Ob-R mRNA Research Articles

Leptin Promotes Prostate Cancer Proliferation and Migration by Stimulating STAT3 Pathway

To better understand the link between obesity and prostate cancer (PC) aggressiveness, we investigate the role of leptin, an obesity associated adipokine, and its receptor (Ob-R) in PC cells migration. The migration assay (Wound-healing) was used to study the leptin impact on DU-145 and PC3 cells lines. STAT3 activation was performed by Western Blot. E-cadherin expression was studied using fluorescence microscopy and Ob-R expression in PC and benign prostatic Hyperplasia (BPH) biopsies was assessed by RT-PCR. In this study we demonstrate that high dose of leptin promotes PC cells migration and EMT transition via the stimulation of STAT3 pathway. In addition, we report that although Ob-R mRNA is expressed by ADK and BPH resections biopsies, significant higher levels were observed for ADK patients. Finally, we found a positive association between Ob-R mRNA expression and worse PC prognosis. A better understanding of the molecular processes of leptin signaling is crucial for identifying appropriate approaches for treatment of obesity-related PC patients.

Dietary protein restriction increases hepatic leptin receptor mRNA and plasma soluble leptin receptor in male rodents.

Leptin is an adipokine that regulates adipose tissue mass through membrane-anchored leptin receptor (Ob-R). Extracellular domain of Ob-R in plasma is called soluble leptin receptor (sOb-R), and is the main leptin-binding protein. Based on a previous DNA microarray analysis that showed induction of hepatic Ob-R mRNA in low-protein diet-fed mice, this study aimed to clarify the effect of dietary protein restriction on hepatic Ob-R mRNA and plasma sOb-R levels. First, the effect of protein restriction on hepatic Ob-R mRNA level was examined together with fasting and food restriction using male rats as common experimental model for nutritional research. Hepatic Ob-R mRNA level was increased by feeding low-protein diet for 7 d, although not significantly influenced by 12-h fasting and sixty percent restriction in food consumption. Then, effect of protein restriction on liver Ob-R and plasma sOb-R was investigated using male mice because specific sOb-R ELISA was more available for mice. Hepatic Ob-R mRNA level was also increased in protein restricted-mice although it did not increase in hypothalamus. Hepatic Ob-R protein was decreased, whereas plasma sOb-R was increased by protein restriction. Because the concentration of sOb-R increased without changing plasma leptin concentration, free leptin in plasma was significantly reduced. The direct effect of amino acid deprivation on Ob-R mRNA level was not observed in rat hepatoma cells H4IIE cultured in amino acid deprived medium. In conclusion, dietary protein restriction increased hepatic Ob-R mRNA, resulting in increased plasma sOb-R concentration, which in turn, reduces plasma free leptin level and may modulate leptin activity.

The Effect of Notch, IL‐1 and Leptin (NILCO) Inhibition in Xenograft Colorectal Cancer

The aim of this study was to investigate the effects of inhibition of Notch, IL‐1 and leptin systems on tumorigenesis and angiogenesis in xenograft colorectal cancer model. This study was conducted with the approval of the EU/2010/63 directive and the Greek Presidential Decree 56/2013 (Approval No. 5542/228006/1). In each group, 40 scid (NOD/SCID) mice with 6–8 weeks of age were studied in 5 groups. For tumor formation 1 × 107 human colon cancer cells HCT‐15 were injected subcutaneously into the dorsal flank of the mice. When the tumor size was 100–250 mm3, treatment was started. Group 1: Leptin receptor antagonist ALLO‐aca was injected with 1mg / kg / day subcutaneously (s.c) for 15 days (n: 8). Group 2: γ‐secretase inhibitor DAPT was injected with 10 mg / kg / day subcutaneously (s.c) for 15 days (n: 8). Group 3: Recombinant human IL‐1 receptor antagonist (IL‐1Ra) was injected with 1mg / kg / day subcutaneously (s.c) for 15 days (n: 8). Group 4: 4% DMSO/Safflower Oil (DAPT solvent) was injected subcutaneously (s.c) for 15 days. (n: 8). Group 5: 10% DMSO/0,9%NaCl (Allo aca and IL‐1 iınhibitor solvent) was injected subcutaneously (s.c) for 15 days. (n: 8). After 15 days, tissues were collected and gene expression experiments were performed. mRNA abundance of leptin, leptin OB‐RL receptor, Notch1 receptor, JAGGED1 ligand, IL‐1β, IL‐1R tI, VEGF‐A, VEGFR2 conducted by qRT‐PCR. According to PCR results, leptin, its receptor Ob‐R, Notch1, JAGGED1, VEGF‐A and VEGFR1 mRNA abundance were decreased only in the DAPT group (respectively; p<0.05, p<0.05, p<0.05, p<0.01, p<0.05, p<0.01). The mRNA abundance of VEGFR2 was decreased only in the IL‐1 inhibitor group (p<0.05). According to the results, DAPT at 10 mg / kg / day dose Notch receptor and its ligand was reduced as well as decreased mRNA abundance of leptin and VEGF systems that pro‐angiogenogenic factors. The results of our study support the oncogenic role of Notch, IL‐1 and Leptin crosstalk in colorectal cancer. Inhibition of NILCO signaling could lead to the development of new adjuvant therapies to reduce or eliminate tumor angiogenesis.Support or Funding InformationThis study is supported by a grant from Eskisehir Osmangazi University Scientific Research Projects (Project no:2017‐11028)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Expression of Leptin Receptor and Effects of Leptin on Papillary Thyroid Carcinoma Cells.

Background Obesity has been hypothesized to contribute to the aggressiveness of thyroid cancer through the production of abnormal levels of serum adipokines. Leptin receptor (OB-R) expression has also been documented in papillary thyroid cancer (PTC). Aim In this translational study, we analyzed in vitro the effects of leptin on the growth and migration of thyroid cancer cells (TPC-1 and K1), the molecular mechanisms underlying leptin's action, and the influence of prolonged leptin exposure on cell response to a protein kinase inhibitor lenvatinib. The expression levels of OB-R mRNA and protein were also investigated in vivo in a series of aggressive PTCs divided into two groups based on the presence of the BRAF mutation. Results In TPC-1 and K1 cells, prolonged treatment with leptin (500 ng/ml for 96 h) resulted in a mild increase in the proliferation (about 20% over control only in K1 cells, p < 0.05) and in the migration of both cancer cell lines. Immunoblot analysis revealed a slight increase in the phosphorylation of AKT, but no effect on β-catenin and phospho-ERK expressions. The inhibitory effects of lenvatinib on the viability of both cell lines were not influenced by the leptin treatment. OB-R transcript (in fresh tissues) and proteins (in formalin-fixed and paraffin-embedded specimens) were expressed in all PTC tissues examined, with no significant differences between BRAF-mutated and BRAF-wild-type tumors. Conclusions These results demonstrate leptin's role in mildly increasing the aggressive phenotype of PTC cells but without influencing the action of lenvatinib. Further studies will clarify whether it is possible to target OB-R, expressed in all aggressive PTCs, as an adjuvant treatment approach for these malignancies.

Leptin/leptin receptor system in the regulation of reproductive functions and stress response in the European beaver.

The European beaver (Castor fiber L.) is the largest free-living rodent in Eurasia. The present work aimed to determine sex- and season-related changes in leptin receptor (Ob-R) expression in the hypothalamic–pituitary–gonadal/adrenal axes and uterus of beavers during breeding- (April), post-breeding- (July), and pre-breeding- (November) periods. The expression of Ob-R gene and protein was found in all analyzed tissues. The expression of Ob-R mRNA remained constant in the hypothalamus of both sexes during the analyzed stages. Sex- and season-related changes were found in the pituitary gland; the greatest level was observed in July in both sexes. The same expression pattern was noted in the testis, whereas in the ovary a lack of seasonal changes was found. In uterine tissues, the greatest expression occurred in November. The impact of season was also demonstrated in the adrenal cortex. In females, a higher Ob-R transcript level was noted in April, while in males, an increased mRNA abundance was noted in November than July. Our study suggests that in the beaver, leptin acting via the Ob-R can be an important endocrine factor engaged in the regulation of reproductive functions and stress response.

Effect of valsartan on the expression of leptin, leptin receptor and collagen in rats with hepatic fibrosis

Objective: To investigate the effects of angiotensin II type 1 receptor antagonist valsartan on leptin, leptin receptor and collagen in rats with hepatic fibrosis. Methods: Thirty-six male wistar rats were randomly divided into control group, model group and drug-treated group, with 12 rats in each group. Liver fibrosis models were made by subcutaneous injection of carbon tetrachloride on the dorsal of the rats, simultaneously gastric gavage with Valsartan and were killed at the end of 8th week. The degree of liver fibrosis was observed by HE and Masson staining. The serum leptin (LP) and TGFβ1 were determined by ELISA. Liver LP mRNA and leptin receptor mRNA (OB-R mRNA) were detected by RT-PCR. Liver LP, OB-R and collagen I were detected by Western blot. The data of multiple groups were analyzed by one-way analysis variance (ANOVA), and linear correlation was performed between serum LP and TGF β1. Results: After the intervention of valsartan, HE and Masson staining showed that the degree of liver fibrosis was significantly reduced. The levels of serum LP and TGFβ1 in the control group were (18.92 ± 7.10) ng/ml and (9.13 ± 1.58) pg/ml respectively, which were significantly lower than those in the model group (46.92 ± 28.54) ng/ml and (16.39 ± 3.56) pg/ml, And (29.27 ± 7.27) ng/ml and (12.24 ± 2.94) pg/ml in the drug-treated group, respectively. The F values were 7.864 and 20.057 respectively. The P values were < 0.05. The differences were statistically significant. The relative expression levels of LP and OB-R mRNA in the control group were 0.35 ± 0.18 and 0.62 ± 0.18, respectively, which were significantly lower than those in the model group (1.79 ± 1.79 and 1.52 ± 1.44, and drug-treated group 0.48 ± 0.34 and 0.75 ± 0.26, respectively), F values = 6.914,3.894, P values were < 0.05, the differences were statistically significant. The relative expression levels of LP, OB-R and collaten I in liver were 0.71 ± 0.13, 0.81 ± 0.11 and 0.76 ± 0.13 in the model group, 0.97 ± 0.06, 1.04 ± 0.06, and 1.05 ± 0.04 respectively in the drug-treated group and 0.74 ± 0.05, 0.93 ± 0.05 and 0.91 ± 0.05. The F values were 15.425, 13.757 and 19.130 respectively in three groups (P < 0.001), the difference was statistically significant. Conclusion: Valsartan, an angiotensin II type 1 receptor antagonist, can reduce the expression of leptin and leptin receptor, reduce the production of TGFβ1 and collaten I, and play an anti-hepatic fibrosis effect.

Abstract A09: Role of adipocytokines in obesity associated Triple Negative Breast Cancer

Abstract Background: Triple-negative breast cancer (TNBC), accounts for a large percentage of breast cancer cases in India including Northeast India. TNBC has an unclear molecular aetiology and hence limited targeted therapies. Human breast is comprised of glandular, ductal, connective, and adipose tissues. Adipose tissue is composed of adipocytes. The adipocytes apart from being energy storage depots are also active sources of adipocytokines and/or adipokines. The role of adipokines in breast cancer including TNBC has been sporadically documented. Two adipokines in particular, leptin and adiponectin, have come to be recognized for their influence on breast cancer risk and tumor biology. Therefore, the aim of this study was to understand the association of differential expression of critical adipokines and associated cellular mechanism in the susceptibility and severity of TNBC in northeast Indian population. Materials and Methods: We collected 68 TNBC and 63 controls cases and examined for serum leptin and adiponectin levels using enzyme linked immunosorbent assay (ELISA). Leptin Receptor (Ob-R) mRNA expression was determined by real-time polymerase chain reaction (RT-PCR) assay. Differential Ob-R mRNA expression and correlation with cancer stem cell (CSC) markers oct4 and nanog was evaluated, and correlated with severity. Results: The serum leptin levels were significantly associated with TNBC severity, while the adiponectin levels were comparative. The serum leptin levels correlated inversely with the adiponetin levels. Serum leptin levels were unaffected with difference in parity. The difference in leptin levels in pre and post menopausal cases were found to be statistically non-significant. Higher leptin levels were also found to be associated obesity, mortality and recurrence. Obesity was found to be a factor for TNBC pathogenesis and severity. Increased Ob-R mRNA expression was associated with TNBC, significantly with TNBC severity, and was significantly higher in obese patients with higher grade TNBC cases. The Ob-R gene mRNA expression was significantly higher in the obese TNBC cases showing recurrence or mortality. The higher Ob-R gene mRNA expression correlated significantly with higher serum leptin levels and lower serum adiponectin levels in TNBC cases. The Ob-R mRNA expression with associated with modulation of CSC. Conclusions: In conclusion, the present study is first of its kind on TNBC from northeast India, indicates that adipocytokines does play a role in TNBC pathogenesis. Thus, the understanding of molecular mechanisms of both leptin and adiponectin and their interplay in TNBC offer the prospects for new therapeutic approaches targeting similar signaling pathways. Note: This abstract was not presented at the conference. Citation Format: Rizwana Sultana, Amal Ch Kataki, Bibhuti Bhusan Barthakur, Sujoy Bose. Role of adipocytokines in obesity associated Triple Negative Breast Cancer [abstract]. In: Proceedings of the AACR International Conference: New Frontiers in Cancer Research; 2017 Jan 18-22; Cape Town, South Africa. Philadelphia (PA): AACR; Cancer Res 2017;77(22 Suppl):Abstract nr A09.

Imbalance in leptin-adiponectin levels and leptin receptor expression as chief contributors to triple negative breast cancer progression in Northeast India

BackgroundTriple-Negative breast cancer (TNBC), accounts for a large percentage of breast cancer cases in India including Northeast India. TNBC has an unclear molecular aetiology and hence limited targeted therapies. Human breast is comprised of glandular, ductal, connective, and adipose tissues. Adipose tissue is composed of adipocytes. The adipocytes apart from being energy storage depots, are also active sources of adipocytokines and/or adipokines. The role of adipokines in breast cancer including TNBC has been sporadically documented. Two adipokines in particular, leptin and adiponectin, have come to be recognized for their influence on breast cancer risk and tumour biology. Therefore, the aim of this study was to understand the association of differential expression of critical adipokines and associated cellular mechanism in the susceptibility and severity of TNBC in northeast Indian population. Materials and methodsWe collected 68 TNBC and 63 controls cases and examined for serum leptin and adiponectin levels using enzyme linked immunosorbent assay (ELISA). Leptin Receptor (Ob-R) mRNA expression was determined by real-time polymerase chain reaction (RT-PCR) assay. Differential Ob-R mRNA expression and correlation with cancer stem cell (CSC) markers was evaluated, and correlated with severity. ResultsThe serum leptin levels were significantly associated with TNBC severity, while the adiponectin levels were comparative. The serum leptin levels correlated inversely with the adiponetin levels. Serum leptin levels were unaffected with difference in parity. The difference in leptin levels in pre and post menopausal cases were found to be statistically non-significant. Higher leptin levels were also found to be associated obesity, mortality and recurrence. Obesity was found to be a factor for TNBC pathogenesis and severity. Increased Ob-R mRNA expression was associated with TNBC, significantly with TNBC severity, and was significantly higher in obese patients with higher grade TNBC cases. The Ob-R gene mRNA expression was significantly higher in the obese TNBC cases showing recurrence or mortality. The higher Ob-R gene mRNA expression correlated significantly with higher serum leptin levels and lower serum adiponectin levels in TNBC cases. The Ob-R mRNA expression with associated with modulation of CSC oct4 and nanog. ConclusionsIn conclusion, the present study is first of its kind on TNBC from northeast India, indicates that adipocytokines does play a role in TNBC pathogenesis. Thus, the understanding of molecular mechanisms of both leptin and adiponectin and their interplay in TNBC offer the prospects for new therapeutic approaches targeting similar signalling pathways.

Knockdown of the AKT3 (PKBγ), PI3KCA, and VEGFR2 genes by RNA interference suppresses glioblastoma multiforme T98G cells invasiveness in vitro.

Glioblastoma multiforme (GBM) is the most common primary brain malignancy, having a very poor prognosis and is characterized by extensive brain invasion as well as resistance to the therapy. The phosphoinositide 3-kinase (PI3K)/Akt/PTEN signaling pathway is deregulated in GBM. Besides, florid vascularization and aberrantly elevated vascular endothelial growth factor (VEGF) occur very often. The present study was designed to examine the inhibitory effect of AKT3, PI3KCA, and VEGFR2 small interfering RNAs (siRNAs) on GBM cell invasiveness. T98G cells were transfected with AKT3, PI3KCA, and/or VEGFR2 siRNAs. VEGFR2 protein-positive cells were identified by flow cytometry using specific monoclonal anti-VEGFR2 antibodies. Alterations in messenger RNA (mRNA) expression of VEGF, VEGFR2, matrix metalloproteinases (MMPs) (MMP-2, MMP-9, MMP-13, MMP-14), tissue inhibitors of metalloproteinases (TIMPs) (TIMP-1, TIMP-3), c-Fos, c-Jun, hypoxia-inducible factor-1α (HIF-1α), ObRa, and cathepsin D genes were analyzed by qRT-PCR. Cells treated with specific siRNA were also analyzed for invasion using the Matrigel invasion assay. We have found significantly lower mRNA levels of MMPs, cathepsin D, VEGF, VEGFR2, HIF-1α, and c-Fos/c-Jun ratio, as well as significantly higher mRNA level of TIMPs in AKT3 and PI3KCA siRNA transfected cells compared to untransfected cells, while significantly lower mRNA levels of MMPs (MMP-2, MMP-9, MMP-14) and TIMP-1, as well as significantly higher mRNA level of TIMP-3, were shown only in cells transfected with VEGFR2 siRNA. The positive correlation between MMP-13 and ObRa mRNA copy number has been found. Summarizing, transfection of T98G cells with AKT3, PI3KCA, or VEGFR2 siRNAs leads to a significant reduction in cell invasiveness. The siRNA-induced AKT3, PI3KCA, and VEGFR2 mRNA knockdown may offer a novel therapeutic strategy to reduce the invasiveness of GBM cells.

Leptin receptor is expressed by epidermis and skin appendages in dog

Leptin is a polypeptide secreted by adipocytes which binds to a specific receptor (Ob-R) that is expressed in various tissues. The wide distribution of the Ob-R suggests that leptin might exert diverse biological functions, not only by regulating energy metabolism and appetite, but also by acting as a mitogen in many cell types, including keratinocytes.In this study, the presence and localization of Ob-R was investigated in the skin of the dog using RT-PCR and immunohistochemical techniques. RT-PCR revealed the presence of Ob-R m-RNA in the skin specimens collected from the dorsal region of two smooth coat breed dogs. Through immunohistochemistry performed on the skin of five dogs, the expression of the receptor was observed in the basal layer of the epidermis, in the hair follicles as well as in the apocrine sweat and sebaceous glands. No staining for Ob-R was detected in the suprabasal epidermis layers. Strong positive signals were observed in many cells of the outer root sheath of hair follicles in growing and in regressive phases. The identification of Ob-R in the above targets suggests that leptin may play a role in the regulation of cyclic renewal of the epidermis and skin appendages in dog. This study represents an important contribution to understand the complex mechanisms that are involved in the skin biology in this species.

Leptin Induces a Proliferative Response in Breast Cancer Cells but Not in Normal Breast Cells

Obesity is a risk factor for breast cancer in postmenopausal women. Leptin, a hormone excessively produced during obesity, is suggested to be involved in breast cancer. The aim of the study was to investigate procarcinogenic potential of leptin by evaluating influence of leptin on cell proliferation, cell cycle, apoptosis, and signaling on numerous breast cells lines, including 184B5 normal cells, MCF10A fibrocystic cells and MCF-7, MDA-MB-231, and T47D cancer cells. Expressions of leptin and Ob-R were analyzed using qRT-PCR and immunohistochemistry, proliferation using fluorimetric resazurin reduction test and xCELLigence system, apoptosis and cell cycle by flow cytometry, and effect of leptin on different signalling pathways using qRT-PCR and Western blot. Cells were exposed to increasing concentrations of leptin. All cell lines expressed mRNA and protein of leptin and Ob-R. Leptin stimulated proliferation of all cell lines except for 184B5 and MDA-MB-231 cells. Leptin inhibited apoptosis but didn't alter proportion of cells within cell cycle in MCF7 cells. Leptin induced overexpression of leptin, Ob-R, estrogen receptor, and aromatase mRNA in MCF-7 and T47D cells. Autoregulation induced by leptin, relationship with estrogen pathway, and proliferative and antiapoptic activity in breast cancer cells may explain that obesity-associated hyperleptinemia may be a breast cancer risk factor.

Effects of Zinc Deficiency and Supplementation on Leptin and Leptin Receptor Expression in Pregnant Mice

Leptin is an adipose-derived hormone that primarily regulates energy balance in response to nutrition. Human placental cells produce leptin, whereas murine placental cells produce soluble leptin receptors (Ob-R). However, the roles of these proteins during pregnancy have not been elucidated completely. As an essential metal, zinc (Zn) is central to insulin biosynthesis and energy metabolism. In the present study, the effects of Zn deficiency and supplementation on maternal plasma leptin and soluble Ob-R regulation in pregnant mice placentas were examined using enzyme-linked immunosorbent assay, reverse transcription-polymerase chain reaction, and Western blotting. Nutritional Zn deficiency significantly reduced plasma insulin concentrations and fetal and placental weights in pregnant mice. Plasma leptin concentrations in pregnant mice also increased 20- to 40-fold compared with those in non-pregnant mice. Although dietary Zn deficiency and supplementation did not affect plasma leptin concentrations in non-pregnant mice, Zn-deficient pregnant mice had significantly reduced plasma leptin concentrations and adipose leptin mRNA expression. In contrast, Zn-supplemented pregnant mice had increased plasma leptin concentrations without increased adipose leptin mRNA expression. Placental soluble Ob-R mRNA expression also decreased in Zn-deficient mice and tended to increase in Zn-supplemented mice. These results indicate that Zn influences plasma leptin concentrations by modulating mRNA expression of soluble Ob-R in the placenta, and leptin in visceral fat during pregnancy. These data suggest that both adipose and placenta-derived leptin system are involved in the regulation of energy metabolism during fetal growth.

Leptin receptor mRNA expression in Taleshi ram gonads

Leptin is secreted mainly by fat, which is involved in energy metabolism and reproduction. Leptin and its receptor (Ob-R) have been detected in human spermatozoa and testis, thus it can be concluded leptin involves in male physiology. The goal of this study was determination of the presence of leptin receptor mRNA in the ram testis, epididymis and spermatozoa by reverse transcription polymerase chain reaction (RT-PCR). Ejaculated spermatozoa from ten fertile Taleshi ram were collected by means of an artificial vagina. Testes, placental cotyledons and adrenal glands were obtained from abattoir. Placental cotyledons and adrenal glands were used as the positive control. Epididymal spermatozoa recovery was performed from epididymis. To purify spermatozoa, motile sperm were isolated by the swim-up procedure. Total RNA was isolated from epididymal spermatozoa, ejaculated spermatozoa, adrenal gland and placental cotyledon and then they were purified. The mRNA for the long form (Ob-Rb) and the short form (Ob-Ra) of leptin receptor was detected in testis. RT-PCR analysis of total RNA from epididymal spermatozoa and ejaculated spermatozoa revealed the presence of leptin mRNA in these cells. The mRNA for Ob-Rb was observed in epididymis and epididymal spermatozoa, but the Ob-Ra mRNA was absent. The presence of Ob-Ra mRNA was found in ejaculated spermatozoa, whereas Ob-Rb mRNA did not exist. It can be concluded that the mRNA for leptin receptor is present in ram gonads and spermatozoa.

Hunting for the functions of short leptin receptor isoforms

Hunting for the functions of short leptin receptor isoforms

Leptin receptor (Ob-R) mRNA expression and serum leptin concentration in patients with colorectal and metastatic colorectal cancer

The objective of the present study was to investigate the effect of leptin on the progression of colorectal carcinoma to metastatic disease by analyzing the serum leptin concentration and Ob-R gene expression in colon cancer tissues. Tissue samples were obtained from 31 patients who underwent surgical resection for colon (18 cases) and metastatic colon (13 cases) cancer. Serum leptin concentration was determined by an enzyme-linked immunosorbent assay (ELISA) and Ob-R mRNA expression by real-time polymerase chain reaction (RT-PCR) for both groups. ELISA data were analyzed by the Student t-test and RT-PCR data were analyzed by the Mann-Whitney U-test. RT-PCR results demonstrated that mRNA expression of Ob-R in human metastatic colorectal cancer was higher than in local colorectal cancer tissues. On the other hand, mean serum leptin concentration was significantly higher in local colorectal cancer patients compared to patients with metastatic colorectal cancer. The results of the present study suggest a role for leptin in the progression of colon cancer to metastatic disease without weight loss. In other words, significantly increased Ob-R mRNA expression and decreased serum leptin concentration in patients with metastatic colon cancer indicate that sensitization to leptin activity may be a major indicator of metastasis to the colon tissue and the determination of leptin concentration and leptin gene expression may be used to aid the diagnosis.

Pressure mediated hypertrophy and mechanical stretch up-regulate expression of the long form of leptin receptor (ob-Rb) in rat cardiac myocytes

BackgroundHyperleptinemia is known to participate in cardiac hypertrophy and hypertension, but the relationship between pressure overload and leptin is poorly understood. We therefore examined the expression of leptin (ob) and the leptin receptor (ob-R) in the pressure-overloaded rat heart. We also examined gene expressions in culture cardiac myocytes to clarify which hypertension-related stimulus induces these genes.ResultsPressure overload was produced by ligation of the rat abdominal aorta, and ob and ob-R isoform mRNAs were measured using a real-time polymerase chain reaction (PCR). We also measured these gene expressions in neonatal rat cardiac myocytes treated with angiotensin II (ANGII), endothelin-1 (ET-1), or cyclic mechanical stretch. Leptin and the long form of the leptin receptor (ob-Rb) gene were significantly increased 4 weeks after banding, but expression of the short form of the leptin receptor (ob-Ra) was unchanged. ob-Rb protein expression was also detected by immunohistochemistry in hypertrophied cardiac myocytes after banding. Meanwhile, plasma leptin concentrations were not different between the control and banding groups. In cultured myocytes, ANGII and ET-1 increased only ob mRNA expression. However, mechanical stretch activated both ob and ob-Rb mRNA expression in a time-dependent manner, but ob-Ra mRNA was unchanged by any stress.ConclusionsWe first demonstrated that both pressure mediated hypertrophy and mechanical stretch up-regulate ob-Rb gene expression in heart and cardiac myocytes, which are thought to be important for leptin action in cardiac myocytes. These results suggest a new local mechanism by which leptin affects cardiac remodeling in pressure-overloaded hearts.

Upregulation of Astrocytic Leptin Receptor in Mice with Experimental Autoimmune Encephalomyelitis

The detrimental role of leptin in experimental autoimmune encephalomyelitis (EAE) is opposite to its neuroprotective role in other neuropathologies. We hypothesize that a shifted cellular distribution of leptin receptors underlies the differential effects of leptin. A robust increase of ObR immunoreactivity was seen along glial fibrillary acidic protein (GFAP)(+) intermediate filaments in reactive astrocytes in the hippocampus and hypothalamus of mice with EAE. Although astrocyte-specific GFAP mRNA and protein were both increased, ObRa mRNA was elevated only after resolution of EAE symptoms, and ObRb mRNA was even decreased at the peak time of symptoms of EAE. A cell type-specific action of leptin may underlie its differential effects.

Estrogen stimulates leptin receptor expression in ATDC5 cells via the estrogen receptor and extracellular signal-regulated kinase pathways

Regulation of the physiological processes of endochondral bone formation during long bone growth is controlled by various factors including the hormones estrogen and leptin. The effects of estrogen are mediated not only through the direct activity of estrogen receptors (ERs) but also through cross talk with other signaling systems implicated in chondrogenesis. The receptors of both estrogen and leptin (OBR (LEPR)) are detectable in growth plate chondrocytes of all zones. In this study, the expression of mRNA and protein of OBR in chondrogenic ATDC5 cells and the effect of 17β-estradiol (E(2)) stimulation were assessed using quantitative PCR and western blotting. We have found that the mRNA of Obr was dynamically expressed during the differentiation of ATDC5 cells over 21 days. Application of E(2) (10(-7) M) at day 14 for 48 h significantly upregulated OBR mRNA and protein levels (P<0.05). The upregulation of Obr mRNA by E(2) was shown to take place in a concentration-dependent manner, with a concentration of 10(-7) M E(2) having the greatest effect. Furthermore, we have confirmed that E(2) affected the phosphorylation of ERK1/2 (MAPK1/MAPK3) in a time-dependent manner where a maximal fourfold change was observed at 10 min following application of E(2). Finally, pretreatment of the cells with either U0126 (ERK1/2 inhibitor) or ICI 182 780 (ER antagonist) blocked the upregulation of OBR by E(2) and prevented the E(2)-induced phosphorylation of ERK. These data demonstrate, for the first time, the existence of cross talk between estrogen and OBR in the regulation of bone growth whereby estrogen regulates the expression of Obr in growth plate chondrocytes via ERs and the activation of ERK1/2 signaling pathways.

Leptin Receptor in Ram Epididymal Spermatozoa

This experiment was designed to investigate leptin receptors (Ob-R and Ob-Rb) mRNA in ram epididymal spermatozoa by RT-PCR. Ten testes were obtained from abattoir and epididymal spermatozoa recovery was performed. To purify spermatozoa, motile sperm were isolated by the swim-up procedure. Total RNA was isolated from epididymal spermatozoa and placental cotyledon, as a positive control, and then they were purified. Specific bands (98 and 308 bp) for Ob-R and Ob-Rb were detected after RT-PCR in both epididymal spermatozoa and placental cotyledon. We may conclude that Ob-R and Ob-Rb mRNA are present in ram epididymal spermatozoa and leptin perhaps exerts physiological effects, as already demonstrated in human.

Expression of leptin and leptin receptor isoforms in the rat and human carotid body

Leptin is known to play a role in the modulation of metabolism and control of breathing acting mainly on central nervous structures, although additional actions on peripheral arterial chemoreceptors have also been suggested in the literature. We therefore examined by means of immunohistochemistry the expression of leptin and leptin receptors in the carotid bodies of rats and humans. Leptin expression and relative expression of leptin receptor isoforms were also studied in rats by real-time PCR. No leptin or leptin receptor immunoreactivities were visible in the type II cells of either series. In rat carotid bodies, diffuse positive stainings for leptin and leptin receptors (both with antibody recognizing all receptor isoforms and antibody specific for Ob-Rb) were observed in type I cells. In human carotid bodies, the mean percentage (±standard error) of leptin immunoreactive type I cells was 39.4%±5.1% and the percentages of leptin receptor immunoreactive type I cells were 57.3%±3.9% with antibody recognizing all receptor isoforms and 33.3%±4.2% with antibody specific for isoform Ob-Rb. Double immunofluorescences with anti-tyrosine hydroxylase (type I cell marker) and anti-glial fibrillary acidic protein (type II cell markers) confirmed the selective location of leptin and Ob-Rb in type I cells. Real-time PCR showed the expression of leptin and Ob-Ra, Ob-Rb, Ob-Rc and Ob-Rf isoform mRNA in the rat carotid body, levels of expression being Ob-Rf>Ob-Rc>>Ob-Ra>>Ob-Rb. Ob-Re mRNA was not detected. The above findings suggest a role of circulating or locally produced leptin in the regulation of chemoreceptor discharge and/or metabolic sensing function, by means of direct action on type I cells.