The role of MAPK11/12/13/14 (p38 MAPK) protein in dopamine agonist-resistant prolactinomas.

BACKGROUND: Prolactinoma is a functional pituitary adenoma that secretes excessive prolactin. Dopamine agonists (DAs) such as bromocriptine (BRC) are the first-line treatment for prolactinomas, but the resistance rate is increasing year by year, creating a clinical challenge. Therefore, it is urgent to explore the molecular mechanism of bromocriptine resistance in prolactinomas. Activation of the P38 MAPK pathway affects multidrug resistance in tumours. Our previous studies have demonstrated that inhibiting MAPK14 can suppress the occurrence of prolactinoma, but the role of MAPK11/12/13/14 (p38 MAPK) signalling in dopamine agonist-resistant prolactinomas is still unclear. METHODS: A prolactinoma rat model was established to determine the effect of bromocriptine on MAPK11/12/13/14 signalling. DA-resistant GH3 cells and DA-sensitive MMQ cells were used, and the role of MAPK11/12/13/14 in bromocriptine-resistant prolactinomas was preliminarily verified by western blot, RT-qPCR, ELISA, flow cytometry and CCK-8 experiments. The effects of MAPK11 or MAPK14 on bromocriptine-resistant prolactinomas were further verified by siRNA transfection experiments. RESULTS: Bromocriptine was used to treat rat prolactinoma by upregulating DRD2 expression and downregulating the expression level of MAPK11/12/13/14 in vivo experiments. The in vitro experiments showed that GH3 cells are resistant to bromocriptine and that MMQ cells are sensitive to bromocriptine. Bromocriptine could significantly reduce the expression of MAPK12 and MAPK13 in GH3 cells and MMQ cells. Bromocriptine could significantly reduce the expression of MAPK11, MAPK14, NF-κB p65 and Bcl2 in MMQ but had no effect on MAPK11, MAPK14, NF-κB p65 and Bcl2 in GH3 cells. In addition, knockdown of MAPK11 and MAPK14 in GH3 cells by siRNA transfection reversed the resistance of GH3 cells to bromocriptine, and haloperidol (HAL) blocked the inhibitory effect of bromocriptine on MAPK14, MAPK11, and PRL in MMQ cells. Our findings show that MAPK11 and MAPK14 proteins are involved in bromocriptine resistance in prolactinomas. CONCLUSION: Bromocriptine reduces the expression of MAPK11/12/13/14 in prolactinomas, and MAPK11 and MAPK14 are involved in bromocriptine resistance in prolactinomas by regulating apoptosis. Reducing the expression of MAPK11 or MAPK14 can reverse bromocriptine resistance in prolactinomas.

Relation among Aromatase P450 and Tumoral Growth in Human Prolactinomas.

The pituitary gland is part of hypothalamic-pituitary-gonadal axis, which controls development, reproduction, and aging in humans and animals. In addition, the pituitary gland is regulated mainly by hormones and neurotransmitters released from the hypothalamus and by systemic hormones secreted by target glands. Aromatase P450, the enzyme responsible for the catabolization of aromatizable androgens to estrogens, is expressed in different parts of body, including the pituitary gland. Moreover, aromatase P450 is involved in sexual dimorphism where alteration in the level of aromatase can initiate a number of diseases in both genders. On the other hand, the direct actions of estrogens, mainly estradiol, are well known for stimulating prolactin release. Numerous studies have shown that changes in the levels of estrogens, among other factors, have been implicated in the genesis and development of prolactinoma. The pituitary gland can produce estradiol locally in several types of endocrine cells, and it is possible that aromatase could be responsible for the maintenance of the population of lactotroph cells and the modulation of the action of central or peripheral regulators. Aromatase overexpression due to inappropriate gene regulation has clinical effects such as the pathogenesis of prolactinomas. The present study reports on the synthesis of pituitary aromatase, its regulation by gonadal steroids, and the physiological roles of aromatase on pituitary endocrine cells. The involvement of aromatase in the pathogenesis of pituitary tumors, mainly prolactinomas, through the auto-paracrine production of estradiol is reviewed.

How effective is temozolomide for treating pituitary tumours and when should it be used?

Temozolomide (TMZ) has been shown as an effective treatment option in aggressive pituitary adenomas and carcinomas. This review analyses the published case series and demonstrates 42 % of patents show a radiological response and 27 % experience stable disease following TMZ. Prolactinomas and corticotroph tumours respond best to TMZ, showing approximately a 50 % response rate, with non-functioning tumours responding only half as frequently. Other factors that may predict the tumour's TMZ response include MGMT and MSH status, but neither is sufficiently robust to determine treatment decisions. TMZ has an accepted role in treating pituitary carcinoma and adenomas if radiation and surgery have failed to control tumour growth. To use TMZ on the basis of anticipated future aggression, as a primary therapy, or in preference to radiotherapy remains controversial.

Matrix metalloproteinase-9 and tissue inhibitor of matrix metalloproteinase-2: Prognostic biological markers in invasive prolactinomas.

In this study, the predictive roles of matrix metalloproteinase (MMP)-9 and tissue inhibitor of matrix metalloproteinase (TIMP)-1 and 2 in invasive and noninvasive prolactinomas were examined. Prognostic biomarkers to distinguish between invasive and noninvasive pituitary adenomas are required for the effective treatment of pituitary adenoma patients. We analyzed 57 prolactinoma patients classified as having invasive and noninvasive adenomas for MMP-9, TIMP-1 and TIMP-2 expression using immunohistochemical methods. Significantly higher MMP-9 expression was detected in invasive prolactinomas (p=0.004). There was also a significant relationship between TIMP-2 expression and invasive behavior (p=0.004) and TIMP-2 expression and recurrence (p=0.005). Because MMP-9 expression is significantly increased in invasive prolactinomas, MMP-9 has potential as a marker for invasion. TIMP-2 may be a marker for both invasion and recurrence. These findings require further examination in large scale prospective studies.

Aromatase cytochrome P450 enzyme expression in prolactinomas and its relationship to tumor behavior.

The aim of the study was to evaluate the presence of aromatase cytochrome P450 enzyme (P450AROM) expression in normal pituitary tissues and tumor tissues of patients with prolactinoma and to examine the impact of the P450AROM expression on clinical outcome. Twenty-six consecutive human pituitary tissue samples were obtained from autopsies performed at the Institute of Forensic Medicine. Sixty-four patients who had an adenomectomy between 2000 and 2009 after prolactinoma diagnosis with histologically confirmed pituitary tumor tissues were retrospectively included in this study. The slices from the pituitary tissues were subjected to immunohistochemical staining for evaluation of P450AROM and estrogen receptor beta (ER beta) subunit. Immunohistochemistry results were compared according to age, gender, remission rate, resistance and invasion status of the patients. Higher than normal P450AROM expression was found in the pituitary tissues of the patients with prolactinoma (p < 0.001). P450AROM intensity had no relation to resistance or remission in patients with prolactinoma (p = 0.44, p = 0.45, respectively). The subgroup analysis showed that compared to males without invasive adenoma, males with invasive adenoma had higher P450AROM expression (p = 0.048). ER beta was found to have an impact on resistance (p = 0.049). This study shows that P450AROM expression is present in the pituitary tissues of patients with prolactinoma and that this presence could be important in development and tumor behavior of prolactinomas.

Loss of 15-hydroxyprostaglandin dehydrogenase indicates a tumor suppressor role in pituitary adenomas.

15-hydroxyprostaglandin dehydrogenase (15-PGDH) may function as a tumor suppressor that antagonizes the action of the cyclooxygenase-2 (COX-2) oncogene in several types of tumors. However, it is unknown if it has a role in the pituitary. Recently, our group found that 15-PGDH expression was low in prolactin (PRL) secreting adenomas (prolactinomas) and growth hormone (GH) secreting adenomas (GHomas) using fiber-optic BeadArray technology. In this study, we examined the relative expression of 15-PGDH and COX-2 mRNA in clinical specimens and examined the effects of 15-PGDH on GH3 rat pituitary tumor cell proliferation, apoptosis and hormone secretion. 15-PGDH expression was lower and COX-2 expression was higher in prolactinomas and GHomas compared with normal controls. Overexpressed 15-PGDH inhibited tumor cell proliferation and induced apoptosis. It had a significant suppressive effect on mRNA levels and on the secretion of PRL and GH in GH3 cells. The inhibition of cell proliferation was accompanied by the decreased expression of cox-2, matrix metalloproteinase-9 (MMP-9) and B cell leukemia/lymphoma-2 (Bcl-2). These data are suggestive of a previously unrecognized pathway in pituitary tumorigenesis, and this novel observation may shed light on therapeutic strategies for pituitary tumors.

Bromocriptine induces parapoptosis as the main type of cell death responsible for experimental pituitary tumor shrinkage.

Bromocriptine (Bc) produces pituitary tumoral mass regression which induces the cellular death that was classically described as apoptosis. However, recent works have related that other mechanisms of cell death could also be involved in the maintenance of physiological and pathological pituitary homeostasis. The aim of this study was to evaluate and characterize the different types of cell death in the involution induced by Bc in experimental rat pituitary tumors. The current study demonstrated that Bc induced an effective regression of estrogen induced pituitary tumors by a mechanism identified as parapoptosis. This alternative cell death was ultrastructurally recognized by extensive cytoplasmic vacuolization and an increased cell electron density, represented around 25% of the total pituitary cells counted. Furthermore, the results obtained from biochemical assays did not correspond to the criteria of apoptosis or necrosis. We also investigated the participation of p38, ERK1/2 and PKC delta in the parapoptotic pathway. An important observation was the significant increase in phosphorylated forms of these MAPKs, the holoenzyme and catalytic fragments of PKC delta in nuclear fractions after Bc administration compared to control and estrogen treated rats. Furthermore, the immunolocalization at ultrastructural level of these kinases showed a similar distribution pattern, with a prevalent localization at nuclear level in lactotrophs from Bc treated rats. In summary, we determined that parapoptosis is the predominant cell death type involved in the regression of pituitary tumors in response to Bc treatment, and may cause the activation of PKC delta, ERK1/2 and p38.

Heregulin regulates prolactinoma gene expression.

To investigate the role of p185(her2/neu)/ErbB3 signaling in pituitary tumor function, we examined these receptors in human prolactinomas. Immunofluorescent p185(her2/neu) was detected in almost all (seven of eight), and ErbB3 expression in a subset (four of eight) of tumors (seven adenomas and one carcinoma). Quantitative PCR also showed abundant ErbB3 mRNA in tumor specimens derived from a rarely encountered prolactin-cell carcinoma. Activation of p185(c-neu)/ErbB3 signaling with heregulin, the ErbB3 ligand, in rat lacto-somatotroph (GH4C1) tumor cells specifically induced prolactin (PRL) mRNA expression approximately 5-fold and PRL secretion approximately 4-fold, whereas growth hormone expression was unchanged. Heregulin (6 nmol/L) induced tyrosine phosphorylation and ErbB3 and p185(c-neu) heterodimerization, with subsequent activation of intracellular ERK and Akt. The Akt signal was specific to ErbB3 activation by heregulin, and was not observed in response to epidermal growth factor activation of epidermal growth factor receptor. Gefitinib, the tyrosine kinase inhibitor, suppressed heregulin-mediated p185(c-neu)/ErbB3 signaling to PRL. Heregulin induction of PRL was also abrogated by transfecting cells with short interfering RNA directed against ErbB3. Pharmacologic inhibition of heregulin-induced phosphoinositide-3-kinase/Akt (with LY294002) and ERK (with U0126) signaling, as well as short interfering RNA-mediated mitogen-activated protein kinase-1 down-regulation, showed ERK signaling as the primary transducer of heregulin signaling to PRL. These results show ErbB3 expression in human prolactinomas and a novel ErbB3-mediated mechanism for PRL regulation in experimental lactotroph tumors. Targeted inhibition of up-regulated p185(c-neu)/ErbB3 activity could be useful for the treatment of aggressive prolactinomas resistant to conventional therapy.

Reduction of retinaldehyde dehydrogenase 1 expression and production in estrogen-induced prolactinoma of rat.

Retinoic acid (RA) plays a critical role in normal development and tissue maintenance and is also a regulatory factor of anterior pituitary cells. We previously demonstrated that a retinoic acid-synthesizing enzyme, retinaldehyde dehydrogenase 1 (RALDH1), is expressed in prolactin cells of adult rats and that estrogen suppressed RALDH1 expression. It is suggested that RA plays a role as a paracrine and/or autocrine signaling molecule in the anterior pituitary gland. However, the presence of RALDH1 in pituitary tumors has not been demonstrated. In this study, we examined the expression of RALDH1 in diethylstilbestrol-induced prolactinoma of LEXF RI rats. Quantitative analysis of mRNA levels by real-time PCR demonstrated drastic reduction of RALDH1 expression in the prolactinoma. We have also detected both mRNA expression and production by in situ hybridization and immunohistochemistry. Both mRNA-expressing cells and immunopositive cells remarkably decreased after 4 weeks of treatment with diethylstilbestrol. Fluorescence double immunohistochemistry of RALDH1 and prolactin revealed that prolactin-immunopositive cells do not colocalize with RALDH1 in the prolactinoma. These results suggest that the reduction of local RA generation relates to cell proliferation and tumorigenesis of lactotrophs.

Expression of aromatase P450 is increased in spontaneous prolactinomas of aged rats.

We have recently reported the presence of aromatase P450 in the rat hypophysis. This enzyme is responsible for the aromatization of testosterone to estradiol. Since the induction of prolactinomas has been demonstrated in the rat following chronic treatment with estradiol, the aim of the present study was to analyze whether a relationship exists between the presence of pituitary aromatase and the appearance of spontaneous prolactinomas in aged rats. Of a series of 90 adenomas studied, 53% showed prolactin immunoreactive cells and were classified as prolactinomas; only 33% of the adenomas were pure prolactinomas and the other 20% were multi-hormonal protactinomas. Moreover, 60% of the adenomas were aromatase-positive tumors. Interestingly, 100% of the pure prolactinomas were aromatase-positive while only 60% of the multi-hormonal prolactinomas expressed the enzyme. Western blotting with anti-aromatase antibodies revealed a 3.8-fold increase in expression of aromatase in pituitary tumors as compared to normal rat pituitary gland. Double immunohistochemical labeling detected the coexistence of prolactin and aromatase P450 in prolactinoma cells. ACTH- and LH-positive adenomas were considered as controls; only multi-hormonal ACTH and LH tumors display aromatase-positive cells and all of these also contained prolactin-positive cells. Our results demonstrate for the first time that aromatase is expressed in pituitary adenomas and that it is related to the functional nature of the tumor, especially in the case of pure prolactinomas, suggesting the possibility that an abnormally high conversion of testosterone into estradiol in pituitary cells may contribute to the genesis of spontaneous prolactinomas in aged rats.

Regulated, adenovirus-mediated delivery of tyrosine hydroxylase suppresses growth of estrogen-induced pituitary prolactinomas.

Prolactin-secreting adenomas are one of the most common types of intracranial neoplasm found in humans. The modalities of clinical treatment currently in use include D(2)-dopamine receptor agonists, surgery, and radiotherapy, and the success rates for treatment are good. However, there are prolactinomas that are difficult to treat. As an alternative, we have developed a gene therapy strategy in which the rate-limiting enzyme in dopamine synthesis, tyrosine hydroxylase (TH), is overexpressed in the anterior pituitary (AP) gland. Because dopamine is known to have an inhibitory effect on lactotroph growth and prolactin secretion, we developed a system that would enable its local synthesis from freely available precursor amino acids. A dual adenovirus tetracycline-regulatable expression system was generated to control the production of TH. In the absence but not presence of the tetracycline analog doxycycline, TH expression was observed in AP tumor cell lines AtT20, GH3, and MMQ. In both primary AP cell cultures and the AP gland, in situ expression of TH was seen in lactotrophs, somatotrophs, corticotrophs, thyrotrophs, and gonadotrophs in the absence but not presence of doxycycline. The ability of this system to inhibit hyperprolactinemia and pituitary lactotroph hyperplasia was then assessed in a model of estrogen- or estrogen/sulpiride-induced pituitary tumors. In the absence but not presence of doxycycline, a 49% reduction in pituitary growth and 58% reduction in the increase of circulating prolactin levels were observed in estrogen, but not estrogen/sulpiride, treated rats. These results indicate that in situ dopamine enhancement gene therapy can be a useful tool for the treatment of prolactinoma. Dopamine synthesis can be tightly regulated and the therapeutic benefit of the system is only inhibited when local dopamine signaling is impaired.

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.

Involvement of p38 mitogen-activated protein kinase activation in bromocriptine-induced apoptosis in rat pituitary GH3 cells.

Bromocriptine, a dopamine D(2) receptor agonist, is a therapeutic agent for patients with prolactinoma and hyperprolactinemia. In this study we demonstrated that bromocriptine induced activation of p38 mitogen-activated protein (MAP) kinase, with concomitant induction of apoptosis in rat pituitary adenoma cell line GH3 cells. Treatment of GH3 cells for 48 h with bromocriptine increased the p38 MAP kinase activity up to 3- to 5-fold and simultaneously increased the number of apoptotic cells. Inclusion in the medium of SB212090 or SB203580, specific p38 MAP kinase inhibitors, completely abolished the bromocriptine-induced activation of p38 MAP kinase and significantly reduced the number of apoptotic cells. The bromocriptine-induced p38 MAP kinase activation was not prevented by S(-)-eticropride hydrochloride, a specific D(2) receptor antagonist. Treatment with either epidermal growth factor (EGF) or thyrotropin-releasing hormone (TRH), which stimulates p44/42 MAP kinase, rescued cells from the bromocriptine-induced apoptosis, with concomitant inhibition of the bromocriptine-induced p38 MAP kinase activation. These results suggest that bromocriptine induces apoptosis in association with p38 MAP kinase activation, and that the p44/42 MAP kinase signaling through EGF and TRH receptors has an opposing effect on p38 MAP kinase activation as well as on apoptosis induced with bromocriptine in GH3 cells.

Mitogen-activated protein kinase activation by stimulation with thyrotropin-releasing hormone in rat pituitary GH3 cells.

We examined whether mitogen-activated protein (MAP) kinase is activated by thyrotropin-releasing hormone (TRH) in GH3 cells, and whether MAP kinase activation is involved in secretion of prolactin from these cells. Protein kinase inhibitors--such as PD098059, calphostin C, and genistein--and removal of extracellular Ca2+ inhibited MAP kinase activation by TRH. A cAMP analogue activated MAP kinase in these cells. Effects of cAMP on MAP kinase activation were inhibited by PD098059. TRH-induced prolactin secretion was not inhibited by levels of PD098059 sufficient to i activation but was inhibited by wortmannin (1 microM) and KN93. Treatment of GH3 cells with either TRH or cAMP significantly inhibited DNA synthesis and induced morphological changes. The effects stimulated by TRH were reversed by PD098059 treatment, but the same effects stimulated by cAMP were not. Treatment of GH3 cells with TRH for 48 h significantly increased the prolactin content in GH3 cells and decreased growth hormone content. The increase in prolactin was completely abolished by PD098059, but the decrease in growth hormone was not. These results suggest that TRH-induced MAP kinase activation is involved in prolactin synthesis and differentiation of GH3 cells, but not in prolactin secretion.

Neuronal nitric oxide synthase gene expression in human pituitary tumours: a possible association with somatotroph adenomas and growth hormone-releasing hormone gene expression.

OBJECTIVE: Nitric oxide (NO) has been implicated in the control of the secretory response to growth hormone-releasing hormone (GHRH) and may also modify GH release in response to excitatory aminoacids. Although rat and mouse pituitary cell lines have been shown to express neuronal NO synthase (nNOS), there has until now been no information on nNOS gene expression in human pituitary adenomas. Our objective was to provide such data and correlate the presence of nNOS transcripts with GHRH transcripts. PATIENTS: Pituitary adenoma tissue was obtained from a random selection of 32 patients with somatotrophadenomas, 16 patients with corticotroph adenomas, 39 patients with endocrinologically inactive adenomas and nine patients with macroprolactinomas undergoing transsphenoidal hypophysectomy. MEASUREMENTS: Transcripts for nNOS and GHRH were identified in frozen tissue sections by in situ hybridization histochemistry using synthetic 35S-labelled oligodeoxynucleotide probes with 100% homology to the target transcript. RESULTS: Neuronal NOS transcripts were identified in one of 16 corticotroph adenomas (6%), one of nine macroprolactinomas (11%), six of 39 endocrinologically inactive adenomas (15%) and 13 of 32 somatotroph adenomas (41%). GHRH transcripts were found in a similar distribution to nNOS transcripts in 10 of the 13 nNOS-expressing somatotroph adenomas, and in three of the four remaining adenomas from which suitable tissue was available. Cross-hybridization of the nNOS and GHRH probes to the same target was excluded by including rat brain sections cut through the arcuate nucleus as hybridization controls. Furthermore, two different nNOS oligodeoxynucleotide probes complementary to different regions of the target transcript produced identical results. CONCLUSIONS: These results suggest that there is a close correlation between nNOS gene expression and 'ectopic' expression of GHRH in human pituitary tumours, especially somatotroph adenomas. The relevance of these findings from a functional or pathologenic viewpoint remains unclear, but the data again emphasize that it is not just GH secretion that distinguishes somatotroph adenomas from other pituitary tumours.

[Immunohistochemical study on the expression of copper and zinc-superoxide dismutase (Cu, Zn-SOD) in human adenohypophysis and pituitary adenomas].

A location of copper and zinc- superoxide dismutase (Cu, Zn-SOD) in adenohypophysis and pituitary adenomas was examined with immunohistochemical technique. Pituitary adenomas include thirteen functioning, five nonfunctioning; functioning adenomas consist seven prolactinomas, four growth hormone (GH) secreting, two adrenocorticotropic hormone (ACTH) secreting adenomas. Three specimens of normal adenohypophysis were used for control study. The Cu, Zn-SOD was localized diffusely in the cytoplasm of normal adenohypophyseal cells and the tumor cells. Sometimes immunoreactive products of Cu, Zn-SOD revealed in the cytoplasm of endothelial cell, neutrophil, macrophage and the cell membrane of erythrocyte in the vessels. The content of Cu, Zn-SOD in normal adenohypophyseal cells and pituitary adenomas was markedly higher in normal cells than adenoma cells. No significant difference of the SOD content was observed not only in non-functioning adenoma but also in functioning adenoma cells including PRL, GH and ACTH cells.

Modulation of ornithine decarboxylase activity by prolactin in seminal vesicles of the rat.

The stimulatory effect of prolactin on sexual accessory glands is well established, though the mechanism of trophic action remains poorly understood. We have therefore assessed the effect of high levels of prolactin on ornithine decarboxylase activity and polyamine content in seminal vesicles (SV) of adult rats. Hyperprolactinaemia was induced by implantation of tissue fragments of a prolactin-secreting tumour 7315a, and the rats killed at 35 days post-inoculation. Serum levels of prolactin were increased significantly (p < 0.001) in tumour-bearing rats. Levels of testosterone in serum were reduced markedly, whereas LH levels remained unchanged. SV weight in the experimental group was not significantly different from that in the corresponding control group. A clear increase in ornithine decarboxylase activity of SV was observed (p < 0.001) in the hyperprolactinaemic rats. However, there was no change in the polyamine content of SV in tumour-bearing rats, compared to the control group. These results indicate that ornithine decarboxylase activity in SV of adult rats is regulated mainly by prolactin. This experimental model may be useful for identification of some of the events involved in the trophic action of prolactin.

The glutathione S-transferases: polymerase chain reaction studies on the frequency of the GSTM1 0 genotype in patients with pituitary adenomas.

The frequency of the GSTM1 0 polymorphism at the glutathione S-transferase M1 locus has been determined in controls and patients with pituitary adenomas by using the polymerase chain reaction to amplify genomic DNA in the exon 4-5 region of the gene. The frequency of the genotype in patients with prolactinomas, non-functional adenomas, corticotrophinomas and somatotrophinomas varied between 52-67% compared with 44% in the controls. In the patients with prolactinomas the frequency of the genotype (67%) was significantly greater than in controls with odds ratio analysis indicating that GSTM1 0 individuals have a 2.56-fold greater risk of developing this adenoma.

Immunohistochemical expression of protein kinase C type III in human pituitary adenomas.

Immunohistochemical expression of the three major isozymes of protein kinase C--Types I, II, and III--was studied in 32 cases of human pituitary adenomas, and the results were compared in detail with their clinical data. Immunoreactivity for the Type I and Type II isozymes was negative in tumor cells of all pituitary adenomas. Moderate to strong cytoplasmic immunoreactivity for the Type III isozyme was constantly seen in acromegaly, Cushing's disease, and nonfunctioning adenomas, which indicated overexpression of the isozyme, since only slight cytoplasmic immunoreactivity was observed in the normal human anterior pituitary cells. Among 13 prolactinomas, 5 cases showed positive immunoreactivity for Type III in all tumor cells, whereas 8 cases showed negative immunoreactivity for the isozyme in all or more than 85% of tumor cells. The sizes of the tumors in this protein kinase C Type III negative group of prolactinomas tended to be smaller than those of the Type III positive prolactinomas. Also, the negative immunoreactivity for Type III was predominantly observed in those cases where prolactinomas were relatively well controlled by continuous oral dosage of dopamine agonists before operation. These results suggest that protein kinase C Type III is closely involved in human pituitary adenomas. The exceptional negativity for the isozyme in prolactinomas may be relevant to the suppression of tumor growth by dopamine agonists.