Different mechanisms for the inhibition of progesterone secretion by ACTH and corticosterone in pregnant rats.

The present study investigated possible sites through which ACTH or corticosterone inhibit progesterone secretion in pregnant rats, and the role of placental factors in blocking the inhibitory effect. The number of conceptuses was adjusted to one (IC group) or more than ten (FC group) on day 7 of pregnancy by aspirating the desired number. Serum concentrations of progesterone, testosterone and oestradiol were significantly (P less than 0.01) lower on day 15 in the 1C group than in the FC group. Corpora lutea (CL) obtained on day 15 were incubated for 6 h with corticosterone or ACTH. Corticosterone (1 mumol/l) significantly (P less than 0.05) inhibited progesterone secretion in the 1C group but not in the FC group. The inhibitory effect of corticosterone in the 1C group was completely blocked by co-addition of 1 mumol testosterone/l or 1 mumol oestradiol/l but not by 1 mumol dihydrotestosterone/l. ACTH (1 microgram/l-1 mg/l) had no direct effect on progesterone secretion in either the 1C or the FC groups, although ACTH apparently decreases progesterone secretion in vivo. Placentae obtained from rats of the FC group on day 15 were incubated for 24 h with or without ACTH (1 mg/l). The supernatant after placental incubation without ACTH significantly (P less than 0.01) increased progesterone secretion by the CL in both the 1C and FC groups, and also eliminated the inhibitory effect of corticosterone in the 1C group. The supernatant after placental incubation with ACTH also increased progesterone secretion in the FC group as effectively as the supernatant from the control incubation, but it had no effect in the 1C group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diuretics and bone loss in rats with aldosteronism

OBJECTIVES: We hypothesized that the increased urinary Ca2+ and Mg2+ excretion and bone loss that accompanies aldosteronism is aggravated with furosemide and is attenuated by spironolactone. BACKGROUND: Furosemide, a loop diuretic, is commonly used in patients with congestive heart failure (CHF), in which chronic, inappropriate (dietary Na+) elevations in plasma aldosterone (ALDO) and a catabolic state that includes bone wasting are expected. METHODS: In age- and gender-matched, untreated controls, four weeks of aldosterone/salt treatment (ALDO/salt, 0.75 microg/h + 1% NaCl/0.4% KCl in drinking water), four weeks of ALDO/salt + furosemide (40 mg/kg in prepared food), and four weeks of ALDO/salt + furosemide + spironolactone (200 mg/kg/day in divided doses by twice-daily gavage), we monitored: 24-h urinary Ca2+ and Mg2+ excretion; plasma-ionized [Ca2+]o and [Mg2+]o, K+, and parathyroid hormone (PTH); and bone mineral density (BMD) in the femur. RESULTS: The ALDO/salt increased (p < 0.05) urinary Ca2+ and Mg2+ excretion (4,969 +/- 1,078 and 3,856 +/- 440 microg/24 h, respectively) compared with controls (896 +/- 138 and 970 +/- 137 microg/24 h, respectively); furosemide co-treatment further increased (p < 0.05) urinary Ca2+ and Mg2+ excretion (6,976 +/- 648 and 6,199 +/- 759 microg/24 h, respectively), whereas spironolactone co-treatment attenuated (p < 0.05) these incremental losses (4,003 +/- 515 and 3,915 +/- 972 microg/24 h). Plasma [Ca2+]o was reduced (p < 0.05) at week 4 ALDO/salt + furosemide and was accompanied by hypokalemia (<3.4 mmol/l) that were rescued by spironolactone. Plasma PTH was increased (p < 0.05) compared with controls (30 +/- 4 vs. 11 +/- 3 pg/ml, respectively), whereas BMD was decreased (p < 0.05) with ALDO/salt and ALDO/salt + furosemide, but not with spironolactone co-treatment. CONCLUSIONS: In aldosteronism, hypercalciuria and hypermagnesuria and accompanying decrease in plasma-ionized [Ca2+]o and [Mg2+]o lead to hyperparathyroidism that accounts for bone wasting. Furosemide exaggerates these losses, whereas its combination with spironolactone attenuates these responses to prevent bone loss.     

Amylin inhibits ovariectomy-induced bone loss in rats

Amylin (AMY), a peptide co-secreted with insulin by pancreatic beta-cells, inhibits bone resorption and stimulates osteoblastic activity. The ovariectomized (OVX) rat is an established animal model for human osteoporosis. Thus, the present experiment was performed to study the effects of AMY on estrogen deficiency-induced bone loss in rats. Thirty-one 6-month-old Wistar rats were randomized by body weight (BW) into two groups. The first underwent surgical OVX (n=21). The second was sham-operated (SH; n=10). Sixty days after surgery, 11 OVX rats were s.c. injected with rat AMY (3 microg/100 g BW/day, for 30 days; OVX+AMY), and 10 with solvent alone in the same way (0.15 ml/100 g BW; OVX). Each rat, housed in an individual cage, was fed daily the mean quantity of diet consumed the day before by SH rats. This diet contained 0.24% calcium and 0. 16% phosphorus. The 31 animals were killed on day 90. No difference in daily weight gain and BW was observed between groups. Neither AMY treatment nor OVX had any significant effect upon femoral morphology, femoral failure load, diaphyseal femoral density (representative of cortical bone) and total femoral calcium content. Nevertheless, both distal metaphyseal (representative of cancellous bone) and total femoral bone densities were higher in SH and OVX+AMY than in OVX rats. The highest plasma osteocalcin concentration was measured in OVX+AMY rats. Simultaneously, urinary deoxypyridinoline excretion was lower in OVX+AMY than in OVX rats. These results indicate that in OVX rats, AMY treatment inhibited trabecular bone loss both by inhibiting resorption and by stimulating osteoblastic activity.     

Leptin reduces ovariectomy-induced bone loss in rats.

Bone mineral density increases with fat body mass, and obesity has a protective effect against osteoporosis. However, the relationship between fat body mass and bone mineral density is only partially explained by a combination of hormonal and mechanical factors. Serum leptin levels are strongly and directly related to fat body mass. We report here the effects of leptin administration compared with estrogen therapy on ovariectomy-induced bone loss in rats. Leptin was effective at reducing trabecular bone loss, trabecular architectural changes, and periosteal bone formation. Interestingly, the combination of estrogen and leptin further decreased bone turnover compared with that in estrogen-treated ovariectomized rats. Leptin also significantly increased osteoprotegerin mRNA steady state levels and protein secretion and decreased RANK ligand mRNA levels in human marrow stromal cells in vitro. Our findings suggest that leptin could modulate bone remodeling in favor of a better bone balance in rats. This study is the first evidence that leptin therapy has a significant effect in preventing ovariectomy-induced bone loss, and this effect may at least in part be mediated by the osteoprotegerin/RANK ligand pathway.     

内质网分子伴侣糖调节蛋白94在大鼠酒精性肝损伤中的高表达和意义

目的检测酒精性肝损伤大鼠肝组织内质网分子伴侣糖调节蛋白94的表达情况,探讨酒精性肝损伤的发病机制.方法24只SD雌性大鼠分成对照组和模型组.模型组大鼠给予乙醇加鱼油灌胃配合高脂饮食8周,建立酒精性肝损伤模型,应用半定量逆转录聚合酶链反应(RT-PCR)法和免疫组化法检测大鼠肝组织内质网分子伴侣糖调节蛋白94 mRNA和蛋白水平.结果与正常组比较,模型组大鼠肝组织糖调节蛋白94 mRNA和蛋白表达明显增强(P＜0.01).结论酒精性肝损伤大鼠存在糖调节蛋白94高表达,这可能是该病的发病机制之一.     

Bariatric surgery,bone loss,obesity and possible mechanisms

Bariatric surgery remains the most effective treatment for severely obese patients. However, the potential long‐term effects of bariatric surgical procedures on health, including bone health, are only partially understood. The goal of this review was to present data on the impact of bariatric surgery on bone metabolism and to analyse possible reasons for the loss of bone mass that frequently occurs after bariatric surgery. Such factors include nutritional deficiencies, rapid weight loss per se, effects of fat‐derived adipokines and gut‐derived appetite‐regulatory hormones. However, the relative roles of these factors in skeletal regulation and the mechanisms by which they work are not yet fully defined. Our review was focussed on the complex relationship between body weight, fat mass and bone mass, as well as peripheral and central mediators potentially involved in the dual regulation of both energy and bone homeostasis. We also review the data on the inverse relationship between central obesity, bone marrow fat and osteoporosis. As the number of bariatric operations increases, it is imperative to recognize mechanisms responsible for bariatric surgery‐induced bone loss, with careful monitoring of bone health including long‐term fracture incidence in patients undergoing these procedures.     

Ghrelin protects against ethanol-induced gastric ulcers in rats: studies on the mechanisms of action

Ghrelin, the endogenous ligand for GH secretagogue receptors, has been reported to influence acid gastric secretion and motility, but its potential gastroprotective effect is unknown. The aims of this study were 1) to examine the effects of central and peripheral administration of ghrelin on ethanol-induced gastric ulcers in conscious rats, and 2) to investigate the possible roles of nitric oxide (NO), vagal nerve, and sensory fibers in the gastric effects of ghrelin. Ghrelin was administered either intracerebroventricularly or sc 30 min before ethanol, and mucosal lesions were examined macroscopically. Additionally, rats were either treated with the inhibitor of NO synthesis N(omega)-nitro-L-arginine methyl ester (L-NAME) or underwent bilateral cervical vagotomy or capsaicin-induced sensory denervation. Conventional histology and immunohistochemistry for ghrelin, gastrin, and somatostatin were performed on gastric specimens from representative rats. Central ghrelin (4-4,000 ng/rat) dose-dependently reduced ethanol-induced gastric ulcers by 39-77%. Subcutaneous ghrelin administration (80 micro g/kg) reduced ulcer depth only. L-NAME and capsaicin, but not vagotomy, prevented the gastroprotective effect of central ghrelin (4000 ng/rat). This is the first evidence that ghrelin exerts a potent central gastroprotective activity against ethanol-induced lesions. The gastroprotective effect of ghrelin is mediated by endogenous NO release and requires the integrity of sensory nerve fibers.     

过氧化物酶体增殖物激活受体γ2活化与大鼠增龄骨丢失关系的研究

目的 观察自然增龄大鼠骨髓过氧化物酶体增殖物激活受体γ2（PPARγ2）表达与氧化低密度脂蛋白（Ox-LDL）及骨量的关系，探讨其在大鼠自然增龄骨丢失发生过程中可能的作用机制。方法 选择不同月龄（6、12及18月龄）大鼠作为研究对象，逆转录PCR（RT—PCR）检测骨髓PPARγ2、核因子-κB活化受体配体（RNAKL）、骨保护素（OPG）、核因子-κB活化受体（RANK）mRNA表达水平；ELISA法检测骨髓上清Ox-LDL、脂肪酸结合蛋白（aP2）水平，图像分析仪测定骨髓脂肪组织含量并检测股骨骨密度。结果 骨密度随增龄进行性减低，组间比较差异有统计学意义（P〈0．05）；骨组织HE染色，6月龄大鼠骨髓腔中脂肪极少，而在18月龄约占40％骨髓腔空间，组间比较差异有统计学意义（P〈0．01），脂肪髓的发生与年龄呈正相关（r=0．762），与骨密度呈负相关（r=0．626）；骨髓中aP2及Ox-LDL浓度随增龄明显增高，组间比较差异有统计学意义（P〈0．01）；PPARγ2表达随增龄明显升高，3组差异有统计学意义（P〈0．05或P〈O．01）；RANKL表达随增龄呈进行性升高，而OPG在18月龄组明显下降，组间比较差异有统计学意义（P〈0．01或P〈0．05），RANK随增龄无明显变化。结论 随增龄Ox-LDL水平升高可激活骨髓PPARγ2转录活性，引起骨髓脂肪生成增多并参与骨量减低的发生，是Ⅱ型原发性骨质疏松发生的可能机制之一。     

Two distinct mechanisms underlie progesterone-induced proliferation in the mammary gland

The mouse mammary gland develops postnatally under the control of female reproductive hormones. Estrogens and progesterone trigger morphogenesis by poorly understood mechanisms acting on a subset of mammary epithelial cells (MECs) that express their cognate receptors, estrogen receptor α (ERα) and progesterone receptor (PR). Here, we show that in the adult female, progesterone drives proliferation of MECs in two waves. The first, small wave, encompasses PR(+) cells and requires cyclin D1, the second, large wave, comprises mostly PR(−) cells and relies on the tumor necrosis factor (TNF) family member, receptor activator of NF-κB-ligand (RANKL). RANKL elicits proliferation by a paracrine mechanism. Ablation of RANKL in the mammary epithelium blocks progesterone-induced morphogenesis, and ectopic expression of RANKL in MECs completely rescues the PR^−/− phenotype. Systemic administration of RANKL triggers proliferation in the absence of PR signaling, and injection of a RANK signaling inhibitor interferes with progesterone-induced proliferation. Thus, progesterone elicits proliferation by a cell-intrinsic and a, more important, paracrine mechanism.     

The mechanisms that underlie glucose sensing during hypoglycaemia in diabetes

Hypoglycaemia is a frequent and greatly feared side-effect of insulin therapy, and a major obstacle to achieving near-normal glucose control. This review will focus on the more recent developments in our understanding of the mechanisms that underlie the sensing of hypoglycaemia in both non-diabetic and diabetic individuals, and how this mechanism becomes impaired over time. The research focus of my own laboratory and many others is directed by three principal questions. Where does the body sense a falling glucose? How does the body detect a falling glucose? And why does this mechanism fail in Type 1 diabetes? Hypoglycaemia is sensed by specialized neurons found in the brain and periphery, and of these the ventromedial hypothalamus appears to play a major role. Neurons that react to fluctuations in glucose use mechanisms very similar to those that operate in pancreatic B- and A-cells, in particular in their use of glucokinase and the K_ATP channel as key steps through which the metabolic signal is translated into altered neuronal firing rates. During hypoglycaemia, glucose-inhibited (GI) neurons may be regulated by the activity of AMP-activated protein kinase. This sensing mechanism is disturbed by recurrent hypoglycaemia, such that counter-regulatory defence responses are triggered at a lower glucose level. Why this should occur is not yet known, but it may involve increased metabolism or fuel delivery to glucose-sensing neurons or alterations in the mechanisms that regulate the stress response.     

Efficiency of bone marrow-derived cells in regeneration of the stomach after induction of ethanol-induced ulcers in rats

Background Bone marrow contains hematopoietic stem cells, nonhematopoietic mesenchymal stem cells, and several precursor cells for osteoblasts, chondrocytes, adipocytes, myocytes, hepatocytes, and even neural cells. Research findings indicate that multipotent stem cells in the adult body may be used to recover the lost functions of damaged tissues. This study examined the involvement of bone marrow-derived cells in the regeneration of the stomach after experimental gastric ulcers were produced in rats.Methods We transplanted the bone marrow of transgenic rats that expressed green fluorescence protein (GFP) throughout the body. Twenty-one days after the bone marrow transplantation (BMT), gastric ulceration was induced, using absolute ethanol. Control animals received saline. After various observation periods, rats harboring GFP-positive bone marrow-derived cells were killed, and the tissues were removed and processed to prepare paraffin-embedded sections. Cells expressing GFP were identified by conventional immunohistochemistry, using anti-GFP antibody. To identify whether cells expressing GFP were epithelial cells or interstitial cells such as fibroblasts, serial sections were examined with anti-cytokeratin antibody or anti-vimentin antibody, respectively. Furthermore, to confirm that cells expressing GFP were epithelial cells or interstitial cells, we used double-staining analysis with anti-GFP antibody or anti-cytokeratin antibody, respectively.Results GFP-positive, bone marrow-derived cells were found in the cytokeratin-positive gastrointestinal epithelium, as well as among vimentin-positive interstitial cells. Interestingly, the proportions of GFP-positive, cytokeratin-positive epithelial cells and vimentin-positive interstitial cells were significantly greater in the ethanol-treated damaged stomachs than in the saline-treated controls.Conclusions The present study clearly demonstrates that bone marrow-derived cells are involved in the regeneration of the stomach after ethanol-induced ulcers in rats.     

Platelet-free calcium and vascular calcium uptake in ethanol-induced hypertensive rats

This study examined the effect of moderate ethanol intake on systolic blood pressure, platelet cytosolic free calcium, aortic calcium, and rubidium-86 uptake in Wistar-Kyoto rats. Twelve Wistar-Kyoto rats, aged 6 weeks, were given 5% ethanol in drinking water the first week followed by 10% ethanol in drinking water for the next 6 weeks. Twelve control animals were given regular tap water. Systolic blood pressure in the ethanol-treated rats was significantly higher (p less than 0.05) than that in controls after 1 week and remained higher throughout the study. At 13 weeks of age, platelet cytosolic free calcium and calcium uptake by aortas were significantly higher (p less than 0.001) in ethanol-treated animals as compared with those in controls. Ethanol intake did not affect aortic ouabain-sensitive 86Rb uptake. The in vitro effect of ethanol on calcium-45 and 86Rb uptake was also investigated in aortas of untreated Wistar-Kyoto rats at 13 weeks of age. In vitro ethanol (2.5-20 mmols/l) did not significantly affect 45Ca and 86Rb uptake in rat aortas. The increases in systolic blood pressure, platelet cytosolic free calcium, and vascular calcium uptake suggest that increases in cytosolic free calcium and calcium uptake mechanisms are associated with ethanol-induced hypertension.     

Verapamil decreases ethanol-induced gastric acid secretion in rats

The present study examined the effect of verapamil, a calcium channel blocker, on gastric acid secretion and circulating gastrin levels in rats after ethanol challenge. Normal saline or verapamil were given intraperitoneally to different groups of rats at 1 min, 1 h or 2 h before the administration of ethanol. One hour later, gastrin and gastric acid concentrations were determined. Regression analysis revealed the relationship between gastric acid output and serum gastrin levels in the group receiving saline intraperitoneally and ethanol orogastrically and the group receiving saline both intraperitoneally and orogastrically is similar. The slope of the regression line of the ethanol-challenged group treated with verapamil, however, was significantly lower than the slopes of the other two groups (P less than 0.001). Furthermore, verapamil decreased gastrin levels and acid output significantly in the ethanol-challenged group (P less than 0.01). When given 10 min prior to ethanol challenge, verapamil had a greater acid suppression effect than when given 60 or 120 min before the challenge. Verapamil at 20 mg/kg was more effective in acid secretion than at 10 mg/kg bodyweight, when administered 60 min before ethanol challenge.     

Ketotifen and nitroxides decrease capsaicin-augmented ethanol-induced gastric damage in rats

Systemic administration of capsaicin aggravates ethanol-induced injury of rat gastric mucosa. We evaluated the effect of subcutaneous administration of capsaicin on the gastric mucosa and on inflammatory mediators in saline- and ethanol-treated rats. Functional ablation of primary afferent C-fibers by capsaicin (total 100 mg/kg subcutaneous) tripled ethanol-induced damage. Pretreatment with ketotifen, a mast cell stabilizer (1 mg/kg) protected rat gastric mucosa from the amplified injury induced by capsaicin and ethanol. Tempol, a selective nontoxic cell-permeable nitroxide, completely prevented the amplified gastric ulceration induced by capsaicin and ethanol. This was accompanied by a significant decrease in leukotriene B₄ and C₄ generation. It is therefore suggested that mast cells and free radicals contribute to the amplified injury observed in rats pretreated with capsaicin and ethanol and that the pharmacological modulation of mast cell release and scavenging of free radicals may be of therapeutic efficacy in the prevention of gastric injury.