Leptin signalling in teleost fish with emphasis in food intake regulation.

Leptin, the product of the obese (ob or Lep) gene, was first cloned in teleost fish in 2005, more than a decade after its identification in mammals. This was because bony fish and mammalian leptins share a very low amino acid sequence identity, which suggests different functionality of the leptin system in fish compared to that of mammals. Indeed, major differences are evident between the mammalian and fish leptin system. Thus, for instance, mammalian leptin is synthesized and released by the adipose tissue in response to the amount of fat depots, while several tissues (mainly the liver) are the main sources of leptin in fish, whose determining factors of production are still unclear. In mammals, the main physiological role for leptin is its involvement in the maintenance of energy balance by decreasing food intake and increasing energy expenditure, although a wide variety of actions have been attributed to this hormone (e.g., regulation of lipid and carbohydrate metabolism, reproduction and immune functions). In fish, available literature also points towards a multifunctional nature for leptin, although knowledge on its functions is limited. In this review, we offer an overview of teleostean leptin structure and mechanism of action, and discuss the available knowledge on the role of this hormone in food intake regulation in teleost fish, aiming to provide a comparative overview between the functioning of the teleostean and mammalian leptin systems.

Salivary concentrations of cytokines and other analytes in healthy children.

BACKGROUND: Blood has been the usual biological fluid for measuring analytes, but there is mounting evidence that saliva may be also useful for detecting cytokines in a noninvasive way. Thus, in this study we aimed to determine concentration of cytokines and other analytes in saliva from a population of healthy children. METHODS: We collected un-stimulated whole saliva samples from clinically healthy children, and concentration of 17 cytokines and 12 other analytes were measured in supernatants. All values were adjusted by albumin content and were log-transformed before multivariate statistical analysis. RESULTS: We included 114 children (53.5% females) between 6.0 and 11.9 years old. The highest concentrations (medians, pg/µg albumin) were seen for visfatin (183.70) and adiponectin (162.26) and the lowest for IL-13 and IL-2 (~0.003). Albumin concentration was associated with age (rS = 0.39, p < 0.001). In the multivariate analysis, five analytes (C peptide, ghrelin, GLP-1, glucagon, leptin) inversely correlated with age and positively with height-for-age. Age was also positively associated with PAI-1, while height-for-age was also positively associated with insulin and visfatin. Finally, BMI-for-age had a positive correlation with GM-CSF and insulin. CONCLUSIONS: Herein, we provided concentration values for 29 analytes in saliva from healthy children that may be useful as preliminary reference framework in the clinical research setting.

A Study on the Immunohistochemical Expressions of Leptin and Leptin Receptor in Clear Cell Renal Cell Carcinoma.

BACKGROUND: The mechanisms that link obesity and cancer development are not well-defined. Investigation of leptin and leptin receptor expressions may help define some of the mechanisms. These proteins are known for associating with the immune response, angiogenesis and, signalling pathways such as JAK2/STAT3, PI3K, and AKT pathways. Tissue proteins can be easily detected with immunohistochemistry (IHC), a technique widely used both in diagnostic and research laboratories. The identification of altered levels of leptin and leptin receptor proteins in tumour tissues may lead to targeted treatment for cancer. OBJECTIVE: The objective of this study was to use IHC to compare leptin and leptin receptor expressions in clear cell renal cell carcinomas (ccRCC) in non-obese and obese patients to determine the association between these proteins with the clinicopathological features and prognosis of ccRCC. Patients and Methods. The study involved 60 patients who underwent nephrectomy of which 34 were obese, as assessed using body mass index (BMI). Nephrectomy samples provided tissues of ccRCC and adjacent non-cancerous kidney. The intensity and localization of leptin and leptin receptor protein expressions were evaluated using IHC and correlated with clinicopathological features and clinical outcomes. Aperio ImageScope morphometry and digital pathology were applied to assess the IHC results. The chi-square test was used to determine if there was any significant association between the proteins and the clinicopathological features. The Kaplan-Meier test was used to determine the overall survival, disease-free survival, and recurrence-free survival. A value of p < 0.05 was considered significant. RESULTS: There was neither significant difference in the overall cellular and nuclear expressions of leptin and leptin receptor between non-cancerous kidney and ccRCC tissues nor in non-obese and obese individuals with ccRCC. CONCLUSION: In this present study, it was revealed that leptin and leptin receptor were not associated with tumour characteristics and progression of ccRCC patients. Interestingly, nuclear expression of leptin was significantly associated with overall survival. However, the significance of these proteins as biomarkers in other RCC histotypes is still unclear.

Evaluation of Fat Accumulation and Adipokine Production during the Long-Term Adipogenic Differentiation of Porcine Intramuscular Preadipocytes and Study of the Influence of Immunobiotics.

The degree of fat accumulation and adipokine production are two major indicators of obesity that are correlated with increased adipose tissue mass and chronic inflammatory responses. Adipocytes have been considered effector cells for the inflammatory responses due to their capacity to express Toll-like receptors (TLRs). In this study, we evaluated the degree of fat accumulation and adipokine production in porcine intramuscular preadipocyte (PIP) cells maintained for in vitro differentiation over a long period without or with stimulation of either TNF-α or TLR2-, TLR3-, or TLR4-ligands. The cytosolic fat accumulation was measured by liquid chromatography and the expression of adipokines (CCL2, IL-6, IL-8 and IL-10) were quantified by RT-qPCR and ELISA at several time points (0 to 20 days) of PIP cells differentiation. Long-term adipogenic differentiation (LTAD) induced a progressive fat accumulation in the adipocytes over time. Activation of TLR3 and TLR4 resulted in an increased rate of fat accumulation into the adipocytes over the LTAD. The production of CCL2, IL-8 and IL-6 were significantly increased in unstimulated adipocytes during the LTAD, while IL-10 expression remained stable over the studied period. An increasing trend of adiponectin and leptin production was also observed during the LTAD. On the other hand, the stimulation of adipocytes with TLRs agonists or TNF-α resulted in an increasing trend of CCL2, IL-6 and IL-8 production while IL-10 remained stable in all four treatments during the LTAD. We also examined the influences of several immunoregulatory probiotic strains (immunobiotics) on the modulation of the fat accumulation and adipokine production using supernatants of immunobiotic-treated intestinal immune cells and the LTAD of PIP cells. Immunobiotics have shown a strain-specific ability to modulate the fat accumulation and adipokine production, and differentiation of adipocytes. Here, we expanded the utility and potential application of our in vitro PIP cells model by evaluating an LTAD period (20 days) in order to elucidate further insights of chronic inflammatory pathobiology of adipocytes associated with obesity as well as to explore the prospects of immunomodulatory intervention for obesity such as immunobiotics.

The energy budget and fat accumulation in striped hamsters (Cricetulus barabensis) during post-lactation.

Adaptive adjustments of energy intake and body fat play an important role in allowing animals' to meet the energy demands of thermoregulation during cold conditions and reproduction. Body fat is usually metabolized during lactation, which is one of the most energetically demanding activities of female mammals, however the effect of this on the energy budget and body fat regulation after lactation remains unclear. We compared the energy intake and body fat of female striped hamsters (Cricetulus barabensis) fed either a high-fat or low-fat diet for 21 days after the end of lactation (post-lactation, PL) to those of virgin controls. Serum leptin levels and the expression of hypothalamic orexigenic and anorexigenic neuropeptide genes were also measured and compared. Although lactating females consumed significantly more food, they had significantly lower body fat than virgin controls. The energy intake and body fat levels of the PL females were, however, significantly higher than those of virgin females. This was particularly true for the PL females that were fed high-fat diet. These females had significantly higher serum leptin concentrations, but lower hypothalamic leptin receptor gene expression, than virgin females. Neither orexigenic nor anorexigenic neuropeptide levels in the hypothalamus differed significantly between the PL and virgin females. This suggests that a negative energy balance during lactation drives fat accumulation after lactation. Furthermore, leptin resistance may occur after the end of lactation, causing females to consume more food, and accumulate more fat, than virgin females.

Association between expression of AMPK pathway and adiponectin, leptin, and vascular endothelial function in rats with coronary heart disease.

OBJECTIVE: The aim of this study was to explore the association between the expression of adenosine monophosphate-activated protein kinase (AMPK) pathway and adiponectin (APN), leptin, and vascular endothelial function in rats with coronary heart disease (CHD). MATERIALS AND METHODS: Experimental rats were divided into three groups, including: control (Col) group, CHD model (CHD) group, and CHD+AMPK activator (CHD+AICAR) group. Except those in Col group, all rats were fed with high-fat diet and intraperitoneally injected with pituitrin to establish the CHD model. The levels of serum APN, leptin, and endothelin-1 (ET-1) were determined via enzyme-linked immunosorbent assay (ELISA). The content of serum nitric oxide (NO) was detected using the nitrate reductase method. Meanwhile, the expression of AMPK pathway-related protein AMPKα in vascular endothelial tissues was detected via Western blotting (WB). Aortic vascular endothelial cells (VECs) were cultured with AICAR or ET-1 in vitro. Subsequently, the expressions of AMPK pathway and protein kinase B (AKT) pathway-related proteins were determined through co-immunoprecipitation and WB. Moreover, the expression level of NO in VECs was determined using the DAF-FM DA fluorescence probe. RESULTS: Compared with Col group, CHD group showed significantly decreased levels of serum APN and NO (p<0.05), significantly increased the levels of leptin and ET-1 (p<0.05), as well as remarkably decreased protein expression of p-AMPKα in vascular endothelial tissues (p<0.05). After injection of AMPK activator AICAR (200 mg/kg), the protein expression of p-AMPKα in CHD rats was significantly activated (p<0.05). The levels of serum APN and NO were remarkably upregulated (p<0.05), while the levels of leptin and ET-1 were significantly reduced (p<0.05). Besides, AICAR could evidently activate the activity of AMPK pathway in VECs in vitro, upregulate the protein levels of p-eNOS (Ser1177) and p-AMPKα, and promote the secretion of NO (p<0.05). In addition, AICAR remarkably inhibited ET-1-induced expression of AKT pathway (p<0.05). CONCLUSIONS: Activating the AMPK pathway may play a positive role in the normal function of VECs and exert a certain curative effect on CHD in rats.

Expression of ghrelin and leptin in the chemosensory system of adult zebrafish.

Numerous data show that the chemosensory system seems to be modulated by changes in the circulating levels of different molecules such as ghrelin, orexin, leptin, NPY, CCK. The chemosensory system of the zebrafish is represented by the taste buds (skin, oral and oropharyngeal), the olfactory rosette and the solitary chemosensorial cells (SCCs). The purpose of our study was to analyze the distribution of two peripheral hormones such as ghrelin and leptin in the chemosensory organs of the zebrafish. Our results demonstrated the presence of immunoreaction for all antibodies used in the zebrafish chemosensory organs even if with different distribution. In particular, IR was observed for ghrelin in the olfactory rosette while IR for leptin was found in the olfactory rosette, in the skin and oropharyngeal taste buds and in the gills. Both these hormones were detected in the intestine, used as a control.

CD90 serves as differential modulator of subcutaneous and visceral adipose-derived stem cells by regulating AKT activation that influences adipose tissue and metabolic homeostasis.

BACKGROUND: White adipose tissue includes subcutaneous and visceral adipose tissue (SAT and VAT) with different metabolic features. SAT protects from metabolic disorders, while VAT promotes them. The proliferative and adipogenic potentials of adipose-derived stem cells (ADSCs) are critical for maintaining adipose tissue homeostasis through driving adipocyte hyperplasia and inhibiting pathological hypertrophy. However, it remains to be elucidated the critical molecules that regulate different potentials of subcutaneous and visceral ADSCs (S-ADSCs, V-ADSCs) and mediate distinct metabolic properties of SAT and VAT. CD90 is a glycosylphosphatidylinositol-anchored protein on various cells, which is also expressed on ADSCs. However, its expression patterns and differential regulation on S-ADSCs and V-ADSCs remain unclear. METHODS: S-ADSCs and V-ADSCs were detected for CD90 expression. Proliferation, colony formation, cell cycle, mitotic clonal expansion, and adipogenic differentiation were assayed in S-ADSCs, V-ADSCs, or CD90-silenced S-ADSCs. Glucose tolerance test and adipocyte hypertrophy were examined in mice after silencing of CD90 in SAT. CD90 expression and its association with CyclinD1 and Leptin were analyzed in adipose tissue from mice and humans. Regulation of AKT by CD90 was detected using a co-transfection system. RESULTS: Compared with V-ADSCs, S-ADSCs expressed high level of CD90 and showed increases in proliferation, mitotic clonal expansion, and adipogenic differentiation, together with AKT activation and G1-S phase transition. CD90 silencing inhibited AKT activation and S phase entry, thereby curbing proliferation and mitotic clonal expansion of S-ADSCs. In vivo CD90 silencing in SAT inhibited S-ADSC proliferation, which caused adipocyte hypertrophy and glucose intolerance in mice. Furthermore, CD90 was highly expressed in SAT rather than in VAT in human and mouse, which had positive correlation with CyclinD1 but negative correlation with Leptin. CD90 promoted AKT activation through recruiting its pleckstrin homology domain to plasma membrane. CONCLUSIONS: CD90 is differentially expressed on S-ADSCs and V-ADSCs, and plays critical roles in ADSC proliferation, mitotic clonal expansion, and hemostasis of adipose tissue and metabolism. These findings identify CD90 as a crucial modulator of S-ADSCs and V-ADSCs to mediate distinct metabolic features of SAT and VAT, thus proposing CD90 as a valuable biomarker or target for evaluating ADSC potentials, monitoring or treating obesity-associated metabolic disorders.

Effects of hydrocortisone administration on leptin and adiponectin synthesis in dogs.

OBJECTIVE: To determine effects of hydrocortisone administration on serum leptin and adiponectin concentrations, abdominal fat distribution, and mRNA expression of leptin and adiponectin in abdominal adipose tissue of dogs. ANIMALS: 12 healthy dogs. PROCEDURES: Dogs received hydrocortisone (8.5 mg/kg; n = 6) or a placebo (6) orally every 12 hours for 90 days. Serum leptin and adiponectin concentrations were measured with a canine-specific ELISA on the day before (day 0; baseline) and during (days 1, 3, 7, 30, 60, and 90) administration. On days 0, 30, 60, and 90, abdominal fat mass was quantified with CT, and mRNA expression of leptin and adiponectin in abdominal fat was analyzed by use of a PCR assay. RESULTS: Hydrocortisone administration resulted in an increase in visceral fat mass on days 60 and 90, compared with the mass at baseline. Visceral fat mass at the level of L3 increased during hydrocortisone administration. Serum leptin concentration began to increase on day 1 and was significantly higher than the baseline concentration on days 30 and 60. Serum adiponectin concentration on days 30, 60, and 90 was significantly lower than the baseline concentration. Leptin and adiponectin mRNA expression in abdominal fat was greater on day 30, compared with expression at baseline, but lower on days 60 and 90, compared with expression on day 30. Serum leptin concentration and visceral fat mass were correlated. CONCLUSIONS AND CLINICAL RELEVANCE: Hydrocortisone administration affected abdominal fat distribution and serum leptin and adiponectin concentrations through dysregulation of leptin and adiponectin expression.

An immunometabolic pathomechanism for chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is an inflammatory condition associated with abnormal immune responses, leading to airflow obstruction. Lungs of COPD subjects show accumulation of proinflammatory T helper (Th) 1 and Th17 cells resembling that of autoreactive immune responses. As regulatory T (Treg) cells play a central role in the control of autoimmune responses and their generation and function are controlled by the adipocytokine leptin, we herein investigated the association among systemic leptin overproduction, reduced engagement of glycolysis in T cells, and reduced peripheral frequency of Treg cells in different COPD stages. These phenomena were also associated with an impaired capacity to generate inducible Treg (iTreg) cells from conventional T (Tconv) cells. At the molecular level, we found that leptin inhibited the expression of forkhead-boxP3 (FoxP3) and its splicing variants containing the exon 2 (FoxP3-E2) that correlated inversely with inflammation and weakened lung function during COPD progression. Our data reveal that the immunometabolic pathomechanism leading to COPD progression is characterized by leptin overproduction, a decline in the expression of FoxP3 splicing forms, and an impairment in Treg cell generation and function. These results have potential implications for better understanding the autoimmune-like nature of COPD and the pathogenic events leading to lung damage.

A structure-function approach identifies L-PGDS as a mediator responsible for glucocorticoid-induced leptin expression in adipocytes.

Leptin is an adipokine predominantly secreted by adipocytes and has many physiological roles, including in energy homeostasis. We identified that AM630, a cannabinoid receptor 2 (CB2) antagonist, down-regulated leptin expression in mature adipocytes differentiated from either stromal vascular fractions isolated from inguinal fat pads of C57BL/6J mice or 3T3-L1 preadipocytes. However, the leptin-suppressive effects of AM630 preserved in CB2-deficient adipocytes indicated the off-target activity of AM630 in leptin expression. Pharmacological and genetic studies, cheminformatics, and docking simulation were applied to identify the potential protein target of AM630 that modulates leptin expression in differentiated primary preadipocytes. Screening of the reported off-targets of AM630 identified a synthetic cannabinoid WIN55212-2 exerting the same function. Target deconvolution and docking simulation suggested that AM630 and WIN55212-2 were both inhibitors of lipocalin-type prostaglandin D2 synthase (L-PGDS). Further studies showed that L-PGDS positively regulates leptin expression. Although glucocorticoid and aldosterone were previously reported to induce expression of both L-PGDS and leptin, our data demonstrated that L-PGDS mediates only glucocorticoid-induced leptin expression in differentiated primary preadipocytes. No effect was observed after aldosterone treatment. This newly discovered glucocorticoid - L-PGDS - leptin pathway may provide insights into current clinical use of glucocorticoid and management of their undesired effects such as obesity.

Leptin produced by obesity-altered adipose stem cells promotes metastasis but not tumorigenesis of triple-negative breast cancer in orthotopic xenograft and patient-derived xenograft models.

BACKGROUND: Breast cancer is the second leading cause of cancer deaths in the USA. Triple-negative breast cancer (TNBC) is a clinically aggressive subtype of breast cancer with high rates of metastasis, tumor recurrence, and resistance to therapeutics. Obesity, defined by a high body mass index (BMI), is an established risk factor for breast cancer. Women with a high BMI have increased incidence and mortality of breast cancer; however, the mechanisms(s) by which obesity promotes tumor progression are not well understood. METHODS: In this study, obesity-altered adipose stem cells (obASCs) were used to evaluate obesity-mediated effects of TNBC. Both in vitro and in vivo analyses of TNBC cell lines were co-cultured with six pooled donors of obASCs (BMI > 30) or ASCs isolated from lean women (lnASCs) (BMI < 25). RESULTS: We found that obASCs promote a pro-metastatic phenotype by upregulating genes associated with epithelial-to-mesenchymal transition and promoting migration in vitro. We confirmed our findings using a TNBC patient-derived xenograft (PDX) model. PDX tumors grown in the presence of obASCS in SCID/beige mice had increased circulating HLA1+ human cells as well as increased numbers of CD44+CD24- cancer stem cells in the peripheral blood. Exposure of the TNBC PDX to obASCs also increased the formation of metastases. The knockdown of leptin expression in obASCs suppressed the pro-metastatic effects of obASCs. CONCLUSIONS: Leptin signaling is a potential mechanism through which obASCs promote metastasis of TNBC in both in vitro and in vivo analyses.

Differential expression of adipokines in the synovium and infrapatellar fat pad of osteoarthritis patients with and without metabolic syndrome.

Objective: To evaluate the expression levels of adipokines in the synovium and infrapatellar fat pad (IPFP) in osteoarthritis (OA) patients with and without metabolic syndrome (MetS). Methods: 120 female patients with OA were enrolled, and 60 healthy women matched body mass index, age, and sex, served as controls. Adipokines levels were measured using a sandwich enzyme-linked immunosorbent assay of the serum of all participants and synovial fluid (SF) of OA patients. Local expression levels of adipokines in the synovium and IPFP were examined by immunohistochemical analysis. The amount of adipokine proteins was analyzed using Western blot, and adipokine mRNA expressions were determined via quantitative real-time polymerase chain reaction (qRT-PCR). Results: Serum leptin levels were significantly higher in the non-MetS-OA group than those in controls (7.97 vs. 4.24 ng/ml, p< 0.001), and even higher leptin levels were found in the MetS-OA group (19.05 ng/ml; p< 0.001 for both). Serum adiponectin levels were significantly lower in the MetS-OA group than those in controls (8.09 vs. 10.07 µg/ml, respectively; p= 0.001). The synovium and IPFP in the MetS-OA group secreted more leptin and less adiponectin than those in the non-MetS-OA group (Leptin: 5.32 vs. 1.28 in synovium, respectively; p= 0.028; 6.44 vs. 0.88 in IPFP, respectively; p= 0.017. Adiponectin: 1.12 vs. 0.12 in synovium, respectively; p= 0.042; 1.07 vs. 0.09 in IPFP, respectively; p= 0.027). Resistin expression levels in the serum, SF, and articular tissues were similar among the groups. Conclusions: Expressions of adipokines were different in the synovium and IPFP of OA patients with and without MetS.

Differences in leptin biosynthesis in the stomach and in serum leptin level between men and women.

BACKGROUND AND AIM: Although the stomach has been identified as an important source of leptin, the detailed biosynthesis sites of leptin in human gastrointestinal tract have not been fully elucidated. The study objective was to compare leptin distribution and expression in the stomach and its serum level between healthy men and women. METHODS: Nineteen subjects (organ donors; 10 men and 9 women) with normal gastric mucosa histology were recruited. Research material contained gastric samples from the cardia, fundus, and pyloric regions. Gastric mucosa leptin content and leptin gene expression were determined by immunohistochemistry and real-time polymerase chain reaction method. Plasma leptin level was measured using ELISA method. RESULTS: In the stomach of healthy adult subjects, leptin-immunoreactive cells were mainly found in the fundus, and the number of immunoreactive cells was higher in women than in men. Leptin-containing cells were less numerous in the cardia and pylorus mucosa. Similarly, leptin gene expression was the highest in the fundus and higher in women than in men. Serum leptin level was higher in women than in men and was found to correlate positively with body mass index and weight in both sexes. A negative correlation between leptin level and age was noted in women, but not in men. CONCLUSIONS: The current study is the first to provide evidence for the presence of leptin-containing cells in all segments of the human stomach. The differences in gastric leptin biosynthesis and serum leptin levels between men and women suggest that leptin secretion can be controlled by sex hormones or other unknown factors.

The Effect of the Leptin and Leptin Receptor Expression on the Efficacy of Neoadjuvant Chemotherapy in Breast Cancer.

BACKGROUND The purpose of the present study was to evaluate the effect of leptin and leptin receptor (LEPR) expression on the efficacy of neoadjuvant chemotherapy in breast cancer. MATERIAL AND METHODS There were 325 breast cancer patients with complete data enrolled in this study. Patients were categorized into 3 groups: pathological complete response group, non-pathological complete response group, and progressive disease group. Immunohistochemistry was performed to determine leptin and its receptor LEPR expression levels that were compared among the 3 groups. RESULTS Compared with the non-pathological complete response group, patients in the pathological complete response group had increased leptin and LEPR expression, although the difference was not statistically significant (P=0.194, P=0.110). In addition, the expression of leptin and LEPR in the pathological complete response group was also higher than that in the progressive disease group, and the difference of LEPR expression was statistically significant (P=0.008) while the leptin expression was not (P=0.065). There were more HER2+ breast cancer patients in the pathological complete response group categorized into strong positive, and positive expression of leptin and LEPR compared with the progressive disease group (P<0.05). There were significant differences of leptin and LEPR expression among breast cancer patients under different molecular subtypes HER2+, HR+, and triple negative, in which the triple negative patients had the highest expression of leptin and LEPR. In addition, patients in the progressive disease group had high and low expression of leptin and LEPR: 13.25% versus 11.32% and 13.1% versus 10.42% respectively. CONCLUSIONS Overexpression of leptin and LEPR improved the therapeutic efficacy of neoadjuvant chemotherapy for patients with breast cancer, especially for those with HER2+ subtype. Overexpression of leptin and LEPR was distinct among the different molecular subtypes of breast cancer, suggesting a certain predictive value for breast cancer prognosis.

Hydrogen Sulfide Inhibits Formaldehyde-Induced Senescence in HT-22 Cells via Upregulation of Leptin Signaling.

It has been previously demonstrated that hydrogen sulfide (H2S) prevents formaldehyde (FA)-induced neurotoxicity. However, the exact mechanisms underlying this protection remain to be fully elucidated. Neuronal senescence is involved in FA-induced neurotoxicity. Leptin signaling has anti-aging function. The present work was to investigate the protection of H2S against FA-induced neuronal senescence and the mediatory role of leptin signaling. FA-exposed HT-22 cells were used as the vitro model of FA-induced neuronal senescence. The senescence-associated ß-galactosidase (SA-ß-Gal) positive cell was detected by ß-galactosidase staining. The expressions of P16INK4a, P21CIP1, leptin, and lepRb (leptin receptor) were measured by western blot. The proliferation, viability, and apoptosis of cells were evaluated by Trypan blue exclusion assay, Cell Counting Kit-8 (CCK-8) assay, and Flow cytometry analysis, respectively. We found that H2S suppressed FA-induced senescence, as evidenced by the decrease in SA-ß-Gal positive cells, the downregulations of P16INK4a and P21CIP1, as well as decrease in cell growth arrest, in HT-22 cells. Also, H2S upregulated the expressions of leptin and lepRb in FA-exposed HT-22 cells. Furthermore, leptin tA (a specific inhibitor of the leptin) abolished the protective effects of H2S on FA-induced senescence and neurotoxicity (as evidenced by the increase in cell viability and the decrease in cell apoptosis) in HT-22 cells. These results indicated that H2S prevents FA-induced neuronal senescence via upregulation of leptin signaling. Our findings offer a novel insight into the mechanisms underlying the protection of H2S against FA-induced neurotoxicity. FA upregulates the expressions of P16INK4a and P21CIP1 via inhibiting leptin signaling, which in turn induces senescence in HT-22 cells; H2S downregulates the expressions of P16INK4a and P21CIP1 via reversing FA-downregulated leptin signaling, which in turn prevents FA-induced senescence in HT-22 cells.

Egr1 mediates the effect of insulin on leptin transcription in adipocytes.

In mammals, leptin production in adipocytes is up-regulated by feeding and insulin. Although this regulatory connection is central to all physiological effects of leptin, its molecular mechanism remains unknown. Here, we show that the transcription factor early growth response 1, Egr1, is rapidly but transiently induced by insulin in adipose cells both in vitro and in vivo, and its induction is followed by an increase in leptin transcription. ChIP and luciferase assays demonstrate that Egr1 directly binds to and activates the leptin promoter. Interestingly, the lipid droplet protein FSP27 may work as a co-factor for Egr1 in regulating leptin expression. By using siRNA-mediated knockout of Egr1 along with its overexpression in adipocytes, we demonstrate that Egr1 is both necessary and sufficient for the stimulatory effect of insulin on leptin transcription.

Iron down-regulates leptin by suppressing protein O-GlcNAc modification in adipocytes, resulting in decreased levels of O-glycosylated CREB.

We previously reported that iron down-regulates transcription of the leptin gene by increasing occupancy of phosphorylated cAMP response element-binding protein (pCREB) at two sites in the leptin gene promoter. Several nutrient-sensing pathways including O-GlcNAcylation also regulate leptin. We therefore investigated whether O-glycosylation plays a role in iron- and CREB-mediated regulation of leptin. We found that high iron decreases protein O-GlcNAcylation both in cultured 3T3-L1 adipocytes and in mice fed high-iron diets and down-regulates leptin mRNA and protein levels. Glucosamine treatment, which bypasses the rate-limiting step in the synthesis of substrate for glycosylation, increased both O-GlcNAc and leptin, whereas inhibition of O-glycosyltransferase (OGT) decreased O-GlcNAc and leptin. The increased leptin levels induced by glucosamine were susceptible to the inhibition by iron, but in the case of OGT inhibition, iron did not further decrease leptin. Mice with deletion of the O-GlcNAcase gene, either via whole-body heterozygous deletion or through adipocyte-targeted homozygous deletion, exhibited increased O-GlcNAc levels in adipose tissue and increased leptin levels that were inhibited by iron. Of note, iron increased the occupancy of pCREB and decreased the occupancy of O-GlcNAcylated CREB on the leptin promoter. These patterns observed in our experimental models suggest that iron exerts its effects on leptin by decreasing O-glycosylation and not by increasing protein deglycosylation and that neither O-GlcNAcase nor OGT mRNA and protein levels are affected by iron. We conclude that iron down-regulates leptin by decreasing CREB glycosylation, resulting in increased CREB phosphorylation and leptin promoter occupancy by pCREB.

The correlation between oxidative stress level and intra-abdominal fat in obese males.

This study aims to investigate the correlation between oxidative stress and intra-abdominal fat (IAF) in obese young and middle-aged males.The present study included 136 male examinees in the Examination Center of the First Hospital of Qinhuangdao from October 10, 2015 to December 10, 2015. Then, clinical data, oxidative stress indices (8-iso-prostaglandin F2α [8-iso-PGF2α], malondialdehyde [MDA], and superoxide dismutase [SOD]), and IAF area were recorded. All subjects were assigned into 3 groups according to body mass index (BMI): obese group (BMI ≥ 28 kg/m, 43 subjects), overweight group (24 ≤ BMI < 28 kg/m, 46 subjects), and control group (BMI < 24 kg/m, 47 subjects). Then, statistical analysis was performed.There were significant differences in IAF area, leptin, adiponectin, 8-iso-PGF2α, MDA, SOD, fasting insulin (FINS), fasting blood glucose (FBG), and homeostasis model assessment-insulin resistance (HOMA-IR) among these 3 groups (P < .05). Male subjects in the obese group had higher leptin, 8-iso-PGF2α, MDA, FINS, and HOMA-IR levels, compared to subjects in the overweight and control groups. Furthermore, subjects in the overweight group had a larger IAF area and higher 8-iso-PGF2α, MDA, and FBG levels, when compared to controls. In addition, SOD was significantly lower in the obese and overweight groups than in the control group. However, there were no statistical differences in age, systolic and diastolic blood pressure, lipids, and islet ß-cell secretion function (homeostasis model assessment-ß) among these 3 groups (P ≥ .05). Moreover, the IAF area was positively correlated to 8-iso-PGF2α and MDA, and negatively correlated to SOD.Oxidative stress is significantly associated with the IAF area in obese males, and abdominal obesity could increase oxidative stress level and insulin resistance.

Expression of Three Adipokines (Adiponectin, Leptin and Resistin) in Normal Canine Skin: a Pilot Study.

Adipokines are biologically active cytokines that are mainly produced in adipose tissue. There is evidence, in man and mice, that some adipokines may be secreted in other tissues including the vascular endothelium, epithelia and sebaceous glands. Moreover, modified serum levels of adipokines have been detected in people with acne vulgaris or psoriasis; it is suspected that adipokines could contribute to local and systemic inflammatory conditions. We aimed to evaluate the expression of three adipokines (i.e. leptin, adiponectin and resistin) in normal canine skin. Formalin-fixed, paraffin wax-embedded punch biopsy samples were obtained from the sparsely-haired skin of the caudal ventral abdomen of a single clinically healthy dog with no history of skin disease. Immunohistochemistry was applied, using rabbit polyclonal primary antibodies specific for leptin, adiponectin and resistin. Adipokines were not expressed in normal canine dermis or hypodermis. In contrast, they were detected in the keratinocytes of all epidermal layers and hair follicle segments, sebocytes, apocrine gland cells and in the vascular endothelium. This is the first report on the expression of adipokines in normal canine skin, a first step in studying their role in the skin physiology and inflammatory skin diseases of dogs.