estradiol-17 beta for 1, 2, 4, and 8 weeks caused a time-dependent increase in pituitary protein, prolactin, and prolactin mRNA levels and in plasma prolactin levels, suggesting that estrogen enhanced lactotropic proliferation in anterior pituitary glands. The levels of TGF-beta 1 protein and mRNA in anterior pituitary tissues were reduced over time after estrogen treatment during the development of pituitary tumors. The mRNA and protein levels of T beta R-II decreased markedly during the development of pituitary tumors. In addition, two transformed lactotropes, GH3 and PR1 cell lines, showed markedly reduced levels of TGF-beta 1 as well as T beta R-II mRNA. Comparison of the antiproliferative effects of TGF-beta in transformed and normal lactotropes in cultures revealed that the sensitivity of GH3 cells is reduced, and that PR1 cells are virtually resistant to TGF-beta 1.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cell Division; Cell Line, Transformed; Estradiol; Female; Pituitary Gland, Anterior; Pituitary Neoplasms; Prolactin; Protein Serine-Threonine Kinases; Rats; Rats, Inbred F344; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; RNA, Messenger; Transforming Growth Factor beta; Tumor Cells, Cultured

A rat pituitary tumor cell line (GH3) has been reported to express transforming growth factor-beta (TGF-beta) binding components of 70-74 kDa (ligand included), denoted TGF-beta type IV receptor. We investigated whether the type IV receptor corresponds to any of the recently cloned type I receptors for proteins in the TGF-beta super-family. TGF-beta type I receptor (T beta R-I) complexes of 69-72 kDa formed a heteromeric complex with T beta R-II in GH3 cells, as detected by immunoprecipitation. In addition, TGF-beta formed complexes of 72-74 kDa, which were different from T beta R-I and the other known type I receptors, and were not dependent on T beta R-II for binding. The GH3 cells were resistant to the growth inhibitory activity of TGF-beta, but a transcriptional response was activated by TGF-beta in this cell line, presumably through the T beta R-II and T beta R-I complex. These results indicate that GH3 cells have T beta R-I and T beta R-II and, in addition, other binding protein(s) which form 72-74-kDa complexes with TGF-beta; the function of the latter component(s) remains to be elucidated.

Topics: Activins; Animals; Base Sequence; Cells, Cultured; DNA Primers; Inhibins; Membrane Proteins; Molecular Sequence Data; Pituitary Neoplasms; Polymerase Chain Reaction; Protein Binding; Rats; Receptors, Transforming Growth Factor beta; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta; Tumor Cells, Cultured

We have shown that growth of F4Z2 cells and F4Z2 tumors was stimulated by estradiol, that of MtTF4 and F4P tumors was inhibited and that of F4P cells remained insensitive. In the present work we explore the possible role of transforming growth factor-beta (TGF-beta) as a mediator of estradiol action in these pituitary tumors and cell lines. In vivo, estradiol treatment increased the concentration of TGF-beta 1 mRNAs in tumors whose growth was inhibited by estradiol (MtTF4 and F4P) but not in tumors whose growth was stimulated (F4Z2). F4Z2 and F4P cell lines also contained TGF-beta 1 transcripts. These cells and tumors differed by two points: the level of TGF-beta 1 transcript was higher in F4Z2 than in F4P cells while the opposite situation was observed in vivo and the concentration of TGF-beta 1 mRNA in cultured cells was insensitive to estradiol (1 or 100 x 10(-9) M). Moreover, the secretion of TGF-beta like activity assayed by two different methods was estradiol insensitive and the growth of both cell lines was dose-dependently inhibited by TGF-beta 1 (ED50:2 x 10(-11) M). Since estradiol increases TGF-beta 1 mRNA in the tumors MtTF4 and F4P whose growth is inhibited by estradiol and that TGF-beta 1 inhibits the proliferation of F4P cells it is proposed as a working hypothesis that TGF-beta 1 is one of the mediators of the inhibitory effect of estradiol in pituitary tumors. No data favor the hypothesis that estradiol stimulates pituitary tumor proliferation by decreasing TGF-beta production.

Topics: Animals; Blotting, Northern; Cell Division; Estradiol; Female; Pituitary Neoplasms; Rats; Rats, Inbred F344; Transforming Growth Factor beta; Tumor Cells, Cultured

Transforming growth factor-beta (TGF beta) is a member of a large family of growth factors, several of which regulate pituitary function. TGF beta has recently been reported to reduce PRL production by GH4 cells. We have examined the effect of TGF beta on PRL gene expression in rat pituitary tumor GH3 cells. TGF beta 1 or TGF beta 2 reduced both basal and Ca(2+)-stimulated PRL mRNA levels. This inhibition was specific, as the mRNA levels for GH, glucose-regulated protein 78, and histone-3 were unaffected by TGF beta. Inhibition of PRL gene expression by TGF beta was dose dependent in the range of 0.5-10 ng/ml. TGF beta inhibited run-on PRL gene transcription in nuclei from treated cells to the same extent that it reduced PRL mRNA levels, indicating a transcriptional mechanism of action. However, TGF beta did not affect Pit-1 mRNA levels or run-on transcription of the Pit-1 gene. Thus, TGF beta does not appear to act through modification of Pit-1 gene expression. The PRL promotor contains two regions of homology, with a consensus sequence found in the promoters of other TGF beta-inhibited genes. These findings are consistent with other studies that have demonstrated transcriptional repression by TGF beta. The potency and specificity of the effects of TGF beta on PRL gene expression suggest that it may be a physiological regulator of lactotroph function.

Topics: Animals; Cell Line; Cell Nucleus; Cytoplasm; Dexamethasone; DNA-Binding Proteins; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Kinetics; Pituitary Neoplasms; Prolactin; Rats; RNA, Messenger; RNA, Neoplasm; Transcription Factor Pit-1; Transcription Factors; Transcription, Genetic; Transforming Growth Factor beta

The mechanisms that restrict cell proliferation play an important regulatory role in differentiation and tumorigenesis. The growth of PRL-secreting cells of the anterior pituitary is known to be highly estrogen dependent; however, estrogen may act indirectly via growth regulatory polypeptides. We have used the GH4C1 rat pituitary cell line to investigate the action of two classes of growth regulatory polypeptides, transforming growth factor-alpha (TGF alpha) and TGF beta. TGF alpha and TGF beta each inhibit GH4 cell proliferation, as measured by cell number and [3H]thymidine incorporation, and given together arrest GH4 cell proliferation. The growth inhibitory action of TGF alpha is concentration dependent (IC50 = 100 pM) and saturable. Activin-A, a TGF beta-related polypeptide, also inhibits proliferation, but is less effective than TGF beta. TGF alpha and TGF beta each alter GH4 cell cycle distribution by decreasing in the percentage of S phase cells (74% and 34%, respectively) and increasing proportionally G0-G1 phase cells. The growth inhibitory action of TGF alpha differs from that of TGF beta in that TGF alpha also causes a temporary accumulation of cells in G2-M phases. We next initiated experiments to evaluate the role of protein kinase-C in the growth inhibitory actions of TGF alpha and TGF beta. The alpha- and beta-isoforms of protein kinase-C were down-regulated by pretreatment with 12-O-tetradecanoylphorbol-13-acetate, yet TGF alpha and TGF beta still substantially inhibited GH4 cell proliferation. We next compared the actions of TGF alpha and TGF beta on two other well characterized prolonged GH4 responses. TGF alpha and TGF beta each increased GH4 cell adhesion, but differed in their effects on PRL production. This indicates that TGF alpha and TGF beta activate different signaling pathways in GH4 cells. Activin-A acted like TGF beta by enhancing cell-substratum adhesion and inhibiting PRL production, consistent with an interaction at a common receptor site. Taken together these results identify biological functions for TGF alpha, TGF beta, and activin-A on PRL cells and open the possibility that they may represent the direct in vivo mediators of estrogen action to regulate the growth of PRL cells in the anterior pituitary gland.

Topics: Activins; Animals; Cell Adhesion; Cell Count; Cell Division; DNA; Inhibins; Interphase; Mitosis; Phorbol 12,13-Dibutyrate; Pituitary Neoplasms; Prolactin; Protein Kinase C; Rats; Recombinant Proteins; S Phase; Tetradecanoylphorbol Acetate; Transforming Growth Factor alpha; Transforming Growth Factor beta; Tumor Cells, Cultured