Project description:Leptin is a pleiotropic hormone that is released by adipocytes and tightly controls energy balance by regulating a range of physiological processes. While leptin is best known for signaling through the hypothalamic arcuate nucleus to regulate feeding, it also supports the immune system by binding its cognate receptor Ob-R to activate various immunocyte populations. Indeed, patients with congenital leptin or leptin receptor deficiency not only display severe obesity but also exhibit increased mortality from infections. Leptin supplementation to leptin deficient mice and humans increases CD4+ cell numbers and augments proinflammatory Th1 responses ex vivo. However, whether this leptin-induced immune response is necessary for viral clearance in vivo is unknown. To better delineate the role of leptin in infection, we inoculated leptin deficient ob/ob mice with influenza and acutely supplemented leptin in physiologically relevant doses prior to infection. Although ob/ob mice and diet-induced obese (DIO) mice have comparable levels of insulin resistance and adiposity, a sublethal dose of influenza virus in DIO mice causes mortality in approximately 80% of ob/ob mice, which was completely reversed by leptin supplementation. Serum cytokine profiling and whole tissue RNA sequencing of spleen, lung, and visceral adipose tissue demonstrated increased inflammatory and anti-viral responses in leptin-supplemented mice and no signs of excessive immunopathology in either group. Further flow cytometric analysis indicated an elevated Th1 immune response locally and systemically in leptin-supplemented mice, along with a two-fold increase in spleen sizes and an expansion in the proportion of splenic B cells. Despite augmented anti-viral responses in leptin-supplemented mice, leptin deficient ob/ob mice were equally capable of clearing the virus. Moreover, pulse oximetry showed diminished oxygen saturation in both groups, a finding further corroborated by similar levels of histopathologic lung damage. Interestingly, leptin-supplementation in ob/ob mice prevented profound hypothermia and bradycardia, which occurred prior to death in control ob/ob mice. Taken together, we provide evidence that leptin is dispensable for immunologic control of viral infection and may instead confer a survival benefit by maintaining autonomic stability. These findings open new avenues for research on the role of leptin signaling in infection.

Project description:Caloric Restriction in Leptin Deficiency Worsens Myocardial Steatosis: Failure to Upregulate PPAR gamma and Thermogenic Glyecrolipid/Fatty Acid Cycling Growing evidence supports an anti-lipotoxic role for leptin in preventing inappropriate peripheral tissue lipid deposition. Obese, leptin deficient ob/ob mice develop left ventricular (LV) hypertrophy and myocardial steatosis with increased apoptosis and decreased longevity. Here we investigated the cardiac effects of caloric restriction in leptin deficiency. Echocardiography was performed on C57Bl/6 wild-type mice (WT) and 7-month-old ob/ob mice fed ad lib, leptin-repleted (LR-ob/ob), or calorie-restricted (CR-ob/ob) for four weeks. Ventricular tissue was examined by electron microscopy (EM), mitochondrial coupling assay, and microarray expression profiling. LR and CR-ob/ob mice showed decreased body weight, heart weight, and LV wall thickness compared to ad lib ob/ob mice. LV fractional shortening was decreased in ad lib ob/ob mice, but restored to WT levels in LR and CR groups. However, EM revealed severe cardiac steatosis in the CR-ob/ob group compared to only moderate steatosis in ad lib ob/ob . Despite marked cardiac steatosis, CR (like LR) restored mitochondrial coupling to WT levels. CR up-regulated genes associated with oxidative stress and cell death, changes suggestive of cardiac lipotoxicity. LR, but not CR was shown to induce core genes involved in glycerolipid/free fatty acid cycling, a highly thermogenic pathway that can reduce intracellular lipid stores. LR, but not CR up-regulated and restored PGC1 and PPARto wild type levels; CR paradoxically further suppressed cardiac PPAR. Thus, leptin is essential in protecting the heart from lipotoxicity, and the inability to up-regulate the thermogenic glycerolipid/free fatty acid cycling pathway may impair the response of leptin deficient animals to the lipotoxic stress of calorie restriction. 6 month aged ob/ob mice were either leptin repleted with osmotic mini-pumps, calorie restricted to match the caloric intake of the leptin repleted mice, or fed ad lib for one month. 6-8 month C57Bl/6J mice were aged to serve as controls.

Project description:Metabolic perturbations resulting from excessive hepatic fat accumulation are poorly understood. Thus, in this study, leptin-deficient ob/ob mice, a mouse model of fatty liver disease, were used to investigate metabolic alterations in more detail. Metabolites were quantified in intact liver tissues of ob/ob (n=8) and control (n=8) mice using high-resolution magic angle spinning (HR-MAS) 1H-NMR. In addition, after demonstrating that HR-MAS 1H-NMR does not affect RNA integrity, transcriptional changes were measured by quantitative real-time PCR on RNA extracted from the same specimens after HR-MAS 1H-NMR measurements. Importantly, the gene expression changes obtained agreed with those observed by Affymetrix microarray analysis performed on RNA isolated directly from fresh-frozen tissue. In total, 40 metabolites could be assigned in the spectra and subsequently quantified. Quantification of lactate was also possible after applying a lactate-editing pulse sequence that suppresses the lipid signal, which superimposes the lactate methyl resonance at 1.3 ppm. Significant differences were detected for creatinine, glutamate, glycine, glycolate, trimethylamine-N-oxide, dimethylglycine, ADP, AMP, betaine, phenylalanine, and uridine. Furthermore, alterations in one-carbon metabolism, supported by both metabolic and transcriptional changes, were observed. These included reduced demethylation of betaine to dimethylglycine and the reduced expression of genes coding for transsulfuration pathway enzymes, which appears to preserve methionine levels, but may limit glutathione synthesis. Overall, the combined approach is advantageous as it identifies changes not only at the single gene or metabolite level but also deregulated pathways, thus providing critical insight into changes accompanying fatty liver disease. Graphical abstract A Evaluation of RNA integrity before and after HR-MAS 1H-NMR of intact mouse liver tissue. B Metabolite concentrations and gene expression levels assessed in ob/ob (steatotic) and ob/+ (control) mice using HR-MAS 1H-NMR and qRT-PCR, respectively.

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