Normal Anterior Pituitary Research Articles

Expression of members of the interleukin-6 family of cytokines and their receptors in human pituitary and pituitary adenomas.

There is increasing evidence for the role of members of the Interleukin (IL)-6 family in pituitary function, particularly in the regulation of the hypothalamo-pituitary-adrenal axis. However, there is only a limited amount of data available on the expression in human normal and tumorous pituitary tissue. In this study we investigated the expression of members of the IL-6 family of cytokines and their receptors in normal human pituitaries as well as pituitary adenomas using the RT-PCR technique. Eighteen pituitary adenoma biopsies removed in transsphenoidal surgery (six corticotrophic adenomas, four nonfunctioning adenomas, four somatotrophinomas, four prolactinomas) and six normal anterior pituitaries were examined for the expression of IL-6 receptor (-R), leukemia inhibitory factor (LIF), LIF-R, IL-11, IL-11-R, oncostatin M (OSM), OSM-R, ciliary neurotrophic factor (CNTF), CNTF-R and cardiotrophin-1 (CT-1). All pituitaries and pituitary adenomas expressed OSM transcripts, whereas no expression of CT-1 was found. The expression of all other cytokines (LIF, IL-11, CNTF) and receptors (IL-6-R, LIF-R, IL-11-R, OSM-R, CNTF-R) was found in different patterns in the adenoma subtypes and normal pituitaries. However, we did not detect expression of LIF-, IL-11-, IL-6-R and CNTF-R in prolactinomas, of CNTF in normal pituitaries and of OSM-R in ACTH-secreting adenomas. In conclusion, our study provides further evidence for a role of the members of the IL-6 family of cytokines in pituitary function.

Thyrotrophin receptor protein expression in normal and adenomatous human pituitary.

Thyrotrophin (TSH) synthesis and secretion is under the positive control of thyrotrophin releasing hormone and under the negative control of the thyroid hormones. However, it is hypothesised that TSH has a direct effect on the regulation of its own synthesis through an intrapituitary loop mediated by pituitary TSH receptors (TSH-R). The aim of this investigation was to study the expression of TSH-R in normal human pituitary at mRNA and protein levels, and to compare the pattern of protein expression between different pituitary adenomas. Using RT-PCR we were able to detect TSH-R mRNA in the normal pituitary, and immunohistochemical studies showed TSH-R protein expression in distinct areas of the anterior pituitary. Double immunostaining with antibodies against each of the intrapituitary hormones and S100 revealed that TSH-R protein is present in thyrotrophs and folliculostellate cells. Examination of 58 pituitary adenomas, including two clinically active and two clinically inactive thyrotroph adenomas, revealed TSH-R immunopositivity in only the two clinically inactive thyrotroph adenomas. This study shows, for the first time, the presence of TSH-R protein in the normal anterior pituitary and in a subset of thyrotroph adenomas. The expression of TSH-R in the thyrotroph and folliculostellate cell subpopulations provides preliminary evidence of a role for TSH in autocrine and paracrine regulatory pathways within the anterior pituitary gland.

Detection of gonadotropin-releasing hormone receptor in normal human pituitary cells and pituitary adenomas using immunohistochemistry.

Gonadotropin-releasing hormone (GnRH), which is a well-known regulator of gonadotroph function, has recently been considered to be a paracrine factor involved in the control of somatotroph, lactotroph, and corticotroph cells. GnRH action is initiated by binding to a specific cell surface receptor, the gonadotropin-releasing hormone receptor (GnRHR), which is expressed by follicle-stimulating hormone/luteinizing hormone (FSH/LH) cells. Using in situ hybridization techniques, GnRHR messenger ribonucleic acid (mRNA) has recently been detected in normal human anterior pituitary gland and in various pituitary adenomas, including FSH/LH-cell, growth hormone (GH)-cell, adrenocorticotropic hormone (ACTH)-cell, and null-cell adenomas. However, immunohistochemical studies indicating the specific cell distribution of GnRHR in normal pituitary cells have never been reported. The aim of the present investigation was to evaluate the immunohistochemical expression of GnRHR in different types of normal pituitary cells and related tumors. Using double-label immunohistochemical techniques on formalin-fixed and paraffin-embedded tissues and specific antibodies directed against pituitary hormones and GnRHR, we found GnRHR immunoreactivity not only in FSH/LH cells, but also in GH- and thyroid-stimulating hormone (TSH) cells. GnRHR was detected in FSH/LH-cell, GH-cell, mixed GH- and prolactin (PRL)-cell, and alpha-subunit (alpha-SU)/null-cell adenomas. The findings of this study suggest that the interaction between GnRH and GnRHR may play a role in paracrine/autocrine regulation of different types of normal pituitary cells and pituitary adenomas.

Expression analysis of cyclins in pituitary adenomas and the normal pituitary gland.

The molecular events involved in pituitary tumour development are still poorly understood. The cyclins play an important role in the control of the cell cycle during cell proliferation and over-expression of the cyclins has been shown in many different tumour types. The aim of this study was to investigate whether, in comparison to the normal gland, ectopic expression of cyclins occurs in pituitary tumours, and whether differences in cyclin expression are seen with different pituitary tumour types or in association with different tumour behaviour. In contrast to work on cyclin D there are no published data on cyclin B, A and E in human pituitary tumours. Sixty-seven surgically removed pituitary tumours and 10 specimens of normal human anterior gland were studied using immunohistochemistry to detect the nuclear expression of cyclin A, B, D and E. The microvascular density (as a measure of angiogenesis), Ki-67 labelling index (to assess cell proliferation) and bcl-2 expression had previously been investigated in this cohort. All tumours studied contained cells that immunostained positively for cyclin A, B, D and E. However the proportion of positive cells in each tumour type was different. In contrast, there were no cyclin D positive cells in the normal anterior pituitary gland studied, and labelling indices (LI) for cyclins A, B and E were significantly lower in the normal gland than in pituitary adenomas. The cyclin LIs for A, B, D and E were significantly higher in macroadenomas when compared to microadenomas. Non-functioning pituitary tumours (NFA) generally showed the highest cyclin LI. In particular, both recurrent and nonrecurrent NFA showed significantly higher cyclin D LI than other tumours. The ratio of cells expressing cyclin B compared to those expressing cyclin A was significantly higher in functionless tumours that regrew when compared to NFAs that did not (P<0.05). Cyclin D LI and the overall Ki-67 LI as a measure of cell proliferation were related (R2 = 11.4, P = 0.0033) and bcl-2 positive tumours had significantly higher cyclin D LI compared with bcl-2 negative tumours. There was a weak relationship between angiogenesis and the relative proportion of cells expressing D when compared to those in S phase (D/A ratio) (r2 = 10.5, P = 0.02). We have demonstrated that ectopic expression of cyclin D and over-expression of cyclins A, B and E, regulating different stages of the cell cycle is common in pituitary adenomas. In addition, cyclin expression was related to size and to pituitary tumour regrowth. The differences between functionless tumours that regrow and those that do not, may be due to reduced bcl-2 expression, increased cell proliferation, more cells at the G2/M stage (B/A ratio) and reduced cell differentiation with more aggressive subsequent tumour behaviour. Cyclin D expression and cell proliferation were related indicating that the cells entering the cycle become 'committed' to cell cycle progression. There was no relative over-expression of individual cyclins, and therefore no evidence of relative increase in cell cycle phase, indicating that the increased cyclin expression is more likely to be due to constant mitogenic stimulation rather than cell cycle regulatory failure. Although nuclear cyclin expression is a good marker of tumour growth and aggressive behaviour, the growth signal that leads to cyclin expression remains to be identified.

Role of matrix metalloproteinase 9 in pituitary tumor behavior.

The matrix metalloproteinases (MMPs) are a family of zinc-containing endopeptidases that are able to degrade the extracellular matrix and allow angiogenesis and tumor invasion. The vast majority of pituitary tumors are benign and do not metastasize to distant sites, although they may invade locally. The aim of this study was to determine whether expression of the collagenase MMP-9 may play a role in allowing angiogenesis and invasion by different pituitary tumor types. Tumor expression of MMP-9 was investigated using a monoclonal antibody on a series of well-characterized paraffin-embedded sections of pituitary tumors. Invasive macroprolactinomas (n = 11) were significantly more likely to express MMP-9 than noninvasive macroprolactinomas (n = 8) (P = 0.003). Invasive macroprolactinomas showed higher-density MMP-9 staining than noninvasive tumors (P < 0.05). MMP-9 expression did not differ between noninvasive tumors and normal pituitary gland, or between different sized prolactinomas. MMP-9 expression was related to aggressive tumor behavior. It was higher in invasive macroprolactinomas (P = 0.003) when compared with noninvasive macroprolactinomas or the normal anterior pituitary gland. In addition, although there was no difference in whether MMP-9 was present or not when nonfunctioning adenomas that recurred were compared with those that did not, samples of recurrent tumor at the second presentation were more likely to express MMP-9 (P = 0.01). Pituitary carcinomas were significantly more likely to be MMP-9 positive compared with normal anterior pituitary gland (P = 0.05), but there was no difference from invasive adenomas. Angiogenesis assessed by vascular density was related to MMP-9 expression (P < 0.05). In summary, we have shown the presence of MMP-9 expression in some invasive and recurrent pituitary adenomas, and in the majority of pituitary carcinoma. The mechanisms whereby MMP-9 expression influences tumor recurrence and invasiveness, and its association with angiogenesis, remains to be elucidated. However, these observations suggest that a future potential therapeutic strategy for some pituitary tumors may be administration of a synthetic MMP-9 inhibitor.

Angiogenesis in pituitary adenomas and the normal pituitary gland.

Angiogenesis is essential for tumor growth beyond a few millimeters in diameter, and the intratumoral microvessel count that represents a measure of angiogenesis has been correlated with tumor behavior in a variety of different tumor types. To date no systematic study has assessed pituitary tumors of different secretory types, correlating vascular count with tumor size. The vascular densities of pituitary tumors and normal anterior pituitary were therefore assessed by counting vessels labeled using the vascular markers CD31 and ulex europaeus agglutinin I. One hundred and twelve surgically removed pituitary adenomas (30 GH-secreting, 25 prolactinomas, 15 ACTH-secreting, and 42 nonfunctioning tumors) were compared with 13 specimens of normal anterior pituitary gland. The vascular counts in the normal anterior pituitary gland were significantly higher (P < 0.05) than those in the tumors using both CD31 and ulex europaeus agglutinin I. In addition, microprolactinomas were significantly less vascular (P < 0.05) than macroprolactinomas, although there was no such difference between vascular densities of microadenomas and macroadenomas producing GH. ACTH-secreting tumors were, like microprolactinomas, of much lower vascular density than the normal pituitary and other secreting and nonsecreting tumor types. In marked contrast to other tumors, pituitary adenomas are less vascular than the normal pituitary gland, suggesting that there may be inhibitors of angiogenesis that play an important role in the behavior of these tumors.

Adenovirus-mediated herpes simplex virus type-1 thymidine kinase gene therapy suppresses oestrogen-induced pituitary prolactinomas.

We tested the hypothesis that gene transfer using recombinant adenovirus vectors (RAds) expressing herpes simplex virus type 1 thymidine kinase (HSV1-TK) might offer an alternative therapeutic approach for the treatment of pituitary prolactinomas that do not respond to classical treatment strategies. HSV1-TK converts the prodrug ganciclovir (GCV) to GCV monophosphate, which is in turn further phosphorylated by cellular kinases to GCV triphosphate, which is toxic to proliferating cells. One attractive feature of this system is the bystander effect, whereby untransduced cells are also killed. Our results show that RAd/HSV1-TK in the presence of GCV is nontoxic for the normal anterior pituitary (AP) gland in vitro, but causes cell death in the pituitary tumor cell lines GH3, a PRL/GH-secreting cell line, and AtT20, a corticotrophic cell line. We have used sulpiride- and oestrogen-induced lactotroph hyperplasia within the rat AP gland as an in vivo animal model. Intrapituitary infection of rats bearing oestrogen-induced lactotroph hyperplasia, with RAd/ HSV1-TK and subsequent treatment with GCV, decreases plasma PRL levels and reduces the mass of the pituitary gland. More so, there were no deleterious effects on circulating levels of other AP hormones, suggesting that the treatment was nontoxic to the AP gland in situ. In summary, our results show that suicide gene therapy using the HSV1-TK transgene could be further developed as a useful treatment to complement current therapies for prolactinomas.

DEXAMETHASONE TREATMENT OF CONGENITAL ADRENAL HYPERPLASIA IN UTERO: AN EXPERIMENTAL THERAPY OF UNPROVEN SAFETY

Congenital adrenal hyperplasia (CAH) due to steroid 21-hydroxylase deficiency is a common cause of genital virilization in female infants resulting from inappropriate fetal adrenal androgen secretion. Some investigators have advocated treating pregnant women who are at risk for carrying a CAH fetus with dexamethasone to suppress fetal adrenal androgen synthesis. Experience to date shows that this treatment can be effective in ameliorating the genital virilization in female fetuses. However, the doses used are supraphysiological, the mechanism of dexamethasone action in the fetus is unclear and no long-term followup studies have been done. To be effective the treatment would need to be started by week 6 of gestation but the genetic diagnosis cannot be made until week 12. If the mother has had a previous CAH child, only 1 in 4 pregnancies will be affected and only the female fetuses stand to benefit from treatment, thus, 7 of 8 fetuses will be treated needlessly. In view of these and other concerns, the prenatal treatment of CAH remains an experimental therapy and, hence, must only be done with fully informed consent in controlled prospective trials approved by human experimentation committees at centers that see enough of these patients to collect meaningful data.

Galanin within the normal and hyperplastic anterior pituitary gland: localization, secretion, and functional analysis in normal and human growth hormone-releasing hormone transgenic mice.

Studies evaluating estrogen-induced anterior pituitary tumors revealed a strong direct correlation between expression of the peptide galanin and tumor growth. To evaluate further the potential roles of galanin in the hyperplastic pituitary, we used a model of estrogen-independent anterior pituitary tumor formation, the male human GH-releasing hormone (hGHRH) transgenic mouse. Pituitaries of hGHRH transgenic mice are characterized by a hyperplasia of somatotrophs and contain markedly elevated levels of galanin. We examined the population of galanin-producing pituitary cells in 4- to 6-month-old male hGHRH transgenic mice and their nontransgenic siblings. The percentage of galanin-containing pituitary cells was significantly increased within the anterior pituitaries of hGHRH transgenic mice. By using the cell immunoblot assay we found that the basal secretion of galanin and GH from individual pituitary cells of hGHRH transgenic mice was significantly greater than that from pituitary cells of nontransgenic mice. By modifying the cell immunoblot assay, we determined that somatotrophs from both hGHRH transgenic and normal mice that were positive for galanin immunoreactivity secreted significantly greater amounts of GH than those somatotrophs devoid of galanin immunoreactivity. Moreover, immunoneutralization of galanin significantly decreased GH secretion from pituitary cells obtained from hGHRH transgenic mice. Thus, we now show that the increased levels of galanin peptide within the hyperplastic pituitaries of hGHRH transgenic mice are due to an increase in the population of cells containing galanin, and that galanin participates in the augmented secretion of GH from hyperplastic proliferating pituitary cells.

Galanin within the Normal and Hyperplastic Anterior Pituitary Gland: Localization, Secretion, and Functional Analysis in Normal and Human Growth Hormone-Releasing Hormone Transgenic Mice

Galanin within the Normal and Hyperplastic Anterior Pituitary Gland: Localization, Secretion, and Functional Analysis in Normal and Human Growth Hormone-Releasing Hormone Transgenic Mice

Pituitary adenylate cyclase-activating polypeptide, interleukin-6 and glucocorticoids regulate the release of vascular endothelial growth factor in pituitary folliculostellate cells

There is increasing evidence that hormones play an important role in the control of endothelial cell function and growth by regulating the production of vascular endothelial growth factor (VEGF). VEGF regulates vascular permeability and represents the most powerful growth factor for endothelial cells. In the normal anterior pituitary, VEGF has been detected only in folliculostellate (FS) cells. In the present study, the regulation of the release of VEGF from FS-like mouse TtT/GF cells, and from FS cells of rat pituitary monolayer cell cultures was investigated using a specific VEGF ELISA. Basal release of VEGF was demonstrated in cultures of both TtT/GF cells and rat pituitary cells. Interestingly, the VEGF secretion was stimulated by both forms of pituitary adenylate cyclase-activating polypeptide (PACAP-38 and PACAP-27), indicating that this hypothalamic peptide regulates endothelial cell function and growth within the pituitary. VEGF secretion was also stimulated by interleukin-6 (IL-6) whereas basal, IL-6- and PACAP-stimulated secretion was inhibited by the synthetic glucocorticoid dexamethasone. The inhibitory action of dexamethasone was reversed by the glucocorticoid receptor antagonist RU486, suggesting that in FS cells functional glucocorticoid receptors mediate the inhibitory action of glucocorticoids on the VEGF secretion. The endocrine and auto-/paracrine control of VEGF production in pituitary FS cells by PACAP, IL-6 and glucocorticoids may play an important role both in angiogenesis and vascular permeability regulation within the pituitary under physiological and pathophysiological conditions.

Immunocytochemical demonstration of oncocytes in normal adenohypophysis.

Immunocytochemical examination for mitochondrial protein and cytochrome oxidase was performed to demonstrate oncocytes in normal adenohypophysis obtained from 28 patients of various age. A small number of solitary large epithelial cells showed intense cytoplasmic granular immunoreactivities for mitochondrial protein and cytochrome oxidase. The proportions of the cells positive for the former and the latter ranged from 0% to 5.9% (mean+/-SD; 1.5+/-1.7%) and from 0% to 4.9% (1.4+/-1.6%), respectively. These cells were either absent or extremely rare in young patients (under 10 years) but tended to increase in number with age (P < 0.0001). On the other hand, the mirror section technique showed that most of these cells were negative for adenohypophysial hormones, but a few of them were faintly positive for: alpha-subunit (8.0%), beta-subunits of follicle-stimulating hormone (4.8%), luteinizing hormone (2.5%), thyroid-stimulating hormone (1.0%), and growth hormone (0.5%), and were negative for prolactin and adrenocorticotropic hormone. We considered that these cells represent oncocytes that exist in varying numbers in normal adenohypophysis. It was suggested that oncocytes in normal adenohypophysis share various common features with tumorous oncocytes of pituitary oncocytomas.

Leukemia Inhibitory Factor as an Intrapituitary Mediator of ACTH Secretion

In the normal anterior pituitary, intercellular interactions are important for the expression of hormones including adrenocorticotropin (ACTH). Leukemia inhibitory factor (LIF) is secreted by anterior pituitary cells and stimulates both basal and corticotropin-releasing hormone (CRH)-stimulated ACTH secretion in AtT₂₀ cells. To determine the effects of LIF on normal pituitary cells, we measured the effects of LIF and an immunoneutralizing antiserum against LIF on ACTH secretion by cultured sheep anterior pituitary cells. In intact populations of anterior pituitary cells, LIF (10 nM) stimulated ACTH secretion (from 0.30 ± 0.06 to 0.77 ± 0.01 ng/well per 3 h) and antiserum to LIF (by itself) had no effect (0.29 ± 0.05 ng/well per 3 h). In marked contrast, following pharmacological elimination of CRH-target cells, a condition known to disinhibit ACTH secretion, basal ACTH secretion was elevated (0.74 ± 0.13 ng/well per 3 h); LIF produced no further stimulation (0.73 ± 0.22 ng/well per 3 h) but immunoneutralization of LIF significantly reduced secretion to 0.50 ± 0.10 ng/well per 3 h. Medium, conditioned by exposure to CRH-target-depleted cultures of anterior pituitary cells, increased net ACTH secretion (from 0.29 ± 0.03 to 6.54 ± 0.71 ng/well per 3 h), when added as a challenge to naive, cultured anterior pituitary cells. Inclusion of antiserum to LIF significantly attenuated (5.29 ± 0.62 ng/well per 3 h) this response. The presence in and secretion of LIF by normal individual pituitary cells was detected using immunocytochemical methods. Seven to 8% of all cells stained positively for LIF, with 66 ± 11% of those secreting detectable amounts of LIF under unstimulated conditions. LIF colocalized with TSH and LH in pituitary cells. Taken together, these data suggest that LIF can stimulate ACTH secretion by normal anterior pituitary cells, and potentially plays a role as an intrapituitary stimulator of ACTH secretion under certain conditions.

Estrogen downregulates neuronal nitric oxide synthase in rat anterior pituitary cells and GH3 tumors.

The anterior pituitary gland produces neuronal nitric oxide synthase (nNOS) and nitric oxide regulates secretion of various anterior pituitary hormones. Estrogen has many functions in anterior pituitary cells including stimulation of prolactin (PRL) cell proliferation and secretion of various anterior pituitary hormones. However, the role of estradiol-17beta (E2) in regulating pituitary nNOS expression has not been previously examined. We studied the regulation of nNOS in normal pituitaries, and neoplastic GH3 pituitary tumors in order to analyze the effects of E2 on nNOS in pituitary cells. GH3 tumors expressed higher levels of nNOS proteins compared to normal pituitaries. Estrogen downregulated nNOS mRNA and protein in both estrogen-treated pituitaries with PRL cell hyperplasia and in GH3 tumors implanted into the flank of rats treated with E2 in silastic tubing. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis demonstrated three alternatively spliced nNOS transcript isoforms--nNOSa, nNOSb, and nNOSc mRNAs--with distinct 5' untranslated first exons that arose from alternative splicing to a common second exon. All three spliced isoforms were found in the normal rat pituitary, whereas nNOSa and nNOSb, but not nNOSc, were expressed in GH3 tumors implanted into Wistar-Furth rats. E2 also downregulated the nNOSa alternative mRNA transcript isoform in vivo. These results indicate that the biological activity of nNOS in the normal rat anterior pituitary and in pituitary tumors is regulated by a complex pattern of alternative splicing and that some of these mRNA isoforms as well as nNOS protein are regulated by estrogen. Our results also indicate that the levels of nNOS and the alternatively spliced nNOS transcript between normal and GH3 pituitary tumors are different.

Growth hormone secretagogue receptor expression in human pituitary tumors.

The GH secretagogue (GHS) receptor (GHS-R) has been characterized and cloned. It is a member of a family of seven transmembrane receptors and is closely related to the neurotensin and TRH receptors. To determine the expression of this receptor in normal anterior pituitary and in 24 human pituitary adenomas, we analyzed GHS-R messenger ribonucleic acid (mRNA) using a RT-PCR assay. We found that normal human pituitary was positive for the GHS-R signal. In addition, all GH-secreting adenomas and the one TSH-secreting adenoma demonstrated the presence of GHS-R mRNA. Three of four ACTH-secreting tumors and three of nine gonadotroph adenomas were also positive for the GHS-R mRNA. To determine the amounts of GHS-R mRNA in normal pituitary and in representative tumors, semiquantitative competitive PCR was performed. We determined that normal pituitary had approximately 750 molecules/L GHS-R mRNA. The acromegalic tumor had approximately 1.5 x 10(5) molecules/L, and the TSH-secreting tumor had approximately 7.5 x 10(3) molecules/L. Other tumor types contained considerably less, with the ACTH-secreting and gonadotroph tumors expressing 7.5 x 10(2) and 3 x 10(2) GHS-R mRNA molecules/L, respectively. These results suggest that GH- and TSH-producing adenomas express GHS-R mRNA at levels 200 and 10 times higher, respectively, than the normal pituitary, and that this receptor expression may be involved in the pathogenesis and growth of these pituitary adenomas.

Differential regulation of MAP kinase activity by corticotropin-releasing hormone in normal and neoplastic corticotropes

Corticotropin-releasing hormone (CRH) plays an important role in regulating the development and function of hypothalamic-pituitary-adrenal axis. The mechanisms by which CRH regulates tissue-specific growth, differentiation and gene expression remain to be established. In the present study, we show that CRH differentially regulates MAP kinase activity in normal ovine anterior pituitary cells and mouse corticotrope AtT20 cells. Incubation of ovine normal anterior pituitary cells with CRH increased MAP kinase activity, an effect mimicked by cAMP and inhibited by the protein kinase A inhibitor H89. In contrast, incubation of mouse pituitary tumor AtT20 cells with CRH inhibited MAP kinase activity, an effect also mimicked by forskolin and inhibited by H89. This decrease in MAP kinase activity occurred with a time course similar to the increase seen in normal anterior pituitary cells. Furthermore, both effects of CRH on MAP kinase activity were inhibited by atrial natriuretic peptide (ANP). ANP also reversed the inhibition of DNA synthesis induced by CRH in AtT20 cells. Thus, CRH may differentially regulate cell growth in sheep normal anterior pituitary and mouse tumor corticotropes by modulating MAP kinase activity through a mechanism dependent on cAMP production and subject to regulation by ANP.

Mutation and expression analysis of corticotropin-releasing factor 1 receptor in adrenocorticotropin-secreting pituitary adenomas.

The present study was designed to investigate a possible role of CRF1 receptors (CRF1-R) in the pathogenesis of Cushing's disease. ACTH-secreting pituitary adenomas and nonsecreting pituitary adenomas have been analyzed for mutations in the CRF1-R gene by PCR and sequencing and been compared with the sequences of normal anterior pituitaries. No mutations affecting the CRF1-R protein have been found in all tumors analyzed. However, we found a significant overexpression of the CRF1-R messenger RNA in ACTH-secreting pituitary adenomas vs. inactive adenomas and normal pituitaries. We conclude that mutations of the CRF1-R are unlikely to be involved in Cushing's disease. We suggest that the overexpression of the CRF1-R messenger RNA may be related to a disturbed receptor regulation in ACTH-secreting pituitary adenomas.

Differential Effects of Somatostatin and Its Analog on Protein Tyrosine Kinases Activity in the Rat Pituitary and the Murine Colonic Tumors

The effects of the native somatostatin-14 (SST-14) and of its analog octreotide (OCT) on the activity of protein tyrosine kinases (PTK) in the normal rat anterior pituitary gland, diethylstilbestrol (DES)-induced rat pituitary tumor and murine colonic cancer Colon 38 were studiedin vitro.PTK activity was estimated in tissue homogenates using gamma-[32P]ATP and poly (Glu80, Tyr20) as a substrate. It was found that both SST-14 and OCT suppressed the PTK activity in all examined tissues. The suppressive effect was more pronounced in DES-induced pituitary tumor than in normal anterior pituitary gland, and in the former, OCT was more effective than SST-14. In contrast, SST-14 stronger suppressed PTK activity in colonic cancer than OCT. We hypothesize that SST-14 acts on PTK activity in colonic cancer mainly via SSTR-1 subtype of somatostatin receptors.

Galanin-like immunoreactive nerve fibres in the anterior pituitary of the normal and adrenalectomized rat

Our previous studies have shown the presence of substantial amounts of substance P and calcitonin gene-related peptide-like immunoreactive nerve fibres in the anterior pituitary of the monkey, dog and rat. Furthermore, synaptic relationships have been demonstrated between these nerve fibres and the gland cells in the dog and rat. The substance P and calcitonin gene-related nerve fibres increase in number following adrenalectomy and ovariectomy, respectively. The present study was aimed to investigate the galanin-containing nerve fibres in the anterior pituitary of normal and adrenalectomized rats. The results showed only a small amount of galanin-like immunoreactive nerve fibres in the normal anterior pituitary, which were present among the gland cells as well as along the blood vessels. Following adrenalectomy, the number of galanin-like immunoreactive nerve fibres increased and ramification appeared more frequently. The results substantiate our hypothesis of a dual neural-humoral regulation of the mammalian anterior pituitary gland.

Immunocytochemical demonstration of the expression and induction of manganese-superoxide dismutase in the adenohypophysis.

Immunocytochemistry for manganese-superoxide dismutase (Mn-SOD) was studied in 12 normal adenohypophyses and 38 various pituitary lesions. The proportions of cells with granular immunoreactivity for Mn-SOD in normal adenohypophysis ranged from 9.8% to 29.6% (mean +/- SD; 18.4+/-6.2%). Some positive cells tended to accumulate in clusters, distribution of which corresponded well with those immunopositive for mitochondrial protein and cytochrome oxidase. The number of Mn-SOD-positive cells increased in adjacent residual adenohypophysis in eight of nine recent infarcts, in two of five old infarcts, in all four cases of lymphocytic hypophysitis, in two of four abscess cases and in one of three metastatic tumour cases, whereas the immunoreactivities of mitochondrial protein- and cytochrome oxidase-positive cells either did not vary or decreased. The intensity of the histological inflammatory reactions showed a positive correlation with reactivity for Mn-SOD in these lesions. Of eight adenomas, the surrounding area of compressed adenohypophysis showed increased numbers of Mn-SOD- and mitochondrial protein-/cytochrome oxidase-positive cells in four and six cases respectively. It is suggested that positivity for Mn-SOD may be related to some functional activity of mitochondria. It is further suggested that adenohypophysial cells have a high potential to induce Mn-SOD by inflammatory and ischaemic stress and, in addition, by enhanced mitochondrial activity.