Human Pituitary Research Articles

TMEFF2: a new endogenous modulator of the CRH signaling in corticotroph cells

We have previously demonstrated that retinoic acid (RA) inhibits CRH signaling in corticotroph cells. In order to elucidate the mechanisms controlling RA action, cDNA microarray analysis was perfomed. From 87 genes regulated by RA in the corticotroph cell line AtT-20, we selected TMEFF2 for further studies. TMEFF2 is a transmembrane protein containing an altered epidermal growth factor (EGF)-like motif, two follistatin-like domains and a cytosolic tail which a putative G protein-activating motif. TMEFF2 is predominantly expressed in brain and prostate and has been implicated in cell signaling, neuronal cell survival and tumor suppression. Although TMEFF2 function is unknown, its structural domains suggest that it may have a role in the regulation of growth factor signalling. Since there is no previous evidence for an involvement of TMEFF2 in Cushing's disease or CRH signalling, we have studied the effects of TMEFF2 on the signal transduction pathway of CRH. Our results show that CRH-signaling is under the inhibitory effects of endogenous TMEFF2. TMEFF2 expression in corticotroph cells inhibits the effects of CRH on the production of intracellular cAMP, CREB and POMC transcriptional activity. Regulation of CRH activity by TMEFF2 does not require the cytoplasmic tail, while deletion of the follistatin and EGF modules abolishes the inhibitory function of TMEFF2. Moreover, a soluble secreted protein containing these three extracellular domains is sufficient for CRH signaling inhibition. TMEFF2-induced inhibition depends on serum components. Furthermore, TMEFF2 seems to regulate non-cannonical Activin/BMP-4 signaling, leading to the inhibition of the PI3K pathway (Akt) and Erk1/2 activation. Whether TMEFF2 acts upstream to Akt to regulate cAMP production and PKA activity remains to be elucidated. We found that TMEFF2 expression in human Cushing's adenoma is reduced as compared to the normal human pituitary, which may indicate that TMEFF2 acts as a tumor suppressor in these adenomas. Consistent with this hypothesis, the overexpression of TMEFF2 decreased cell proliferation in corticotroph cells. Our results also indicate a potential therapeutic use of TMEFF2 or factors that stimulate TMEFF2 activity for the treatment of corticotroph tumors in order to reduce their ACTH secretion and proliferation.

Iatrogenic Creutzfeldt–Jakob disease in Australia: time to amend infection control measures for pituitary hormone recipients?

From 1967, the Australian Human Pituitary Hormone Program offered treatment for short stature and infertility using human cadaver-acquired pituitary hormones (human growth hormone [hGH] and human pituitary gonadotrophin [hPG]). The program was suspended in 1985when a growth-hormone recipient in the United States developed Creutzfeldt-Jakob disease (CJD), an incurable and rapidly progressive neurodegenerative disorder. Since this time, recipients have lived with the significant anxiety that they have an elevated risk of developing CJD. Furthermore, additional CJD infection control measures are required when recipients undergo some types of surgery. As it is 20years since the last Australian pituitary hormone recipient developed CJD, we evaluated the risk for Australian recipients of developing iatrogenic CJD, and compared Australian data with data from New Zealand and selected other countries who had pituitary hormone programs. Our evaluation indicates that pituitary hormone recipients in Australia have the lowest risk of developing iatrogenic CJD, and that Australia is the only country not to have experienced ongoing CJD-related deaths. Thus, we believe that: in the Australian hGH recipient cohort, the risk of developing CJD is sufficiently low for this cohort to no longer require additional infection control measures in the health care setting; and in the Australian hPG recipient cohort, if another 5years elapses with no further occurrence of CJD in this group, the hPG recipient cohort could also be considered as not requiring additional infection control measures in the health care setting. These recommendations should not be misunderstood as implying that there is no ongoing risk, but that the risk is acceptably low and generally in keeping with guidelines that stratify the risk.

Magmas, a Gene Newly Identified as Overexpressed in Human and Mouse ACTH-Secreting Pituitary Adenomas, Protects Pituitary Cells from Apoptotic Stimuli

Pituitary tumors are mostly benign, being locally invasive in 5-35% of cases. Deregulation of several genes has been suggested as a possible alteration underlying the development and progression of pituitary tumors. We here report the identification of a cDNA, corresponding to Magmas gene (mitochondria-associated protein involved in granulocyte-macrophage colony-stimulating factor signal transduction), which is highly expressed in two different ACTH-secreting mouse pituitary adenoma cell lines as compared with normal pituitary as well as in two thirds of 64 examined pituitary adenomas as compared with human normal pituitary. Tim 16, the mitochondrial protein encoded by Magmas, was indeed expressed in a mouse ACTH-secreting pituitary adenoma cell line, AtT-20 D16v-F2 cells, in a subcellular compartment likely corresponding to mitochondria. Magmas silencing determined a reduced rate of DNA synthesis, an accumulation in G1 phase, and a concomitant decrease in S phase in At-T20 D16v-F2 cells. Moreover, Magmas-silenced cells displayed basal caspase 3/7 activity and DNA fragmentation levels similar to control cells, which both increased under proapoptotic stimuli. Our data demonstrate that Magmas is overexpressed in mouse and human ACTH-secreting pituitary adenomas. Moreover, our results show that Magmas protects pituitary cells from apoptosis, suggesting its possible involvement in neoplastic transformation.

Expression of (Pro)renin Receptor in the Human Brain and Pituitary, and Co‐localisation with Arginine Vasopressin and Oxytocin in the Hypothalamus

(Pro)renin receptor [(P)RR], a specific receptor for renin and prorenin, is a 350 amino acid protein with a single transmembrane domain. In the present study, the expression of (P)RR in the human brain and pituitary, and its co-localisation with arginine vasopressin and oxytocin in the human hypothalamus were studied by quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and immunocytochemistry. Human brain and pituitary tissues were obtained at autopsy from the subjects without neurological or endocrinological disorders. The antiserum against (P)RR was raised in a rabbit by injecting the peptide fragment of human (P)RR corresponding to 224-237 amino acids conjugated with bovine serum albumin. Quantitative RT-PCR showed that (P)RR mRNA was widely expressed in every region of brain examined and pituitary, with the highest expression levels found in the pituitary and frontal lobe. Immunocytochemistry showed that (P)RR was expressed in the paraventricular and supraoptic nuclei of human hypothalami, and in anterior pituitary cells. Immunostaining of serial sections showed that (P)RR was co-localised with arginine vasopressin and oxytocin in the magnocellular neurones of the paraventricular and supraoptic nuclei. The preabsorption of the antibody by the antigen peptide abolished the immunostaining of (P)RR in the human hypothalamus. The present study has shown that (P)RR mRNA is widely expressed in the human brain and pituitary, consistent with the hypothesis that (P)RR is related to the various brain functions, such as cognitive function and brain development. Co-localisation of (P)RR with vasopressin in the hypothalamus raised the possibility that (P)RR may be related to the central control of water-electrolyte metabolism and blood pressure.

Mechanisms of FSH synthesis: what we know, what we don't, and why you should care

The pituitary gonadotropin hormones, FSH and LH, are key regulators of reproductive physiology. Though the two hormones are produced by the same cell type, often in response to the same endocrine and paracrine regulators, they sub-serve different biological functions and their synthesis and secretion are differentially regulated. This stems largely from differences in transcriptional, post-transcriptional, and post-translational regulation of their unique beta subunits. That is, both hormones are dimeric glycoproteins and share a common alpha subunit. Their unique beta subunits, however, derive from different genes encoding distinct proteins. Past and recent research indicates synthesis and release of the two hormones are subject to extensive and independent regulation. LH appears to be secreted predominantly via the regulated secretory pathway, whereas FSH release is largely constitutive. As such, investigations of FSH-beta subunit synthesis may lend direct insight into mechanisms underlying patterns of secreted FSH, more so than investigations of the LHbeta subunit. Here, we review recent investigations of transcriptional regulation of the FSH-beta subunit gene from different mammalian species, including humans. The results reveal both conserved and species-specific regulatory mechanisms that might contribute to inter-species variation in FSH release.

Increased Expression of Angiogenic Genes in the Brains of Mouse Meg3-Null Embryos

Maternally expressed gene 3 (MEG3) is a noncoding RNA highly expressed in the normal human brain and pituitary. Expression of MEG3 is lost in gonadotroph-derived clinically nonfunctioning pituitary adenomas. Meg3 knockout mice were generated to identify targets and potential functions of this gene in embryonic development and tumorigenesis. Gene expression profiles were compared in the brains of Meg3-null embryos and wild-type littermate controls using microarray analysis. Microarray data were analyzed with GeneSifter, which uses Kyoto Encyclopedia of Genes and Genomes pathways and Gene Ontology classifications to identify signaling cascades and functional categories of interest within the dataset. Differences were found in signaling pathways and ontologies related to angiogenesis between wild-type and knockout embryos. Quantitative RT-PCR and immunohistological staining showed increased expression of some Vascular Endothelial Growth Factor pathway genes and increased cortical microvessel density in the Meg3-null embryos. In conclusion, Meg3 may play an important role in control of vascularization in the brain and may function as a tumor suppressor in part by inhibiting angiogenesis.

Effect of concomitant progesterone administration or the effect of removal of estrogen capsule on changes caused by long-term estrogen treatment in pituitary VIP immunoreactivities

In the anterior pituitary besides the classical tropic hormones, peptides of a small molecular weight are also synthesized. One of them is the vasoactive intestinal polypeptide (VIP). VIP immunoreactivity is readily detected in human and monkey pituitaries; however, in the rat VIP immunoreactive cells were observed in about 50% of intact rats. In estrogen treated rats VIP immunoreactive cells were observed in the anterior pituitary of all animals. In this work, we have examined the effect of long-term sexual steroid treatments on the VIP immunoreactivity of the anterior pituitary using diethylstilbestrol (DES) or progesterone (P) filled capsules. The effectiveness of steroid treatments was tested by the measurement of plasma prolactin (PRL) level and by the appearance of prolactinoma. DES enhanced the plasma PRL level and 5months later it induced prolactinomas, the concomitant P treatment prevented both the elevation of plasma PRL level and the formation of prolactinomas. These results indicated that there was enough steroid in the capsules. There was a positive correlation between the duration of DES influence and the number of VIP immunoreactive cells. Two months after the implantation of DES there was a considerable number of VIP cells in the anterior pituitary, and 5months after implantation the number of VIP cells was greatly increased so as to form a VIP-oma. Concomitant implantation of P prevented the formation of VIP-oma. Two months after the implantation, the DES capsule was removed. Already 2months after removal the number of VIP cells approximated to the control level. It has been concluded that P can prevent the undesired effect of DES not only on the PRL, but on the VIP immunoreactivity as well.

Folliculo-stellate Cells of the Human Pituitary as Adult Stem Cells: Examples of Their Neoplastic Potential

Two unique, formerly unrecorded sellar neoplasms were observed in two women of 60 and 63 years of age. One lesion consisted of small epithelial cells and the other was a large-cell oncocytic tumor, yet they had the same simple cytoplasmic organization with dominance of polyribosomes and a sprinkle of glycogen. Striking markers shared by the neoplasms: (1) network of typical pituitary follicles, and (2) unexpected similarity to fetal human pituitary tissue at different gestational ages of 6 and 10–12 weeks. The latter showed appreciable endocrine differentiation. The assumed parent cell is the folliculo-stellate cell as pluripotent adult stem cell.

Maternally Expressed Gene 3 (MEG3) Noncoding Ribonucleic Acid: Isoform Structure, Expression, and Functions

Maternally expressed gene 3 (MEG3) is an imprinted gene highly expressed in the human pituitary. However, MEG3 expression is lost in human gonadotroph-derived pituitary adenomas and most human tumor cell lines. Expression of MEG3 in tumor cells results in growth suppression, p53 protein increase, and activation of p53 downstream targets. The MEG3 gene encodes a noncoding RNA of approximately 1700 nucleotides. There are 12 different MEG3 gene transcripts, generated by alternative splicing. They contain the common exons 1-3 and exons 8-10, but each uses one or more exons 4-7 in a different combination in the middle. MEG3 isoform expression patterns are tissue and cell type specific. Functionally, each isoform stimulates p53-mediated transactivation and suppresses tumor cell growth. We analyzed the secondary RNA folding structure of each MEG3 isoform, using the computer program mfold. All MEG3 RNA isoforms contain three distinct secondary folding motifs M1, M2, and M3. Deletion analysis showed that motifs M2 and M3 are important for p53 activation. Furthermore, a hybrid MEG3 RNA, containing a piece of artificially synthesized sequence different from the wild type but folding into a similar secondary structure, retained the functions of both p53 activation and growth suppression. These results support the hypothesis that a proper folding structure of the MEG3 RNA molecule is critical for its biological functions. This study establishes for the first time the structure-function relationship of a large noncoding RNA and provides a first look into the molecular mechanisms of the biological functions of a large noncoding RNA.

Study on chang of histopathology and molecular biology of NPY in human pituitary adenomas

Objective To investigate the localization of neuropeptide Y(NPY) in the subcellular structure and expression of NPY in human pituitary adenomas, and their specificity and relationship. Method The tissues of pituitary adenomas were obtained from 57 patients with pituitary adenomas who underwent surgery in the department of neurosurgery in General hospital of Tianjin Medical University. The localization of NPY in pituitary adenoma cells was studied by immuno-electron microscopy. The expression of NPY in all patients with pituitary adenomas were studied by immunohischemical analysis and RT-PCR. Results (1)Immuno-electron microscopy demonstrated that the positive NPY was located mainly in the secretory granulas in cell matrix of pituitary adenomas. Gold particles were sparsely detected in the rough endoplasmic reticulum and cell matrix. (2) Immunohistochemical analysis showed that NPY was immunodetected in 59.6% (34/57) patients with pituitary adenomas. NPY expression was significantly different among different types of pituitary adenomass (P<0.05). NPY was immunodetected in 91.7% of gonadotrophic adenomas, 66.7% of GH and ACTH adenomas, 50% of null cell adenoma and plurihormonal adenoma, 25% of prolaetinomas. There was significant difference among them(χ~2=11.64, P=0.04). (3) RT-PCR showed that there was signification differentiation (F=4.21, P<0.01) among the expression of NPY in different types of pituitary adenoma. The NPY expression in gonadotrophic adenomas was the highest (7.00±8.32),with 9.3-fold(q=5.2616, P<0.01)compared with prolactinomas(0.75±0.11),5.8-fold (q=5.0143, P<0.01) with non-thnctional adenomas (1.79±2.13),2.7-fold (q=4.07, P<0.05) with GH adenomas (2.59±3.06). There was no significant differentiation among the rest groups. Conclusions (1)NPY really exists in human pituitary adenomas. (2)NPY expression is significantly different among different types of pituitary adenomas. The NPY expression level in gonadotrophic adenomas is the highest, the lowest in prolactinomas. (3)These results will provide sound bases for further research of the action of NPY in human pituitary. Key words: Pituitary neoplasms; Neuropeptide Y; Subcellular structure; Gene

Immunohistochemical detection of somatostatin receptor subtype 5 (SSTR-5) in cushing adenoma

The actions of somatostatin hormone are mediated by specific transmembrane receptors (somatostatin receptors). These receptors are G-protein linked receptors encoded on five different chromosomes: SSTR1–SSTR5. At the protein level, SSTR-5 was found to have the highest level of expression in corticotroph adenomas [1, 2]. These data suggest that SSTR-5 may be a potential in situ target for medical therapies of adrenocorticotrophic hormone (ACTH) producing corticotroph adenomas [3]. As Cushing adenomas tend to be small and often are not identified by MR imaging [4–6], we hypothesized, based on the previous data, that SSTR-5 may be a potential target for localization of unidentified Cushing adenomas on MRI. Here, we present the results of a pilot study on immunohistochemical staining of 3 Cushing adenomas using the monoclonal SSTR-5 antibodies. Three paraffin blocks containing ACTH producing adenomatous and normal periadenomatous tissue of human pituitary were utilized. For immunohistochemistry, all mouse monoclonal anti-SSTR-5 antibodies were commercially obtained from Advanced Targeting Systems (San Diego, CA, USA). Paraffin sections were dewaxed in xylene and rehydrated through a graded series of ethanol, the antigen was retrieved in 0.01 M citrate buffer (with EDTA added) for 25 min, and the sections were incubated in 3% hydrogen peroxide for 5 min. Sections were then rinsed in TRIS buffer saline pH 7.8 twice for 3 min and blocked in mouse serum for 10 min. Sections were subsequently incubated for 30 min in mouse monoclonal SSTR-5 antibodies at 2ug/ml, rinsed in TRIS buffer saline twice for 3 min, incubated in anti-mouse IgG, biotinylated for 30 min, and rinsed again in TRIS buffer saline twice for 3 min. Sections were then incubated in strepavidin-HRP for 30 min, rinsed in TRIS buffer twice for 3 min and visualized with DAB (diaminobenzidine) for 5 min, counterstained with hematoxylin for 5 min, and cover slips were applied. The results were consistent in the 3 samples. Immunoreactivity was detected primarily in the cytoplasm and the cell membrane of the normal periadenomatous tissue. Immunopositivity was strongly correlated with ACTH-producing cells in the normal pituitary tissue as seen in slides immunostained for ACTH that had been obtained at diagnosis. Cushing adenoma was negative in all 3 samples (Fig. 1). In the literature, detection of somatostatin receptor subtypes was limited to qualitative or real-time PCR analysis of their mRNA [1, 2, 7–10]. Immunohistochemical detection and localization of somatostatin receptor subtypes has been reported in prostate tissue from patients with bladder outlet obstruction. To the best of our knowledge, there are no reports of immunohistochemical detection of SSTR-5 in ACTH-producing corticotroph adenomas. In conclusion, immunohistochemical staining was performed using monoclonal SSTR-5 antibodies on three specimens of ACTH-producing adenoma and their adjacent normal periadenomatous tissue. Our results demonstrated negative immunoreactivity for SSTR-5 in the adenomatous tissue. Studies on SSTR-5 epitopes are warranted before any trial of targeting the receptor for either localization or treatment of Cushing adenomas. W. Hassaneen (&) D. P. Cahill G. N. Fuller N. B. Levine The University of Texas, M. D. Anderson Cancer Center, Houston, TX, USA e-mail: whassaneen@mdanderson.org

Expression of mRNA for human type-I LHRH receptor transcript forms in human benign prostatic hyperplasia

The presence of four different isoforms of luteinizing hormone-releasing hormones (LHRH) and one LHRH receptor (LHRH-R) has been reported in vertebrates. In the human genome only LHRH-I and LHRH-II genes have been identified. The human LHRH-I gene is composed of four exons separated by three introns. Three LHRH receptor or receptor-like genes have been demonstrated. The well-established type-I LHRH receptor (LHRH-R-I) gene is composed of three exons separated by two introns. In this study we investigated the expression of transcript forms of LHRH-R-I in human benign prostatic hyperplasia (BPH) with reverse transcriptase-polymerase chain reaction (RT-PCR) using gene specific primers. Thirty-five human BPH specimens were obtained at surgery. Normal human pituitaries collected at autopsy served as control. RNA extraction and RT-PCR with gene-specific primers for LHRH-R-I forward (F1)/reverse (R1), LHRH-R-I F2/R3, LHRH-R-I F1'/R2' were carried out to determine the mRNA expression for LHRH-R-I transcript forms. The expected PCR products amplified with gene specific primers were LHRH-R-I F1/R1 with 319 bp, LHRH-R-I F2/R3 with 309 bp and LHRH-R-I F1'/R2' with 219 bp. PCR products for LHRH-R-I F1/R1 were detected in 21 (60%) and for LHRH-R-I F2/R3 in 5 of 35 (14%) BPH samples. No PCR products for LHRH-R-I F1'/R2' were found. In conclusion, we detected mRNA for LHRH-R-I in human BPH specimens. Our results suggest that LHRH-R-I gene may have more than two splice variants or uncharacterised transcript forms of LHRH-R-I. Our findings support the merit of further investigation of the expression of LHRH-R-I and its transcript forms in human BPH.

OMEGA‐3 POLYUNSATURATED FATTY ACIDS AFFECT LEPTIN RECEPTOR GENE EXPRESSION IN PITUITARY GH4C1 CELL LINE

ABSTRACTLeptin (Ob) and Ob receptor (Ob‐Rb) genes are expressed in human and rodent anterior pituitary, and an autocrine/paracrine loop in the production and regulation of Ob and Ob‐Rb in this tissue has been suggested. As brain membranes contain large amounts of polyunsaturated fatty acids (PUFAs), and PUFA may affect the levels of leptin as well as the receptor's environment, a possible regulatory role of exogenous PUFA on Ob and Ob‐Rb expression in pituitary cell lines was investigated. GH4C1 cells were incubated with arachidonic (ARA), eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids and fatty acid incorporation, viability, and gene expression were measured. The results showed that although fatty acids were incorporated in GH4C1 cells at a minimum level, positively affected cell viability without differences, among the different fatty acids. On the contrary, ARA, EPA and DHA showed different effects on Ob‐Rb gene regulation that was upregulated by EPA, downregulated by DHA and not affected by ARA. The effects of leptin co‐incubation are also reported and discussed.PRACTICAL APPLICATIONSData from this paper consolidate the observations regarding the role of n‐3 polyunsaturated fatty acid in gene expression regulation, and thus they could support the recommendation to enrich diet with fish or fish oil with high PUFA content.

Immunoreactive human chorionic gonadotropin beta core fragment in human pituitary.

The human pituitary has been shown to produce small amounts of immunoreactive human chorionic gonadotropin (hCG) and its alpha- and beta- subunit (hCG alpha, hCG beta). The aim of the present studies was to further examine the various hCG-related materials in the human pituitary, particularly to search for the existence of pituitary hCG beta core fragment (hCG beta cf)--like material. HCG beta cf has been found in the urine of pregnant women, patients with trophoblastic tumors and also in postmenopausal women. Gel chromatography of pituitary extract on Superdex 200 showed three distinct peaks of hCG-related immunoreactivities, i.e. hCG, hCG beta and hCG beta cf, which were distinguishable from hLH and hLH beta peaks. HCG beta cf was recognized by a specific immunometric assay (crossreactivities with hCG beta 0.016%, hLH beta 0.04%), but unreactive in an hLH + hLH beta assay. It was purified and displayed physical properties similar to those of hCG beta cf derived from pregnancy urine. Apart from immunological differences between the small molecular weight forms or fragments of hCG and LH origin, reversed phase HPLC was able to physically discriminate between hCG beta cf and hLH beta fragment. The latter was much more abundant than the former in the pituitary extract. HCG beta cf showed microheterogeneity related to its sialic acid content. In conclusion, the present data indicate that immunoreactive hCG beta cf is present in human pituitary extracts. The physical and immunological properties of pituitary hCG beta cf are distinguishable from those of the more abundant hLH beta and its fragment, and compare favorably with those of urinary hCG beta cf of trophoblastic origin.

Recent advances in the molecular biology of growth-hormone secreting human pituitary tumours.

The application of DNA technology has led to significant progress in our understanding of the aetiology of GH-secreting pituitary tumours. X-chromosome inactivation and RFLP analysis has revealed that GH-secreting tumours are monoclonal in origin and thus arise as a consequence of a somatic mutation within a single cell. Using PCR and sequence analysis, such a mutation has been identified in 30-40% of tumours. This is the so-called gsp mutation in which the gene for the alpha s subunit of the Gs protein is converted to an oncogene the expression of which results in constitutive adenyl cyclase activity and thus excessive cAMP production. In our own studies, we demonstrate that it is unlikely that a defect within the promoter region of the GH gene will prove to be a cause of excessive GH secretion in acromegaly. In contrast, we have shown that the GH gene is hypomethylated in tumour derived DNA and this may account, at least in part, for abnormal GH gene expression.

Expression of leptin receptor mRNA and the long form splice variant in human anterior pituitary and pituitary adenoma.

Expression of leptin receptor (OB-R) mRNA was detected in the human anterior pituitary as well as in ACTH-secreting and nonsecreting pituitary adenomas by RT-PCR with primers recognizing all receptor splice variants. Primers specific to the long splice variant of the leptin receptor (OB-Rb), containing the putative intracellular signalling domain, also revealed a strong expression in normal and adenomatous anterior pituitaries. These results indicate that the pituitary is a possible target tissue of leptin action and might be involved in leptin regulation of pituitary hormone secretion.

Evidence for a progenitor cell population in the human pituitary

The ability to isolate and propagate adult stem/progenitor cells from the human brain opens novel avenues for cell replacement therapy. This will also apply to the pituitary gland, i.e., following tumor induced endocrine deficiency. Herein, we examine autopsy derived pituitaries to unravel a putative stem/progenitor cell population in humans. In tissue sections of the anterior lobe nestin immunoreactive cells co-expressing smooth muscle actin (SMA) were identified in the perivascular space, indicating a pericytic differentiation. Under clonal conditions, this particular cell population generated primary and secondary cell aggregates (spheres). Pituitary cell cultures maintained a stable cell cycle length with a doubling time of 10 days for over eight months. Forskolin treatment induced a prolactin-expressing phenotype in the majority of cell progenies as well as few betaIII-tubulin (Tuj1) expressing cells of putative neuronal lineage. The presence of sphere-forming, nestin-immunoreactive cells and their ability to generate differentiated cell lineages indicates the existence of a progenitor cell population persisting in the adult human pituitary. Further studies are needed to characterize this cell population in more detail and to clarify their potential to initiate neoplastic transformation for example in the cellular pathogenesis of pituitary adenoma.

Folate receptor α regulates cell proliferation in mouse gonadotroph αT3-1 cells

We have previously found that the mRNA and protein levels of the folate receptor alpha (FRα) are uniquely over-expressed in clinically human nonfunctional (NF) pituitary adenomas, but the mechanistic role of FRα has not fully been determined. We investigated the effect of FRα over-expression in the mouse gonadotroph αT3-1 cell line as a model for NF pituitary adenomas. We found that the expression and function of FRα were strongly up-regulated, by Western blotting and folic acid binding assay. Furthermore, we found a higher cell growth rate, an enhanced percentage of cells in S-phase by BrdU assay, and a higher PCNA staining. These observations indicate that over-expression of FRα promotes cell proliferation. These effects were abrogated in the same αT3-1 cells when transfected with a mutant FRα cDNA that confers a dominant-negative phenotype by inhibiting folic acid binding. Finally, by real-time quantitative PCR, we found that mRNA expression of NOTCH3 was up-regulated in FRα over-expressing cells. In summary, our data suggests that FRα regulates pituitary tumor cell proliferation and mechanistically may involve the NOTCH pathway. Potentially, this finding could be exploited to develop new, innovative molecular targeted treatment for human NF pituitary adenomas.

Rab3B Immunoexpression in Human Pituitary Adenomas

Rab3B, a member of the Rab family is a low molecular weight GTP-binding protein that has been implicated in the regulation of exocytosis. To shed light on its presence in the normal human pituitary and in adenomas, a detailed immunohistochemical study of 130 surgically removed human pituitary adenomas was undertaken, including 23 somatotroph, 32 lactotroph, 19 functional corticotroph, 10 silent subtype 1 and 8 silent subtype 2 corticotroph adenomas, 12 gonadotroph hormone producing, 10 thyrotroph, 7 silent subtype 3 adenomas, and 9 null cell adenomas, 5 of the latter being of oncocytic type. Among the 32 prolactin lactotroph adenomas, 10 had been treated preoperatively with bromocriptine, a dopamine agonist. Among the 23 somatotroph adenomas, 10 were pretreated with octreotide, a long acting somatostatin analog. In addition, 10 nontumorous adenohypophyses were also examined. As used by the World Health Organization, the tumors were classified on the basis of their histologic, immunohistochemical, and ultrastructural characteristics. The results showed Rab3B immunopositivity to be strongest in corticotroph adenomas followed by thyrotroph, lactotroph, gonadotroph, null cell, and somatotroph adenomas. No difference was noted between endocrinologically active and silent corticotroph adenomas. Bromocriptine therapy was associated with decreased Rab3B immunoexpression, whereas pretreatment with octreotide induced no significant reduction. Immunopositivity was cytoplasmic and was evenly distributed. No staining was noted in normal adenohypophyses. Our results add new information to the view that Rab3B is involved in the regulation of pituitary hormone secretion.

EFFECTS of growth hormone from human pituitaries.

Journal Article Effects of Growth Hormone from Human Pituitaries Get access Nutrition Reviews, Volume 16, Issue 8, August 1958, Pages 253–255, https://doi.org/10.1111/j.1753-4887.1958.tb00790.x Published: 01 August 1958