Ob-Rb Expression Research Articles

Low expression level of OB-Rb results from constitutive translocational attenuation attributable to a less efficient signal sequence.

OB-Rb is a crucial factor for leptin signaling. This study was initially motivated by the observation that OB-Rb expression is constitutively inhibited in the early secretory pathway. Our analyses reveal that OB-Rb contains a less hydrophobic, but functionally active N-terminal signal sequence. Constitutive translocational attenuation attributable to a less efficient signal sequence proved to be a reason for low protein level of OB-Rb. By contrast, enhanced signal sequence efficiency rescues translocation and cell surface expression of OB-Rb, and eventually potentiates leptin signaling. These observations provide considerable insight into the therapeutic enhancement of OB-Rb translocation as a potential strategy for leptin resistance.

GW4064, a farnesoid X receptor agonist, upregulates adipokine expression in preadipocytes and HepG2 cells.

To investigate the effect of GW4064 on the expression of adipokines and their receptors during differentiation of 3T3-L1 preadipocytes and in HepG2 cells. The mRNA expression of farnesoid X receptor (FXR), peroxisome proliferator-activated receptor-gamma 2 (PPAR-γ2), adiponectin, leptin, resistin, adiponectin receptor 1 (AdipoR1), adiponectin receptor 2 (AdipoR2), and the long isoform of leptin receptor (OB-Rb) and protein levels of adiponectin, leptin, and resistin were determined using fluorescent real-time PCR and enzyme linked immunosorbent assay, respectively, on days 0, 2, 4, 6, and 8 during the differentiation of 3T3-L1 preadipocytes exposed to GW4064. Moreover, mRNA expression of AdipoR2 and OB-Rb was also examined using fluorescent real-time PCR at 0, 12, 24, and 48 h in HepG2 cells treated with GW4064. The mRNA expression of FXR, PPAR-γ2, adiponectin, leptin, resistin, AdipoR1, AdipoR2, and OB-Rb and protein levels of adiponectin, leptin, and resistin increased along with differentiation of 3T3-L1 preadipocytes (P < 0.05 for all). The mRNA expression of FXR, PPAR-γ2, adiponectin, leptin, and AdipoR2 in 3T3-L1 preadipocytes, and AdipoR2 and OB-Rb in HepG2 cells was significantly increased after treatment with GW4064, when compared with the control group (P < 0.05 for all). A similar trend was observed for protein levels of adipokines (including adiponectin, leptin and resistin). However, the expression of resistin, AdipoR1, and OB-Rb in 3T3-L1 cells did not change after treatment with GW4064. The FXR agonist through regulating, at least partially, the expression of adipokines and their receptors could offer an innovative way for counteracting the progress of metabolic diseases such as nonalcoholic fatty liver disease.

The leptin receptor is expressed in the Leydig cells of Merino rams, but its expression is not affected by long-term differences in the plane of nutrition

We studied whether the long-term effects of nutrition on ram reproductive potential are mediated by changes in the expression and distribution of leptin receptor (Ob-Rb) in the testes. Thirty-two young Merino rams were allocated to a 2 × 2 factorial design, with either natural (NP) or improved pastures (IP), with (+S) or without (–S) supplementation. After 14 weeks of treatment, a blood sample was taken to measure leptin concentrations and rams were castrated. The weights of testicles (TW) and epididymides (EW) were recorded before tissue sampling for the detection of Ob-Rb expression by immunohistochemistry. TW was positively affected by the interaction IP × S, but not EW and bodyweight (BW), which were affected either by IP or S (P &lt; 0.05). TW and EW were greatest (g) in IP+S rams (290.6 ± 40.1 and 57.6 ± 2.3), followed by IP–S (284.6 ± 61.4 and 52.5 ± 2.3), NP+S (282.6 ± 37.9 and 55.8 ± 2.3), and NP–S rams (208.5 ± 48.7 and 44.4 ± 2.3; P &lt; 0.01). Expression of Ob-Rb was observed in the cytoplasm of Leydig cells, without any effect of treatments on the percentage of positive Leydig cells or on staining-intensity score. The serum leptin concentrations were similar among groups. The absolute volume of the seminiferous epithelium (mL) was affected by IP and S, as well as by their interaction (IP–S: 179.7 ± 13.1, IP+S; 174.9 ± 9.0; NP+S: 178.0 ± 10.2; NP–S: 120.8 ± 9.8; P &lt; 0.05). We conclude that, under the conditions of the present study, the long-term improvement of nutrition promoted histological changes of increased spermatogenesis of Merino rams, but was not related to changes in the expression of the Ob-Rb receptor in the Leydig cell cytoplasm.

Plumbagin Inhibits Leptin-Induced Proliferation of Hepatic Stellate Cells via JAK2-STAT3 Pathway to Protect against Hepatic Fibrosis

Purpose: To investigate the protective effects of plumbagin against liver fibrosis and explore the influence of plumbagin on the proliferation of hepatic stellate cells (HSCs).Methods: HSC-LX2 cells were divided into blank/control group, 100 ng/ml leptin group, 100 ng/ml leptin + 2 μmol/L plumbagin group, 100 ng/ml leptin + 8 μmol/L plumbagin group and 100 ng/ml leptin + 6.25 μg/ml colchicines group. The expressions of leptin receptor protein (OB-Rb), p-JAK2, p-STAT3, JAK2, STAT3, p-ERK1/2, ERK1/2 and MMP-1 were detected by Western blot assay. The content of type I collagen in the supernatant of HSCs was also measured after stimulation by leptin.Results: Leptin induced OB-Rb expression in HSCs, which reached a peak level at 24 h (p &lt; 0.01). Leptininduced OB-Rb expression was significantly inhibited by plumbagin at concentrations of 2 μmol/L and 8 μmol/L, respectively. Western blot assay revealed that plumbagin significantly decreased pJAK2 expression in leptin-treated HSCs (p &lt; 0.01). Leptin induced expression of pSTAT3 significantly decreased after plumbagin treatment in HSC-LX2 (p &lt; 0.01). p-ERK1/2 expression markedly decreased in plumbagin-treated HSCs (p &lt; 0.01). Plumbagin significantly increased MMP-1 expression in leptin-treated HSCs (p &lt; 0.01).Conclusion: Plumbagin has an anti-fibrotic effect and may decrease the protein expressions of components in JAK2-STAT3 pathway to inhibit HSC proliferation. Thus, plumbagin may be useful in the clinical prevention and treatment of liver fibrosis.Keywords：Plumbagin, Leptin, Hepatic stellate cell, Proliferation, Signal transduction pathway, Antiinflammatory

Exercise training improve leptin sensitivity in peripheral tissue of obese rats

The present study examined thechange to the effect of the leptin sensitivity by leptin resistance-induced leptin receptor (ObRb) and leptin-related suppressor of cytokine signaling 3 (SOCS3) mRNA levels in hypothalamic, liver, muscle and leptin protein levels in blood after eight 8weeks of exercise training and/or dietary control of high fat induced obese rats. After 2weeks of adaptation maintenance, four-week-old male SD rats (n=42) were randomly divided into control (CO) (n=8) and high-fat diet (HF) (n=32) groups. The HF group randomly divided into HF, HF+exercise training (HFT), changed to normal diet (HFND) and changed to normal diet and exercise training (HFNDT) groups. 13weeks of HF group average body weight significantly increased in comparison to the CO group (p<0.05). Plasma leptin levels of the HFT, HFND and HFNDT group were significantly decreased in comparison to the HF group (p<0.05). The mRNA expression of ObRb and SOCS3 in the liver and muscle of the HF group was significantly decreased comparison to that of the HFT, HFND and HFNDT group after 8weeks intervention (p<0.05). In addition, the mRNA expression of ObRb and SOCS3 in the hypothalamus of the HF group was significantly increased comparison to that of the HFT, HFND and HFNDT group (p<0.05). HFND group also was significantly reduced comparison to of the HFT and HFNDT group (p<0.05).These findings suggest that the effect of leptin sensitivity in peripheral may primarily the more relate to combined dietary control and exercise training more than effect of dietary control.

Increased Leptin Response and Inhibition of Apoptosis in Thymocytes of Young Rats Offspring from Protein Deprived Dams during Lactation

We investigated the consequences of mild maternal malnutrition in rat dams, in terms of thymocyte responses and the putative role of leptin. The young progeny of dams submitted to protein deprivation (PD) during lactation showed at 30 days of age lower body and thymus weights, significant alterations in CD4/CD8-defined T cell subsets without modifications in total thymocyte number as well as in proliferative response. Despite, the rats from PD group did not present alterations in leptin circulating levels, the expression of leptin receptor ObRb was enhanced in their thymocytes. This change was accompanied by an increase in leptin signaling response of thymocytes from PD rats, with an increase in JAK2 and STAT3 phosphorylation after leptin stimulation. Thymocytes from PD rats also presented a decreased rate of spontaneous apoptosis when compared to controls. Accordingly, higher expression of anti-apoptotic protein Bcl-2, and lower of pro-apoptotic protein Bax, with no change of pro-apoptotic Bad, and higher pro-caspase 3 content were detected in PD thymocytes. Moreover, thymocytes from PD group exhibited a constitutive higher nuclear content of p65 NF-kB associated to a lower IkB content in the cytoplasm. Finally, although there was no change in ob gene expression in PD thymocytes, a higher mRNA expression for the Ob gene was observed in the thymic microenvironment from PD animals. Taken together, the results show that mild maternal protein deprivation during lactation affects thymic homeostasis, enhancing leptin activity, which in turn protects thymocytes from apoptosis in the young progeny, with possible consequences upon the immune response of these animals in adult life.

Up-Regulation of the Fetal Baboon Hypothalamo-Pituitary-Adrenal Axis in Intrauterine Growth Restriction: Coincidence with Hypothalamic Glucocorticoid Receptor Insensitivity and Leptin Receptor Down-Regulation

Intrauterine growth restriction (IUGR) is an important fetal developmental problem resulting from 2 broad causes: maternal undernutrition and/or decreased fetal nutrient delivery to the fetus via placental insufficiency. IUGR is often accompanied by up-regulation of the hypothalamo-pituitary-adrenal axis (HPAA). Sheep studies show fetal HPAA autonomy in late gestation. We hypothesized that IUGR, resulting from poor fetal nutrient delivery, up-regulates the fetal baboon HPAA in late gestation, driven by hypothalamo-pituitary glucocorticoid receptor (GR) insensitivity and decreased fetal leptin in peripheral plasma. Maternal baboons were fed as ad libitum controls or nutrient restricted to produce IUGR (fed 70% of the control diet) from 0.16 to 0.9 gestation. Peripheral ACTH, cortisol, and leptin were measured by immunoassays. CRH, arginine vasopressin (AVP), GR, leptin receptor (ObRb), and pro-opiomelanocortin peptide expression were determined immunohistochemically. IUGR fetal peripheral cortisol and ACTH, but not leptin, were increased (P < .05). IUGR increased CRH peptide expression, but not AVP, in the fetal hypothalamic paraventricular nucleus (PVN) and median eminence (P < .05). PVN ObRb peptide expression, but not GR, was decreased (P < .05) with IUGR. ObRb and pro-opiomelanocortin were robustly expressed in the anterior pituitary gland, but ∼1% of cells showed colocalization. We conclude that (1) CRH, not AVP, is the major releasing hormone driving ACTH and cortisol secretion during primate IUGR, (2) fetal HPAA activation was aided by GR insensitivity and decreased ObRb expression in the PVN, and (3) the anterior pituitary is not a site for ObRb effects on the HPAA.

Sweet Preference Modified by Early Experience in Mice and the Related Molecular Modulations on the Peripheral Pathway

The sweet taste is of immense interest to scientists and has been intensively studied during the last two decades. However, the sweet preference modification and the related mechanisms are still unclear. In this study, we try to establish a mice model with manipulated sweet taste preference and explore the involved possible molecular mechanisms. The animals were exposed to acesulfame-K via maternal milk during lactation and the sweet preference tests were carried out when they grew to adulthood. Our results showed that the preference thresholds for sweet taste were increased in adults by early acesulfame-K exposure and the preference ratios for sweet tastants at low or preferred concentrations were decreased. Moreover, by means of qRT-PCR and Western blot, we observed the increased expression of leptin receptor Ob-Rb and downregulation of Gα-gustducin protein in the soft palate. Thereby, the sweet taste sensitivity may be modified by early sweetener experience during lactation. Along the peripheral sweet sensory pathway, the sweet regulator receptors Ob-Rb, CB1 and components of sweet transduction signal Gα-gustducin and T1R2 in both the soft palate and tongue may be cooperatively involved in the plastic development of sweet taste.

The role of leptin on the organization and expression of cytoskeleton elements in nucleus pulposus cells

Obesity is an important risk factor for intervertebral disc degeneration and leptin is a biomarker of obesity. However, the expression of leptin receptors has not been determined in disc tissue. It is not known whether leptin has a direct effect on the nucleus pulposus (NP) cells. To determine whether the NP tissues and cells express leptin receptors (OBRa and OBRb) and whether leptin affects the organization and the expression of major cytoskeletal elements in NP cells. Messenger RNA (mRNA) and protein levels of OBRa and OBRb were measured by real-time PCR and Western blot, respectively, in NP tissues and cells. Immunofluorescence and real-time PCR and Western blot were performed to investigate the effect of leptin on cytoskeleton reorganization and expression. Results show that mRNA and proteins of OBRa and OBRb were expressed in all NP tissues and cells, and that OBRb expression was correlated with patients' body weight. Increased expression of β-actin and reorganization of F-actin were evident in leptin-stimulated NP cells. Leptin also induced vimentin expression but had no effect on β-tubulin in NP cells. These findings provide novel evidence supporting the possible involvement of leptin in the pathogenesis of intervertebral disc degeneration. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 847–857, 2013

Protective Role of Astrocytic Leptin Signaling Against Excitotoxicity

Both proconvulsive and anticonvulsive roles of leptin have been reported, suggesting cell-specific actions of leptin in different models of seizure and epilepsy. The goal of our study was to determine the regulation and function of astrocytic leptin receptors in a mouse model of epilepsy and glutamate-induced cytotoxicity. We show that in pilocarpine-challenged mice developing epilepsy with recurrent seizures after a latent period of 2 weeks, hippocampal leptin receptor (ObR) immunofluorescence was increased at 6 weeks. This was more pronounced in astrocytes than in neurons. In cultured astrocytes, glutamate increased ObRa and ObRb expression, whereas leptin pretreatment attenuated glial cytotoxicity by excess glutamate, reflected by better preserved adenosine triphosphate production. The protective role of astrocytic leptin signaling is further supported by the higher lethality of the astrocyte-specific leptin receptor knockout mice in the initial phase of seizure production. Thus, leptin signaling in astrocytes plays a protective role against seizure, and the effects are at least partially mediated by attenuation of glutamate toxicity. Astrocytic leptin signaling, therefore, may be a novel therapeutic target.

Leptin attenuates the anti-estrogen effect of tamoxifen in breast cancer

BackgroundLeptin is a circulating peptide hormone, encoded by the obesity (ob) gene, acting as a regulator of food intake via hypothalamic-mediated effects. Recent studies have shown that leptin and leptin receptor (ObR) are involved in the carcinogenesis and development of breast cancer. In addition, functional cross talk between leptin and estrogen signaling has been registered. Here, we investigated the relation of leptin and ObR expression with survival in women with breast cancer treated with the anti-estrogen tamoxifen, and whether leptin can interfere with the estrogen receptor alpha (ERα) and the effect of tamoxifen in breast cancer cells. MethodsThe protein expression of leptin and ObRb in 114 breast cancer samples was evaluated by immunohistochemistry, quantified by Immunoreactivity Score (IRS) and correlated to survival and other clinicopathological features. The expression of ObRb in ERα positive MCF-7 breast cancer cells was examined by immunofluorescence and western blot. Leptin effect on cell proliferation was determined by MTT assay. The interference of leptin with tamoxifen on ERα degradation was studied by western blot and immunofluorescence. Effects of leptin on the transcriptional activity of ERα were explored using luciferase reporter assays. ResultsPositive staining (Immunoreactivity Score, IRS≥1) of leptin and ObR isoform ObRb in breast cancer tissues were seen in 79.8% and 85.1% of patients respectively. In overall and in tamoxifen-treated breast cancer patients, leptin expression (IRS≥1) correlated with poor prognosis, (log-rank test, P=0.016, overall; P=0.031, tamoxifen-treated). Overexpressed ObRb was found by western blotting andimmunofluorescence in MCF-7 as well as in MDA-MB-231, T47D, and MDA-MB-435 cell lines. Tamoxifen (1000μM) significantly inhibited the proliferation of MCF-7 cells, degraded ERα and reduced ERα-dependent transcription from estrogen response element-containing promoter. On the contrary, simultaneous treatment with leptin (100ng/ml) significantly attenuated these effects, similar to the effects of estradiol. ConclusionsLeptin correlated significantly with poor prognosis in overall and tamoxifen-treated breast cancer patients. Leptin interferes with the action of tamoxifen in MCF-7 cells, at least partly, through inducing increased nuclear expression of ERα. Thus, leptin may contribute to tamoxifen resistance and consequently, leptin suppression could be a novel way of circumventing resistance to anti-estrogen treatment.

Estrogen and Leptin Regulation of Endocrinological Features of Anorexia Nervosa

Anorexia nervosa (AN) is an eating disorder characterized by profound weight loss, osteoporosis, amenorrhea, and low leptin levels. Leptin is a hormone secreted from adipocytes in direct proportion to body adiposity and low levels of leptin, or hypoleptinemia, is a key endocrinological feature of AN. AN is more prevalent in women, and is characterized by severely reduced estradiol levels. Estrogens are critical regulators of reproduction and metabolism that act by binding to estrogen receptors (ERs). ERs are members of the nuclear receptor superfamily and they regulate target genes by binding to estrogen response elements (EREs). There is an ERE on the long form of the leptin receptor (OB-Rb), which allows estrogen to modulate OB-Rb expression (Machinal et al, 1999). This review will focus on the possible therapeutic benefits of combining estrogens and leptin therapy for AN patients. We propose that combination therapy would decrease the effective dose of each drug and this in turn would reduce the anorexic side effects reported at higher doses. ESR1 and ESR2 are genes that code for estrogen receptors ERα and ERβ, respectively (Osterlund and Hurd, 2001), and are highly expressed in hypothalamic regions that regulate body weight and reproduction. ERs and OB-Rbs are colocalized in specific hypothalamic neurons and as estrogen levels increase, leptin transport across the blood–brain barrier is enhanced. In addition, knockdown of ERα from hypothalamic steroidogenic factor-1 and pro-opiomelanocortin neurons increases food intake, or reduces energy expenditure, or influences reproduction (Xu et al, 2011). In healthy females, leptin regulates the minute-to-minute oscillations in the levels of luteinizing hormone (LH) and estradiol, the most potent estrogen (Licinio et al, 1998). In women with AN, leptin administration increases pulsatile LH, resulting in an enlargement of the ovaries, increased number and sizes of follicles, and elevated plasma estradiol levels (Welt et al, 2004). Therefore, leptin may be necessary for the resolution of amenorrhea in AN. In addition, leptin through OB-Rb reduces food intake by activating transcription factors such as phosphorylated STAT3 (pSTAT3) (Munzberg et al, 2005). Estrogens potentiate leptin-induced pSTAT3 activation in the hypothalamus. Leptin is one of the few treatments for AN patients (Welt et al, 2004); however, at high doses leptin increases weight loss. There are no standardized medications approved for the treatment of AN by the Food and Drug Administration; however, research has focused on the potential therapeutic actions of leptin, as leptin may promote restoration of menstrual cycles and prevent osteoporosis. Estrogen is also given as a treatment for AN; however, sudden introduction of sex steroids to hypo-estrogenic girls can be followed by reductions in food intake and heightened manifestations of AN (Kauli et al, 1982). Therefore, our hypothesis is that combinatorial treatment with very low levels of estrogens and leptin will decrease the effective dose of each hormone because of the enhanced activation of intracellular cascades, and at the same time this would reduce the anorexic side effects reported at higher doses. To our knowledge, estrogenic enhancement of leptin function has not previously been tested with respect to AN.

Expression of ghrelin and leptin during the development of type 2 diabetes mellitus in a rat model

The aim of this study was to investigate the syste-matic changes in ghrelin and leptin expression, as well as their correlation with insulin resistance (IR) during the development of type2 diabetes mellitus (T2DM) in a rat model. T2DM was induced in rats fed a high-fat (HF)-diet followed by the intraperitoneal injection of low-dose streptozotocin (STZ, 35mg/kg). Sixty male Sprague-Dawley rats were divided into 4groups: the control, HF-4W (HF diet for 4weeks), HF-8W (HF diet for 8weeks) and the T2DM group. During the development of T2DM, the production of ghrelin in the stomach and leptin in adipose tissue, the blood levels of ghrelin and leptin, and the expression of leptin and ghrelin receptors (OB-Rb and GHS-R1a) in the hypothalamus were measured by enzyme-linked immunosorbent assay (ELISA), radioimmunology assay (RIA), immunohistochemistry (IHC) and real-time reverse transcription-polymerase chain reaction (real-time RT-PCR). IR was assessed using the hyperinsulinemic-euglycemic clamp technique. The production of ghrelin in the stomach, the plasma ghrelin levels and the expression of GHS-R1a in the hypothalamus were significantly reduced in the HF-4W and HF-8W rats compared with the control rats; however, no significant difference was found between the HF-8W and T2DM group rats. Comparing the control to the T2DM group, the production of leptin in the adipose tissue and the serum leptin levels increased, whereas the expression of OB-Rb in the hypothalamus decreased. At the same time, the glucose infusion rate (GIR), indicating the insulin sensitivity, decreased significantly; GIR positively correlated with plasma ghrelin and negatively correlated with serum leptin levels. In conclusion, increased leptin levels are associated with obesity and T2DM, while decreased ghrelin levels are associated with obesity/IR rather than T2DM.

The effect of chronic immobilization stress on leptin signaling in the ovariectomized (OVX) rat

Previous studies have shown that both 17β-estradiol (E2) treatment and chronic stress may attenuate post-OVX weight gain in the female rat. However, the interaction between E2 and stress is unclear. This study examined the effect of E2 treatment and chronic immobilization stress on body weight. Adult OVX Sprague-Dawley rats were randomly assigned to one of four treatment groups in a 2X2 factorial design examining hormone treatment [vehicle (VEH) or E2, sc] and stress (no stress vs stress 60 min/day for 22 days). After 22 days, E2 significantly inhibited weight gain and food intake in OVX rats. In contrast, chronic stress reduced body weight only in control OVX animals but did not affect food intake. E2 reduced circulating leptin levels in non-stressed animals, but not in animals subjected to chronic immobilization. Western blot analysis indicated that E2 treatment increased leptin receptor (Ob-Rb) expression in the medial basal hypothalamus (MBH); however, this treatment also increased suppressor of cytokine signaling 3 (SOCS3), which is an inhibitor of leptin signaling. Chronic immobilization stress blunted the E2-induced increase in Ob-Rb and SOCS3 levels. These results suggest that chronic stress counteracts E2 effects on leptin signaling in the MBH without altering body weight.

Leptin and HER-2 are associated with gastric cancer progression and prognosis of patients

We conducted this study to evaluate the expression of leptin and its receptor, OB-Rb in gastric cancer and their relationship to clinicopathological features, VEGF and HER-2 expression, as well as the prognostic value. One hundred and ten gastric cancer specimens were detected for leptin, OB-Rb, VEGF and HER-2 by immunohistochemistry (IHC), and 96 specimens of normal gastric mucosa served as the control. The expression level of leptin, OB-Rb and HER-2 in gastric tissues were significantly higher than normal tissues (49.1% vs. 34.0%, 60.9% vs. 46.0%, 19.1% vs. 8.0%, P<0.05). There was a correlation between the expression of leptin and HER-2, both of which were significantly associated with invasion depth, lymph node metastasis, AJCC stage and VEGF expression. However, there was no correlation between OB-Rb expression and all clinicopathological features. Cox regression analyses showed that age, tumor size, histological grade, serosa invasion, AJCC stage, chemotherapy, leptin and HER-2 overexpression were prognostic factors. The survival of patients with leptin positive expression was significantly poorer than those with negative expression (OS: 20.0months vs. 32.5months, FPS: 12.0months vs. 18.0months, P<0.01). Leptin, rather than OB-Rb, played an important role in the progression and angiogenesis of gastric cancer, and was a prognostic factor for poor outcome.

CPU86017-RS attenuate hypoxia-induced testicular dysfunction in mice by normalizing androgen biosynthesis genes and pro-inflammatory cytokines

Downregulation of androgen biosynthesis genes StAR (steroidogenic acute regulatory) and 3β-HSD (3β-hydroxysteroid dehydrogenase) contributes to low testosterone levels in hypoxic mice and is possibly related to increased expression of pro-inflammatory cytokines in the testis. The aim of this study is to investigate the effects of CPU86017-RS that block Ca(2+) influx on hypoxia-induced testis insult in mice. Male ICR mice were divided into 5 groups: control group, hypoxia group, hypoxia group treated with nifedipine (10 mg/kg), hypoxia groups treated with CPU86017-RS (60 or 80 mg/kg). Hypoxia was induced by placing the mice in a chamber under 10%±0.5% O2 for 28 d (8 h per day). The mice were orally administered with drug in the last 14 d. At the end of experiment the testes of the mice were harvested. The mRNA and protein levels of StAR, 3β-HSD, connexin 43 (Cx43), matrix metalloprotease 9 (MMP9), endothelin receptor A (ET(A)R) and leptin receptor (OBRb) were analyzed using RT-PCR and Western blotting, respectively. The malondialdehyde (MDA), lactate dehydrogenase (LDH), succinate dehydrogenase (SDH) and acid phosphatase (ACP) levels were measured using biochemical kits. Serum testosterone concentration was measured with radioimmunoassay. Hypoxia significantly increased the MDA level, and decreased the LDH, ACP and SDH activities in testes. Meanwhile, hypoxia induced significant downregulation of StAR and 3β-HSD in testes responsible for reduced testosterone biosynthesis. It decreased the expression of Cx43, and increased the expression of MMP9, ETAR and OBRb, leading to abnormal testis function and structure. These changes were effectively diminished by CPU86017-RS (80 mg/kg) or nifedipine (10 mg/kg). Low plasma testosterone level caused by hypoxia was due to downregulation of StAR and 3β-HSD genes, in association with an increased expression of pro-inflammatory cytokines. These changes can be alleviated by CPU86017-RS or nifedipine.

112 OVARIAN TUMORS AND THE MANNER OF THEIR SURGICAL TRETMENT IN GOC KRAGUJEVAC SERBIA

Objectives: To investigate the molecular mechanisms of leptin on bone, the functions of different leptin receptors (OB-Ra and OB-Rb) on the membrane of osteoblasts and their signal transduction pathways. Material andmethods:Wistar rats were randomly divided into four groups, sham operation with ordinary diet group (n=20), sham operation with high nutrition group (n=20), ovariectomy with ordinary diet group (n=20), and ovariectomy with high nutrition group (n=20). mRNA expressions of OB-Ra and OB-Rb were measured. Results: PCR electrophoresis results showed lost expression of OB-Rb in two high-nutrition groups. In the sham operation with high nutrition group, the loss of OB-Rb expression accounts for 20%. In the ovariectomy with high nutrition group, this number grows to 60%. In sham operation with high nutrition group, ovariectomy with normal diet group and ovariectomy with high-nutrition group, the lost expression of short receptor OB-Ra accounts for 40%, 20%, and 20% respectively. By semi-quantitative analyzing the electrophoresis on mRNA expressions did not show significant differences between different groups. Conclusions: The physiological functions of leptin on bone may be executed through regulating receptors express differentially. Obesity and nutritional factors might influence expression of the same receptors.

Correlation between Obesity and Inflammation in Cardiovascular Diseases—Evaluation of Leptin and Inflammatory Cytokines

Obesity, an important risk factor for cardiovascular disease (CVD), is currently viewed as a pro-inflammatory state with an increase in the expression of inflammatory cytokines including Tumor Necrosis Factor-α (TNF-alpha) and Interleukin-6 (IL-6) along with the increased secretion of leptin. Serum leptin was estimated in 150 CVD patients (belonging to the age group of 35 - 75 years) of different Body Mass Index (BMI) and its correlation to various biochemical parameters including lipids, glucose and CRP was analyzed to study the association of leptin with CVD. Further to understand the relationship between obesity, inflammation and CVD, serum leptin, TNF-alpha, IL-6 was evaluated in 50 CVD and 50 non-CVD subjects. Results were presented as mean ± SD. A distinct correlation between leptin, TNF-alpha, IL-6 and BMI was observed and these cytokines were elevated in CVD subjects. The protein expression of the monomeric form of CRP which is known to be expressed in pathological condition was found to be highly expressed in serum of CVD subjects. Further, to substantiate the role of inflammation in obesity related CVD, different concentrations of leptin were added to Peripheral Blood Mononuclear Cells (PBMC) isolated from twenty healthy volunteers of different BMI and the gene expression of TNF-alpha, IL-6 and Ob-Rb were studied. A significant increase in the level of these cytokines was observed with increasing leptin concentration. The results obtained from the present analysis suggest that inflammation may be a crucial factor involved in obesity associated CVD.

Ingestion of Carbohydrate-Rich Supplements during Gestation Programs Insulin and Leptin Resistance but not Body Weight Gain in Adult Rat Offspring

Prenatal nutritional conditions can predispose to development of obesity and metabolic syndrome in adulthood. Gestation with its important modifications in hormonal status is a period of changes in normal feeding habits with pulses of consumption or avoidance of certain categories of food. We tried to mimic in an animal model some changes in food consumption patterns observed in pregnant women. For this purpose, Long–Evans female rats were fed during the dark period, their usual pre-gestational food quantity, and were allowed to complete their daily intake with either a restricted control (Cr), high-fat (HF), or high-carbohydrate (HC) diet available ad libitum during the light period. Dams fed a control diet ad libitum (Ca) served as controls. Body weight and composition, food intake, and metabolic hormones (insulin, leptin) were recorded in male offspring until 20 weeks after birth. Cr and HC females ate less than Ca females (−16%; p < 0.001) and their offspring presented a weight deficit from birth until 6 (HC group) and 10 (Cr group) weeks of age (p < 0.05 or less). Plasma leptin corresponded to low body weight in Cr offspring, but was increased in HC offspring that in addition, had increased plasma insulin, blood glucose, and subcutaneous adipose tissue mass. HF dams ate more than Ca dams (+13%; p < 0.001), but plasma leptin and insulin were similar in their offspring. Hypothalamic Ob-Rb expression was increased in Cr, HC, and HF offspring (+33–100% vs Ca; p < 0.05 or less). HC supplement ingestion during gestation therefore leads to insulin and leptin resistance in adult offspring independently of lower birth weight. These hormonal changes characterize obesity-prone animals. We therefore suggest that attention should be paid to the carbohydrate snacking and overall carbohydrate content in the diet during the last weeks (or months) preceding delivery to limit development of later metabolic disorders in offspring.

Leanness in postnatally nutritionally programmed rats is associated with increased sensitivity to leptin and a melanocortin receptor agonist and decreased sensitivity to neuropeptide Y.

BackgroundPups of normally nourished dams that are cross-fostered after birth to dams fed a low protein (8% by weight) diet (postnatal low protein; PLP) grow slower during the suckling period and remain small and lean throughout adulthood. At weaning, they have increased expression in the arcuate nucleus (ARC) of the hypothalamus of the orexigenic neuropeptide Y (NPY) and decreased expression of pro-opiomelanocortin (POMC), the precursor of anorexigenic melanocortins.Objectives and methodsWe investigated, using third ventricle administration, whether 3-month-old male PLP rats display altered sensitivity to leptin with respect to food intake, NPY and the melanocortin 3/4-receptor agonist MTII, and using in situ hybridization or laser capture microdissection of the ARC followed by RT-PCR, whether the differences observed were associated with changes in the hypothalamic expression of NPY, or the leptin receptor, NPY receptors, and melanocortin receptors.ResultsPLP rats were smaller and had reduced percentage body fat content and plasma leptin concentration compared to control rats. Leptin (5 μg) reduced food intake over 0-48 h more in PLP than control rats (P<0.05). Submaximal doses of NPY increased food intake less in PLP rats than controls, whilst submaximal doses of MTII reduced food intake more in the PLP rats. Maximal responses did not differ between postnatal low protein and control rats. Leptin (ObRb) and melanocortin-3 (MC3R) receptor expression were increased in both ARC and ventromedial (VMH) hypothalamic nuclei in PLP animals compared to the controls. MC4R, NPY Y1R and Y5R and NPY expression were unchanged.ConclusionPostnatal undernourishment results in food intake in adult rats being more sensitive to reduction by leptin and melanocortins, and less sensitive to stimulation by NPY. We propose that this contributes to increased leptin sensitivity and resistance to obesity. Increased expression of ObRb and MC3R may partly explain these findings but other downstream mechanisms must also be involved.