Suppressive effects of dehydroepiandrosterone and the nuclear factor-kappaB inhibitor parthenolide on corticotroph tumor cell growth and function in vitro and in vivo.

Dehydroepiandrosterone (DHEA) is believed to have an anti-tumor effect, as well as anti-inflammatory, antioxidant, and anti-aging effects. To clarify the possible inhibitory action of DHEA on pituitary tumor cells, we tested the effects of DHEA, alone or in combination with the nuclear factor-kappaB (NF-kappaB) inhibitor parthenolide (PRT), on AtT20 corticotroph cell growth and function both in vitro and in vivo. We found that, in vitro, DHEA and PRT had potent inhibitory effects on pro-opiomelanocortin and NF-kappaB-dependent gene expression. They also suppressed the transcription activity of survivin, a representative anti-apoptotic factor, and induced apoptosis in this cell line. Furthermore, using BALB/C nude mice with xenografts of AtT20 cells in vivo, we found that the combined administration of DHEA and PRT significantly attenuated tumor growth and survivin expression. The treatment also decreased the elevated plasma corticosterone levels and ameliorated the malnutrition induced by tumor growth. Altogether, these results suggested that combined treatments of DHEA and PRT potently inhibit the growth and function of corticotroph tumor cells both in vitro and in vivo. This effect may, at least partly, be caused by the suppressive effects of these compounds, such as survivin and other inhibitor of apoptosis proteins, on NF-kappaB-mediated gene transcription.

Primary amyloidosis with multiple pulmonary nodular lesions and IgA nephropathy-like renal involvement.

A 66-year-old woman demonstrated multiple nodular lesions in the lungs without symptoms, and laboratory tests and transbronchial lung biopsy (TBLB) had been negative for malignancy, tuberculosis and sarcoidosis 15 years ago. She developed proteinuria and hematuria 10 years later. Renal biopsy revealed focal segmental mesangial proliferation with predominant IgA deposition in the paramesangium, suggesting IgA nephropathy. However, electron-microscopic observation revealed 8-12 nm fibril deposits in the interstitium and few in the mesangium that were positively stained with amyloid P protein and negative for amyloid A protein. Re-evaluation of previous TBLB samples showed apple-green birefringence with Congo-red staining that was resistant to potassium permanganate reaction. Electron-microscopic observation with high magnification and immunostaining for amyloid components led to a diagnosis of AL amyloidosis in this patient with predominant mesangial IgA deposition and slowly progressive nodular lesions in the lungs.

Effects of adrenomedullin on adrenocorticotropic hormone (ACTH) release in pituitary cell cultures and on ACTH and oxytocin responses to shaker stress in conscious rat.

Adrenomedullin (AM) is a potent vasodilator peptide, which is initially isolated from tissue of human pheochromocytoma. In addition to the effect on cardiovascular system, previous studies suggest that AM plays some roles as a neuropeptide in the brain. In the present study, we examined the effect of AM on in vitro adrenocorticotropic hormone (ACTH) secretion stimulated by corticotropin-releasing hormone (CRH), vasopressin (VP) or oxytocin (OT) in cultured rat corticotrophs and on the response of plasma ACTH, corticosterone (B) and OT to shaker stress in vivo. In contrast to the previous report, basal or CRH (10(-9) M)-stimulated ACTH secretion was not affected by coincubation with AM. Either of VP (10(-8) M) or OT (10(-8) M) significantly increased ACTH secretion in cultured rat anterior pituitary cells (156.7+/-24.9 in basal incubation vs. 267.8+/-15.0 in VP-stimulation, P<0.05, and 308.6+/-41.3 pg/ml in OT-stimulation, P<0.05). AM (10(-10) M) significantly inhibited OT-stimulated ACTH secretion. AM tended to inhibit VP-stimulated ACTH secretion, although the inhibitory effect was not statistically significant. Thus, it is likely that AM attenuates OT-stimulated ACTH secretion in corticotrophs. In vivo study, male Wistar rats were prepared with a guide cannula in the lateral ventricle and a catheter in femoral artery for blood sampling. AM (0.5, 1.0 microg in 5 microl) or normal saline (5 microl, control) was intracerebroventricularly (i.c.v.) injected in conscious rats. Shaker stress (110 cycles/min for 5 min) produced a significant increase of plasma ACTH (baseline: 106.4+/-48.6; vs. just after stress: 388.9+/-56.1 pg/ml, P<0.05) and B (baseline: 198.6+/-46.8 vs. 15 min after stress: 378.5+/-13.6 ng/ml, P<0.05) in the control group. Plasma OT tended to increase after stress, although the change was not significantly different (baseline: 29.8+/-6.5; just after stress: 65.6+/-18.2 pg/ml). I.c.v. injection of AM at 3 min before the stress did not significantly affect stress-induced changes of plasma ACTH, B and OT. These results suggest that AM has an inhibitory effect on OT-induced ACTH release in vitro and the inhibitory effect may be overwhelmed in ACTH and B response to shaker stress.

Differential neuropeptide responses to starvation with ageing.

During starvation, counterregulatory responses to loss of food (i.e. responses that lead to an increase in appetite) occur in the central nervous system (CNS). This study was designed to examine whether middle-aged rats show greater or smaller behavioural, peripheral and central hormonal responses during starvation compared to young rats. In experiment 1, refeeding following 4 days of starvation was measured in both middle-aged (72-week-old) and young (9-week-old) rats. The level of refeeding was similar to each prestarved level until 3 days after the end of starvation in both groups. From the 4th day, the level of refeeding in young rats increased and reached beyond the prestarved level, whereas refeeding in middle-aged rats remained similar to the prestarved level. Thus, overall refeeding throughout 7 days was greater in young rats than in middle-aged rats. In experiment 2, middle-aged and young rats were starved for 4 days and were killed in the morning. Middle-aged rats showed a smaller plasma corticosterone response than that of young rats. The magnitude of decreases in plasma glucose, insulin and leptin was similar in both groups. In the arcuate nucleus, the starvation-induced increase in neuropeptide Y (NPY) mRNA and the decrease in proopiomelanocortin (POMC) mRNA were smaller in middle-aged rats than in young rats. In contrast, the starvation-induced decrease in corticotrophin-releasing hormone (CRH) mRNA in the hypothalamic paraventricular nucleus was greater in middle-aged rats than young rats. The magnitude of decrease in type-2 CRH receptor mRNA in the ventromedial hypothalamus was similar in both groups. The results indicate that (a) ageing impaired refeeding response (b), middle-aged rats showed the same directional neuropeptide mRNA responses as seen in young rats during starvation and (c) the magnitude of these counterregulatory responses in the CNS in middle-aged versus young rats was not uniform, but rather was site-specific or neuropeptide-specific. This study suggests the importance of NPY and POMC responsiveness in the arcuate nucleus in the age-related differences resulting from starvation-induced refeeding.

Lack of decrease in hypothalamic and hippocampal glucocorticoid receptor mRNA during starvation.

We have shown in a previous study that high corticosterone levels during repeated immobilization stress result in a reduction of glucocorticoid receptor (GR) mRNA in the hypothalamic paraventricular nucleus (PVN) and the hippocampus. The reduction of GR presumably accounts for loss of or decrease in glucocorticoid-negative feedback, and thus hyperfunction of the hypothalamic-pituitary-adrenocortical (HPA) axis persists during chronic stress. Starvation is a stress state in which the counterregulatory responses against the loss of food occur in the central nervous system. We explored the impact of starvation on the HPA axis, GR and mineralocorticoid receptor (MR) mRNAs in the hippocampus, the PVN, and the anterior pituitary (AP) of rats. Rats were starved for 4 days and sacrificed in the morning. Starved rats showed high levels of plasma corticosterone, whereas neither plasma corticotropin (ACTH), AP proopiomelanocortin (POMC) mRNA nor AP type-1 corticotropin-releasing hormone (CRH) receptor mRNA was altered in the starved rats. In the presence of high corticosterone, starvation resulted in a decrease in both CRH mRNA and type-1 CRH receptor mRNA in the PVN. Consistently, the starved rats did not show any changes in GR mRNA in the hippocampus (CA1-2, CA3, and dentate gyrus), the PVN or the AP despite the elevation of plasma corticosterone. A significant decrease in MR mRNA was seen in the dentate gyrus and the AP, but not in CA1-2, CA3 or PVN. The lack of reduction of GR may be one of the organism's counterregulatory responses during starvation, which allows an intact glucocorticoid negative feedback, thereby resulting in decreased anorectic neuropeptide levels, namely CRH, in the PVN. The results also indicate that GR mRNA in the hippocampus and other brain regions is not solely regulated by circulating glucocorticoids. The mechanism underlying the regulation of GR mRNA in the central nervous system remains to be clarified.

Physiological roles of corticotropin-releasing hormone receptor type 2.

Recent investigations of the physiological roles of CRH-R2 are reviewed and summarized in Fig. 5. VMH CRH-R2 is more important than CRH-R1 in mediating anorexic effect of CRH or urocortin (UCN) and stress-induced reduction of food intake. CRH-R2 mediates a central anxiolytic response, opposing the anxiogenic effect of CRH mediated by CRH-R1. Hippocampal CRH-R1 mediates stress-induced enhancement of learning, while CRH-R2 in the lateral intermediate septum may act to impair learning. CRH-R1 mediates CRH-induced blood pressure elevation, while peripheral CRH-R2 mediates the hypotensive effect of systemically administered UCN and CRH. It is likely that CRH-R2 does not play an important role in hypothalamic-pituitary adrenal axis regulation, though it has been reported that CRH-R2-deficient mice showed hyperresponse of ACTH and corticosterone. Peripheral CRH-R2 mediates UCN-induced mast cell degranulation, vascular permeability, and abdominal surgery-induced gastric stasis. These recent investigations have revealed that the existence of two CRH receptors, which mediate some opposite effects, provides the CRH and UCN systems a high flexibility and dynamic role in the adaptation of the body to environmental challenge.

Effect of non-peptide corticotropin-releasing factor receptor type 1 antagonist on adrenocorticotropic hormone release and interleukin-1 receptors followed by stress.

Previous studies demonstrated that ether-laparotomy significantly increased iodine-125-labeled interleukin-1alpha ([125I]IL-1alpha) binding in the mouse anterior pituitary at 2 h after the onset of stress. Corticotropin-releasing factor (CRF) receptor antagonist, D-Phe CRF (12-41), abolished ether-laparotomy-induced increase in [125I]IL-1alpha binding in the pituitary, showing that CRF plays a pivotal role in the regulation of IL-1 receptors under stress conditions. In an attempt to define the effect of CRA 1000 (2-(N-(2-methylthio-4-isopropylphenyl)-N-ethylamino-4-(4-(3-fluorophenyl)-1,2,3,6-tetrahydropyridin-1-yl)-6-methylpyrimidine), a non-peptide CRF receptor type 1 antagonist on the regulation of hypothalamic-pituitary-adrenal (HPA) axis and IL-1 receptors in the mouse, we measured plasma adrenocorticotropic hormone (ACTH) and corticosterone levels, [125I]IL-1alpha binding and the expression of transcripts for type 1 IL-1 receptor (IL-1R1 mRNA) in the pituitary at 2 h after endotoxin lipopolysaccharide (LPS) treatment or ether-laparotomy stress with or without CRA 1000 pretreatment. A single injection of LPS dramatically increased plasma ACTH and corticosterone levels compared with saline injection. In contrast, plasma ACTH levels were significantly attenuated in response to one LPS injection following oral CRA 1000 pretreatment. LPS-induced plasma corticosterone levels tended to be lower after CRA 1000 pretreatment but it did not reach statistical significance. Ether-laparotomy stress significantly increased plasma ACTH and corticosterone levels at 2 h after the onset of stress and CRA 1000 pretreatment did not affect the peak ACTH and corticosterone levels following stress. Ether-laparotomy stress resulted in a robust increase in [125I]IL-1alpha binding and IL-1R1 mRNA levels in the pituitary. CRA 1000 pretreatment significantly decreased ether-laparotomy stress-induced IL-1R1 mRNA levels but did not affect [125I]IL-1alpha binding. Pretreatment with CRA 1000 without stress significantly increased [125I]IL-1alpha binding and IL-1R1 mRNA levels compared with those in vehicle pretreatment. These data demonstrate differential effects of CRA 1000 in HPA axis following endotoxin and ether-laparotomy stress and complex interactions between CRF and IL-1 receptors during stress.

Antipituitary antibodies in patients with lymphocytic hypophysitis.

BACKGROUND: Lymphocytic hypophysitis is one of the causes of hypopituitarism, which is considered an autoimmune reaction in the anterior pituitary. METHOD: We examined antipituitary antibodies in patients with lymphocytic hypophysitis and related diseases by immunoblotting method. RESULTS: Autoantibodies to a 22-kDa human pituitary cytosolic protein were identified in significantly higher frequencies in sera from patients with lymphocytic hypophysitis (11 of 15, 73.3%) and isolated ACTH deficiency (7 of 9, 77.8%) compared with Hashimoto thyroiditis, Basedow's disease and normal control subjects. Also, reactivity against a 49-kDa human pituitary cytosolic protein was seen in 6 of 15 patients (40%) with lymphocytic hypophysitis. N-terminal amino acid sequences of 22-kDa human and rat pituitary cytosolic protein were FPTIPLSVL and FPAMPLSSLFAN, respectively, suggesting that they are human and rat growth hormone, respectively. The pituitary dysfunction (at least one hormone dysfunction) was observed in 11 of 14 patients. Nine of them (82%) showed 22 kDa antibody but 2 of them (18%) did not. CONCLUSION: The present study demonstrated that pituitary autoantibodies could be involved in the pathogenesis of lymphocytic hypophysitis and could be a positive marker for the disease.

Regulation of type-2 corticotropin-releasing hormone receptor mRNA in rat heart by glucocorticoids and urocortin.

A novel subtype of corticotropin-releasing hormone (CRH) receptor, designated type-2 CRH receptor (CRHR-2), has been cloned by a number of laboratories, and its mRNA has been found to be distributed not only in the brain but in peripheral tissues such as heart and skeletal muscle. To date, however, the regulation of CRHR-2 mRNA is poorly understood. Therefore, we examined the effect of glucocorticoid treatment, adrenalectomy, and systemic administration of urocortin, a possible endogenous ligand for CRHR-2, on heart CRHR-2 mRNA levels in male Wistar rats, using in situ hybridization histochemistry. CRHR-2 mRNA in the heart was significantly decreased 9 h after systemic administration of urocortin (5 microg/kg b.w.). Systemic administration of corticosterone (CORT; 10 mg/rat/day for 12 days) or CORT pellet (200 mg) implant for 7 and 14 days also decreased CRHR-2 mRNA in the heart, whereas it was unchanged 7 days after adrenalectomy. Thus, similar regulation of CRHR-2 mRNA in the rat heart by its ligand and glucocorticoids was observed. The precise mechanism of the regulation of CRHR-2 mRNA in the heart and the physiologic significance of cardiac CRHR-2 remains to be elucidated.

A case of lymphocytic infundibuloneurohypophysitis showing diabetes insipidus followed by anterior hypopituitarism associated with thrombasthenia.

We report a case of a 42-year old male patient with diabetes insipidus followed by anterior hypopituitarism associated with thrombasthenia. The patient had been diagnosed with thrombasthenia since the age of 19. He was admitted and diagnosed as diabetes insipidus in 1995. Although T1-weighted image of magnetic resonance imaging (MRI) showed empty sella and partial pituitary stalk hypertrophy, the anterior pituitary functions were normal at that time. Three years later, he was re-admitted after an episode of general malaise and impotence in 1998. Endocrinological studies revealed adrenal insufficiency, hypothyroidism and hypogonadism. T1-weighted image of MRI demonstrated the thickening of pituitary stalk and neurohypophysis. Analysis of anti-pituitary antibodies by immunoblotting identified a major band at 61.5 kDa. The diabetes insipidus was controlled by desmopressin acetate and the shrinkage of pituitary stalk was seen after hormonal replacement therapy including glucocorticoid and thyroid hormone. We suggested that this case represented lymphocytic infundibuloneurohypophysitis, in which a chronic inflammatory process occurred in infundibulum and/or neurohypophysis and that hypopituitarism developed possibly due to damage to the pituitary portal vessels caused by a thickened pituitary stalk, although a pituitary biopsy was not done because of the risk of bleeding in thrombasthenia. The pituitary autoantibodies in sera from patients with hypopituitarism may be helpful to characterize the patient with lymphocytic hypophysitis.

Possible reproductive toxicity of styrene in peripubertal male mice.

Environmental estrogens (endocrine disruptive chemicals) have been shown to affect reproduction in wild life and it has been reported that maternal exposure to those chemicals has adverse effects on the male reproductive tract. However, little is known about the potential effects of prepubertal or pubertal exposure to environmental estrogens on the male reproductive tract. Here we examine plasma hormone levels of mice following 4-week oral administration of styrene. Plasma free testosterone levels were dramatically decreased following 4 weeks of styrene treatment compared with control group. No differences in plasma corticosterone and luteinizing hormone levels were seen between styrene and control groups. Thus, exposure to styrene around pubertal period may directly disrupt the male reproductive tract. These facts suggest that more detailed studies regarding assessment of the risk to the developing human testes from exposure to styrene and other environmental estrogens in prepubertal and pubertal period are warranted.

Effect of orchidectomy on the age-related modulation of IL-1beta and IL-1 receptors following lipopolysaccharide treatment in the mouse.

OBJECTIVE: To compare the effect of orchidectomy (ODX) in 7- and 24-week-old C57BL/6 mice on the age-related responses of the cytokine interleukin (IL)-1beta and its receptor to intraperitoneal injection of the bacterial endotoxin, lipopolysaccharide (LPS). METHODS: We measured IL-1beta concentrations in the plasma, hippocampus, hypothalamus and adrenal gland using ELISA and iodine-125-labeled recombinant human IL-1alpha ([(125)I]IL-1alpha) binding in the hippocampus following the intraperitoneal administration of saline or LPS. RESULTS: There were no significant differences in the concentrations of IL-1beta and its receptors in the brain and peripheral tissues between sham-operated and ODX mice in both age groups injected with saline. LPS induced significantly higher IL-1beta production in the plasma and hippocampus in sham-operated 24-week-old mice than in 7-week-old mice. Coincident with the heightened IL-1beta response to LPS, hippocampal [(125)I]IL-1alpha binding was lower in 24-week-old mice than in 7-week-old mice after LPS injection in the sham-operated group. The age-related differences in the IL-1beta concentrations in the plasma and hippocampus and [(125)I]IL-1alpha binding in the hippocampus in response to LPS administration were abolished by ODX. Although LPS dramatically increased IL-1beta levels in the hypothalamus, no significant age-related differences in IL-1beta concentrations were seen, and ODX did not affect IL-1beta levels. In contrast, there were no significant differences between saline- and LPS-injected 7-week-old mice in relation to concentrations in the adrenal gland. Moreover, although the adrenal IL-1beta concentrations in 24-week-old mice were significantly higher than those in 7-week-old mice, ODX did not abolish these age-related differences in concentrations in the adrenal gland. CONCLUSIONS: Our data suggest the involvement of testosterone (or gonadal product) in plasma and hippocampal IL-1beta regulation in relation to age, and demonstrate the importance of the gonadal development in mediating the effects of infectious challenge on the brain and immune function.

Sarcoidosis with vocal fold involvement and hypercalcaemia.

Sarcoidosis affects many different organ systems. However, laryngeal involvement is rare and most cases with laryngeal involvement affect the supraglottis, occasionally secondarily extending to the vocal fold. The features of sarcoidosis in the supraglottic region have been described in several reports, but vocal fold involvement has not been presented in detail. We report an unusual case of laryngeal sarcoidosis initially involving the vocal folds in a 66-year-old man, associated with hypercalcaemia. Our report describes characteristics of the involved vocal folds. Hypercalcaemia is one of the characteristics of sarcoidosis. We concluded that laryngeal sarcoidosis should be added to the differential diagnosis of vocal fold lesions, particularly in patients with hypercalcaemia.

Glucocorticoid effects on the diurnal rhythm of circulating leptin levels.

It is known that circulating leptin shows diurnal variation with a nocturnal rise; however, the mechanisms generating this rhythm have not been fully elucidated. Glucocorticoids are a potent stimulator of leptin secretion, and there is a reciprocal relationship between circulating leptin and glucocorticoid levels. We hypothesized that glucocorticoids could modulate the diurnal rhythm of circulating leptin. We therefore explored the diurnal variation of leptin under situations in which subjects showed no or some shift of glucocorticoid diurnal rhythm, such as prednisolone-administered humans, and adrenalectomized and corticosterone-replaced (ADX+B) rats. The peak level of plasma cortisol immunoreactivity was shifted from early morning to noon by prednisolone administration. The nocturnal increment of plasma leptin in prednisolone-administered patients (71.2 +/- 14.2% from 08:00 h value) was significantly greater than that in normal volunteers (12.2 +/- 7.5% from 08:00 h value), but the timing of nadir and the peak of plasma leptin was not shifted. In normal rats, the plasma concentration of leptin showed the diurnal rhythm with the bottom at 16:00 h and the top between midnight and early morning. The amplitude of leptin diurnal rhythm was significantly reduced in ADX+B rats (08:00 h: 3.0 +/- 0.2, 16:00 h: 2.7 +/- 0.2, 00:00 h; 3.7 +/- 0.2 ng/ml) compared with sham operated rats (08:00 h: 3.0 +/- 0.2, 16:00 h 2.2 +/- 0.2, 00:00 h: 4.7 +/- 0.4 ng/ml); but ADX+B rats still retained similar timing of nadir and the peak of plasma leptin as observed in sham rats. These results indicate that glucocorticoids enhance the amplitude of leptin diurnal rhythm, and are consistent with previous findings showing that glucocorticoids increase leptin secretion. Glucocorticoids appear to play modulatory, but not essential roles in generating leptin diurnal rhythm.

Significant gene expression of insulin-like growth factor II and proliferating cell nuclear antigen in a rapidly growing recurrent pituitary acth-secreting adenoma.

BACKGROUND: We quantified the expression of various growth-related factors in an adrenocorticotropic hormone (ACTH)-secreting adenoma that had recurred very rapidly as invasive macroadenoma. METHODS/RESULTS: A 43-year-old woman underwent successful transsphenoidal surgery for Cushing's disease. Seven years later, she was admitted to our ward for further endocrine examinations. In spite of a very high plasma ACTH level, the serum cortisol level was normal. Discrepancies between ACTH and cortisol levels were detected on the basis of diurnal rhythms, dexamethasone suppression tests, and corticotropin-releasing hormone test. The patient showed no clinical features of Cushing's disease. Magnetic resonance imaging of the pituitary showed an almost empty sella, and no microadenoma was found. These results, along with those of Sephadex column gel filtration and high-performance liquid chromatography of plasma-immunoreactive ACTH, suggested that the patient's residual corticotrophs secreted biologically inactive ACTH. Two years later, the patient suddenly developed diplopia and right abducens nerve palsy. She was slightly moonfaced and centrally obese. Her plasma ACTH and serum and urinary free cortisol levels were elevated, although discrepancies between ACTH and cortisol still existed. Magnetic resonance imaging revealed a large pituitary mass with suprasellar and cavernous sinus extensions. The tumor was excised, and the proopiomelanocortin gene and the expression of growth-related factors were analyzed. No mutations were found in the ACTH-coding region of the proopiomelanocortin gene. A significant expression of insulin-like growth factor II and proliferating cell nuclear antigen mRNAs was demonstrated. A high MIB-1 antibody labeling index was also detected in the adenoma tissue, suggesting high Ki-67 expression. CONCLUSION: These growth- and proliferation-related factors might be involved in the rapid growth and aggressiveness of this patient's pituitary adenoma.

Exposure with the environmental estrogen bisphenol A disrupts the male reproductive tract in young mice.

Environmental estrogens (endocrine disruptive chemicals) have been shown to affect reproduction in wild life and it has been reported that maternal exposure with those chemicals have adverse effects on the male reproductive tract. However, little is known about the potential effects of prepubertal or pubertal exposure with environmental estrogens on the male reproductive tract. Here we examine plasma hormone levels and histology in the testis of mice following either 4- or 8-week oral administration of bisphenol A. Plasma free testosterone levels were dramatically decreased following 8 weeks of bisphenol A treatment compared with control group and morphologically multinucleated giant cells having greater than three nuclei were found in seminiferous tubules in the testis following the 8-week bisphenol A treatment. No differences in plasma corticosterone and luteinizing hormone levels were seen between bisphenol A and control groups. Thus, exposure with bisphenol A around pubertal period may directly disrupt the male reproductive tract. These facts suggest that more detailed studies will warrant the assessment of the risk to the developing human testis from exposure to bisphenol A and other environmental estrogens in prepubertal and pubertal period.

Decreased type 2 corticotropin-releasing hormone receptor mRNA expression in the ventromedial hypothalamus during repeated immobilization stress.

Chronic or repeated stress results in reduction of food intake and body weight in rats. Stress-induced anorexia has been attributed to increased corticotropin-releasing hormone (CRH) function in the central nervous system. To explore possible roles of other neuropeptides and peripheral hormones involved in food intake and energy utilization during continuing stress, we examined the impact of repeated immobilization stress on expression of mRNAs coding for CRH, neuropeptide Y (NPY), galanin and pro-opiomelanocortin (POMC) mRNAs in such hypothalamic nuclei as the paraventricular nucleus (PVN), arcuate nucleus (ARC) and dorsomedial hypothalamus (DMH), as well as plasma insulin and leptin concentrations. Changes in type 2 CRH receptor (CRHR-2) mRNA in the ventromedial hypothalamus (VMH), a possible target of anorectic CRH effect, were also examined. Rats were immobilized for 2 h daily for 6 days and sacrificed 24 h after the last immobilization. Immobilized rats had lower food intake and body weight and higher levels of PVN CRH mRNA than controls. Repeated immobiliza tion also lowered plasma insulin and leptin concentrations and VMH CRHR-2 mRNA levels. These results provide additional evidence linking VMH CRHR-2 mRNA levels to plasma leptin concentration. ARC NPY and DMH galanin mRNAs increased following repeated immobilization, while ARC POMC mRNA decreased. DMH NPY mRNA and ARC galanin mRNA were unaltered by immobilization. Since NPY and galanin are considered orexigenic, while the POMC-melanocortin-4 receptor system is apparently anorexigenic, the changes in neuropeptide mRNAs and VMH CRHR-2 mRNA may play counterregulatory roles against anorectic CRH effects.

Differential effects of one and repeated endotoxin treatment on pituitary- adrenocortical hormones in the mouse: role of interleukin-1 and tumor necrosis factor-alpha.

The role of endogenous interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) in modulating the hypothalamic-pituitary-adrenal (HPA) axis response was examined in male C57BL/6 mice injected with endotoxin (lipopolysaccharide, LPS) or saline at 24-hour intervals for 4 or 8 consecutive days. The mice were divided into four groups: (1) LPS injections for 4 or 8 days and LPS injection on day 5 or 9, respectively (LPS-LPS); (2) LPS injections for 4 or 8 days and saline injection on day 5 or 9, respectively (LPS-saline); (3) saline injections for 4 or 8 days and LPS injection on day 5 or 9, respectively (saline-LPS), and (4) saline injections for 4 or 8 days and saline injection on day 5 or 9 (saline-saline). The mice were sacrificed by decapitation 2 h after the last injection and plasma levels of hormones and cytokines and tissue levels of IL-1beta were measured. Plasma adrenocorticotropin (ACTH) levels were significantly attenuated in the LPS-LPS group compared with the dramatic increases in the saline-LPS group following 4 or 8 days of endotoxin treatment. Plasma corticosterone concentrations were comparable in the LPS-LPS group after 4 days' treatment, but significantly lower following 8 days of treatment when compared with saline-LPS group. Repeated endotoxin treatment followed by a single saline injection (LPS-saline) did not alter the levels of IL-1beta in plasma or any of the tissues examined. IL-1beta levels in the hippocampus, hypothalamus, adrenal gland and plasma were elevated to comparable levels in the saline-LPS and LPS-LPS groups after 4 days of treatment. In contrast to the plasma IL-1beta response, TNFalpha levels were dramatically increased in the saline-LPS group but not in the LPS-LPS group following the 4-day treatment regimen. Increases in IL-1beta concentrations were seen in all tissues following one endotoxin challenge in the saline-LPS group following the 8-day treatment regimen, while increases were significantly attenuated in the hypothalamus, adrenal gland and plasma in LPS-LPS for 8 days. The sustained increases in tissue levels of IL-1beta following 4-day endotoxin treatment appears to have functional consequences since [125I]IL-1alpha binding was significantly decreased in the LPS-saline group compared with the saline-saline group. Furthermore, [125I]IL-1alpha binding was markedly reduced in the LPS-LPS group compared with the saline-LPS group. There was a significant positive correlation between plasma ACTH and IL-1beta after a single and repeated LPS treatment for 4 days, while a significant correlation was seen between plasma ACTH and TNFalpha following one but not repeated LPS treatment. These data demonstrate a differential regulation of IL-1beta and TNFalpha by repeated endotoxin treatment and suggest that while TNFalpha may be important modulating the attenuated pituitary adrenocortical response following the 4-day endotoxin treatment, IL-1beta appears to be the primary regulator of the response following the 8-day endotoxin treatment in the regulation of the HPA axis.

Leptin effects on the expression of type-2 CRH receptor mRNA in the ventromedial hypothalamus in the rat.

The product of the ob gene, leptin, is thought to act in the hypothalamus to reduce food intake and body weight (b.w.) in rats and mice; however, the mechanisms of leptin action in the brain have not been fully elucidated. Corticotropin-releasing hormone (CRH) is a potent anorectic neuropeptide, and its type-2 receptor (CRHR-2) in the ventromedial hypothalamus (VMH) appears to play an important role in the expression of this anorectic effect. We explored here the impact of systemic leptin administration on CRH mRNA expression in the hypothalamic paraventricular nucleus (PVN) and CRHR-2 mRNA expression in the VMH in male rats, using in-situ hybridization histochemistry. The expression of CRH mRNA in the PVN and CRHR-2 mRNA in the VMH were increased at 2 h and 6 h, respectively, after a single intraperitoneal injection of leptin (1.0 mg/kg). Continuous subcutaneous infusion of leptin (1.2 mg/kg/day) via an osmotic minipump for 5 days increased the expression of CRHR-2 mRNA in the VMH, but not the expression of CRH mRNA in the PVN, compared with vehicle treatment. The rats that received the single or continuous administration of leptin showed reductions of food intake and b.w. compared with vehicle-treated rats. These results are consistent with our previous findings that the expression of CRHR-2 mRNA in the VMH is positively correlated with plasma leptin concentrations under various conditions, and highlight the importance of circulating leptin for the regulation of VMH CRHR-2 mRNA. The present results also raise the possibility that leptin reduces food intake and b.w. at least partially due to the enhancement of the anorectic effect of CRH via increased PVN CRH expression and/or VMH CRHR-2 expression.

Pituitary adenoma showing intermittent secretion of high molecular weight adrenocorticotropin without evidence of Cushing's disease.

A 29-year-old woman was admitted in March 1998 due to high plasma ACTH levels, amenorrhea and uncontrolled diabetes mellitus (DM) which had persisted since 1991. Plasma ACTH levels showed a wide range of changes: they were usually high (59-240 pg/ml), intermittently very high (336-942 pg/ml), and sometimes normal or low. Plasma cortisol levels were usually normal but were sometimes high when the ACTH levels were very high. However, even when the plasma ACTH levels were very high, she did not show any cushingoid features. DM was diagnosed as non-insulin-dependent DM. Plasma ACTH showed an excessive response to CRH, while cortisol showed a delayed response. Plasma cortisol showed a poor response to ACTH-(1-24). ACTH receptor gene analysis revealed no mutations in the ACTH receptor-coding region. MRI showed a nonenhancing mass on the left side of the pituitary. Cavernous sinus sampling showed a very high plasma ACTH level in the left cavernous sinus compared with the levels in the right cavernous sinus and peripheral blood. Sephadex G-75 gel filtration of plasma ACTH immunoreactivity in plasma obtained by cavernous sinus sampling showed mainly high molecular forms of ACTH, probably proopiomelanocortin and ACTH-beta-lipotropin. This case is a very rare form of pituitary adenoma showing intermittent secretion of high molecular ACTH unaccompanied by cushingoid features.