MiR-125b-2 knockout increases high-fat diet-induced fat accumulation and insulin resistance
ABSTRACT: Obese individuals are more susceptible to comorbidities than individuals of healthy weight, including cardiovascular disease and metabolic disorders. MicroRNAs are a class of small and noncoding RNAs that are implicated in the regulation of chronic human diseases. We previously reported that miR-125b plays a critical role in adipogenesis in vitro. However, the involvement of miR-125b-2 in fat metabolism in vivo remains unknown. In the present study, miR-125b-2 knockout mice were generated using CRISPR/CAS9 technology, resulting in mice with a 7 bp deletion in the seed sequence of miR-125b-2. MiR-125b-2 knockout increased the weight of liver tissue, epididymal white fat and inguinal white fat. MiR-125b-2 knockout also increased adipocyte volume in HFD-induced obese mice, while there were no significant differences in body weight and feed intake versus mice fed a normal diet. Additionally, qRT-PCR and western blot analysis revealed that the expression of the miR-125b-2 target gene SCD-1 and fat synthesis-associated genes, such as PPAR? and C/EBP?, were significantly up-regulated in miR-125b-2KO mice (P?
Project description:<b>Objective:</b> Long-term dysregulation of energy balance is the key component of the obesity epidemic. Given the harm of central obesity and the discovery that beige cells appear within white adipose tissue (WAT), enhancing the energy-expending or "browning" ability of visceral adipose tissue (VAT) has become of therapeutic interest. In this study, we focused on the regulating role of microRNA (miRNA)-27b-3p in mice epididymal white adipose tissue (eWAT) browning. <b>Methods:</b> High-fat diet (HFD) induced obese mice model was constructed. Expression of miR-27b-3p and Ucp1 in eWAT was measured during the course of HFD. Through tail vein injection of antimiR-27b-3p, miR-27b-3p expression was inhibited to analyze the potential role of miR-27b-3p in fat browning and metabolism. <b>Results:</b> miR-27b-3p was predominantly expressed in eWAT and browning ability of eWAT in HFD induced obese mice was impaired. Inhibition of miR-27b-3p enhanced browning capacity of eWAT in mice fed an HFD and led to weight loss and insulin sensitivity improvement. <b>Conclusions:</b> High expression of miR-27b-3p in eWAT inhibits browning ability and leads to visceral fat accumulation. It is suggested miR-27b-3p may become a potential therapeutic option for visceral obesity and its associated diseases.
Project description:Background: This study aimed at assessing the effect of a low-fat diet (LFD) in obese mice lacking toll?like receptors (Tlr) and understanding the expression and regulation of microRNAs during weight reduction. Methods: C57BL/6, Tlr5-/-, Tlr2-/- and Tlr4-/- mice were used in this study. A group of mice were fed with a high-fat diet (HFD) (58% kcal) for 12 weeks to induce obesity (diet-induced obesity, DIO). Another group that had been fed with HFD for eight weeks (obese mice) were switched to a low-fat diet (LFD) (10.5% kcal) for the next four weeks to reduce their body weight. The control mice were fed with a standard AIN-76A diet for the entire 12 weeks. The body weight of the mice was measured weekly. At the end of the experiment, epididymal fat weight and adipocyte size were measured. The differentially expressed miRNAs in the fat tissue was determined by next-generation sequencing with real-time quantitative reverse transcription polymerase chain reaction (RT?qPCR). Target prediction and functional annotation of miRNAs were performed using miRSystem database. Results: Switching to LFD significantly reduced the body weight and epididymal fat mass in the HFD-fed C57BL/6 and Tlr5-/- mice but not in Tlr2-/- and Tlr4-/- mice. Weight reduction significantly decreased the size of adipocytes in C57BL/6 but not in the Tlr knockout mice. In Tlr2-/- and Tlr4-/- mice, feeding with HFD and the subsequent weight reduction resulted in an aberrant miRNA expression in the epididymal fat tissue unlike in C57BL/6 and Tlr5-/-. However, target prediction and functional annotation by miRSystem database revealed that all the top 10 Kyoto Encyclopedia of Genes and Genomes (KEGG) database pathways of the dysregulated miRNAs during weight reduction in the C57BL/6 mice were also found in the regulated pathways of Tlr5-/-, Tlr2-/-, and Tlr4-/- strains. However, among these pathways, gene sets involved in arginine and proline metabolism and glutathione metabolism were mainly involved in the Tlr knockout mice but not in the C57BL/6 mice. Conclusions: In this study, we demonstrated that feeding of LFD leads to significant body weight reduction in C57BL/6 and Tlr5-/- mice, but not in Tlr2-/- and Tlr4-/- mice. Significant reduction in the size of adipocytes of epididymal fat was only found in C57BL/6, but not in Tlr5-/-, Tlr2-/-, and Tlr4-/- mice. The dysregulated miRNAs in Tlr2-/- and Tlr4-/- mice were different from those in C57BL/6 and Tlr5-/- strains. Among those miRNA-regulated pathways, arginine and proline metabolism as well as glutathione metabolism may have important roles in the Tlr knockout mice rather than in C57BL/6 mice.
Project description:The synthetic retinoid Fenretinide (FEN) increases insulin sensitivity in obese rodents and is in early clinical trials for treatment of insulin resistance in obese humans with hepatic steatosis (46). We aimed to determine the physiological mechanisms for the insulin-sensitizing effects of FEN. Wild-type mice were fed a high-fat diet (HFD) with or without FEN from 4-5 wk to 36-37 wk of age (preventive study) or following 22 wk of HF diet-induced obesity (12 wk intervention study). Retinol-binding protein-4 (RBP4) knockout mice were also fed the HFD with or without FEN in a preventive study. FEN had minimal effects on HFD-induced body weight gain but markedly reduced HFD-induced adiposity and hyperleptinemia in both studies. FEN-HFD mice gained epididymal fat but not subcutaneous or visceral fat mass in contrast to HFD mice without FEN. FEN did not have a measurable effect on energy expenditure, food intake, physical activity, or stool lipid content. Glucose infusion rate during hyperinsulinemic-euglycemic clamp was reduced 86% in HFD mice compared with controls and was improved 3.6-fold in FEN-HFD compared with HFD mice. FEN improved insulin action on glucose uptake and glycogen levels in muscle, insulin-stimulated suppression of hepatic glucose production, and suppression of serum FFA levels in HFD mice. Remarkably, FEN also reduced hepatic steatosis. In RBP4 knockout mice, FEN reduced the HFD-induced increase in adiposity and hyperleptinemia. In conclusion, long-term therapy with FEN partially prevents or reverses obesity, insulin resistance, and hepatic steatosis in mice on HFD. The anti-adiposity effects are independent of the RBP4 lowering effect.
Project description:Obesity is a growing epidemic in developed countries. Obese individuals are susceptible to comorbidities, including cardiovascular disease and metabolic disorder. Increasing the ability of adipose tissue to expend excess energy could improve protection from obesity. One promising target is microRNA (miR)-155-5p. We demonstrate that deletion of miR-155 (-5p and -3p) in female mice prevents diet-induced obesity. Body weight gain did not differ between wild-type (WT) and miR-155 knockout (KO) mice fed control diet (CD); however, miR-155 KO mice fed high-fat diet (HFD) gained 56% less body weight and 74% less gonadal white adipose tissue (WAT) than WT mice. Enhanced WAT thermogenic potential, brown adipose tissue differentiation, and/or insulin sensitivity might underlie this obesity resistance. Indeed, miR-155 KO mice on HFD had 21% higher heat release than WT HFD mice. Compared to WT adipocytes, miR-155 KO adipocytes upregulated brown (Ucp1, Cidea, Pparg) and white (Fabp4, Pnpla2, AdipoQ, Fasn) adipogenic genes, and glucose metabolism genes (Glut4, Irs1). miR-155 deletion abrogated HFD-induced adipocyte hypertrophy and WAT inflammation. Therefore, miR-155 deletion increases adipogenic, insulin sensitivity, and energy uncoupling machinery, while limiting inflammation in WAT, which together could restrict HFD-induced fat accumulation. Our results identify miR-155 as a novel candidate target for improving obesity resistance.
Project description:Obesity contributes to metabolic disorders such as diabetes and cardiovascular disease. Characterization of differences between the main adipose tissue depots, white (WAT) [including subcutaneous (SAT) and visceral adipose tissue (VAT)] and brown adipose tissue (BAT) helps to identify their roles in obesity. Thus, we studied depot-specific differences in whole transcriptome and miRNA profiles of SAT, VAT and BAT from high fat diet (HFD/45% of calories from fat) fed mice using RNA sequencing and small RNA-Seq. Using quantitative real-time polymerase chain reaction, we validated depot-specific differences in endoplasmic reticulum (ER) stress related genes and miRNAs using mice fed a HFD vs. low fat diet (LFD/10% of calories from fat). According to the transcriptomic analysis, lipogenesis, adipogenesis, inflammation, endoplasmic reticulum (ER) stress and unfolded protein response (UPR) were higher in VAT compared to BAT, whereas energy expenditure, fatty acid oxidation and oxidative phosphorylation were higher in BAT than in VAT of the HFD fed mice. In contrast to BAT, ER stress marker genes were significantly upregulated in VAT of HFD fed mice than the LFD fed mice. For the first time, we report depot specific differences in ER stress related miRNAs including; downregulation of miR-125b-5p, upregulation miR-143-3p, and miR-222-3p in VAT following HFD and upregulation of miR-30c-2-3p only in BAT following a HFD in mice than the LFD mice. In conclusion, HFD differentially regulates miRNAs and genes in different adipose depots with significant induction of genes related to lipogenesis, adipogenesis, inflammation, ER stress, and UPR in WAT compared to BAT.
Project description:Obesity is characterized by chronic inflammation and immune dysregulation, as well as insulin resistance, but the link between obesity and adaptive immunity remains to be fully studied.To elucidate the role of adaptive immunity on body composition, glucose homeostasis and inflammation, recombination-activating gene 1 knockout (Rag1-/-) mice, without mature T-lymphocytes or B-lymphocytes, were maintained on a low- or high-fat diet (LFD and HFD, respectively) for 11 weeks.Rag1-/- mice fed HFD gained significantly more weight and had increased body fat compared with wild type. Downregulation of energy expenditure as well as brown fat uncoupling protein UCP-1 and UCP-3 gene expression were noticed in HFD-fed Rag1-/- mice compared with LFD. HFD mice had significantly decreased energy intake compared with LFD mice, consistent with decreased agouti-related protein and increased pro-opiomelanocortin gene expression levels in the hypothalamus. Moreover, compared with wild type, Rag1-/- mice had lower interleukin (IL)-4 levels, a cytokine recently found to induce browning in white adipocytes, and higher IL-12 levels in HFD-fed Rag1-/- mice. Despite that HFD Rag1-/- mice were more obese, they had similar glucose, insulin and adiponectin levels, while leptin was marginally increased.Mice with deficiency in adaptive immunity are obese, partly owing to decreased energy expenditure, but are metabolically normal, suggesting that mature lymphocytes have necessary roles in the development of obesity-related metabolic dysregulation.
Project description:We studied the effects of weight loss induced by either a low-fat normal diet (ND) or restriction of high-fat diet (HFD) on hepatic steatosis, inflammation in the liver and adipose tissue (AT), and blood monocytes of obese mice.In mice with HFD-induced obesity, weight loss was achieved by switching from HFD to ND and maintaining on ND ad libitum or by restricting HFD intake to match body weight of mice with ND-induced weight loss. After diet interventions for 4 weeks, hepatic steatosis, hepatic and AT inflammation, and blood CD11c(+) monocytes were examined.At 4 weeks after switching diets, body weight was reduced by 23% from baseline. To achieve the same reduced body weight required restricting calorie intake from HFD. Weight loss with either ND or HFD restriction decreased body fat mass and ameliorated liver steatosis; both effects were greater with ND-induced weight loss than HFD restriction-induced weight loss. Weight loss with ND but not HFD restriction normalized blood CD11c(+) monocytes and attenuated hepatic inflammation assessed by chemokine and CD11c expression. In contrast, weight loss with HFD restriction significantly reduced chemokine levels and CD11c(+) cells in AT compared to obese controls, and tended to reduce AT chemokines and CD11c(+) cells more than ND-induced weight loss.In mice with diet-induced obesity, weight loss with ND was superior in alleviating hepatic inflammation and steatosis, whereas weight loss with HFD calorie restriction provided greater amelioration of AT inflammation.
Project description:OBJECTIVES:Safflower yellow (SY) is the main effective ingredient of Carthamus tinctorius L. It has been reported that SY plays an important role in anti-inflammation, anti-platelet aggregation, and inhibiting thrombus formation. In present study, we try to investigate the effects of SY on body weight, body fat mass, insulin sensitivity in high fat diet (HFD)-induced obese mice. METHODS:HFD-induced obese male ICR mice were intraperitoneally injected with SY (120 mg kg(-1)) daily. Eight weeks later, intraperitoneal insulin tolerance test (IPITT), and intraperitoneal glucose tolerance test (IPGTT) were performed, and body weight, body fat mass, serum insulin levels were measured. The expression of glucose and lipid metabolic related genes in white adipose tissue (WAT) were determined by RT-qPCR and western blot technologies. RESULTS:The administration obese mice with SY significantly reduced the body fat mass of HFD-induced obese mice (P < 0.05). IPITT test showed that the insulin sensitivity of SY treated obese mice were evidently improved. The mRNA levels of insulin signaling pathway related genes including insulin receptor substrate 1(IRS1), PKB protein kinase (AKT), glycogen synthase kinase 3? (GSK3?) and forkhead box protein O1(FOXO1) in mesenteric WAT of SY treated mice were significantly increased to 1.9- , 2.8- , 3.3- , and 5.9-folds of that in HFD-induced control obese mice, respectively (P < 0.05). The protein levels of AKT and GSK3? were also significantly increased to 3.0 and 5.2-folds of that in HFD-induced control obese mice, respectively (P < 0.05). Meanwhile, both the mRNA and protein levels of peroxisome proliferator-activated receptorgamma coactivator 1? (PGC1?) in inguinal subcutaneous WAT of SY group were notably increased to 2.5 and 3.0-folds of that in HFD-induced control obese mice (P < 0.05). CONCLUSIONS:SY significantly reduce the body fat mass, fasting blood glucose and increase insulin sensitivity of HFD-induced obese mice. The possible mechanism is to promote the browning of subcutaneous WAT and activate the IRS1/AKT/GSK3? pathway in visceral WAT. Our study provides an important experimental evidence for developing SY as a potential anti-obesity and anti-diabetic drug.
Project description:Growing evidence demonstrates the important role of microRNAs (miRs) in regulating adipogenesis, obesity and insulin resistance. The miR-200b/a/429 cluster has been functionally characterized in mammalian reproduction; however, the potential role of the miR-200 family in adipocytes is poorly understood. The aim of our study was to investigate the physiological function of miR-200b/a/429 in the regulation of whole-body metabolism in terms of the activities and targets of this cluster in adipocytes. We generated adipocyte-specific miR-200b/a/429 knockout (ASKO) mice using a Cre-loxP system in which Cre expression was driven by the aP2 promoter. The ASKO and wild type (WT) littermate were fed a chow diet (CD) or high-fat-diet (HFD), and changes in body composition, metabolic parameters, energy homeostasis, glucose tolerance and insulin sensitivity were analyzed. The miR-200b/a/429 putative target genes were predicted and validated via luciferase reporter assays. We found that the HFD-fed ASKO mice gradually gained more body weight than the WT mice due to the increased adiposity. Decreased glucose tolerance and insulin sensitivity were also observed in the HFD-fed ASKO mice. Notably, the down-regulation of lipolysis-related genes and the decreased response to CL-316,243 stimulation in the HFD-fed ASKO mice suggested that these animals exhibited impaired lipolysis. In addition, the HFD-fed ASKO mice displayed impaired energy expenditure, indicating that the miR-200b/a/429 cluster is essential for developing adaptive responses to stressors such as HFD. For the first time, our studies demonstrated the essential role of miR-200b/a/429 in adipocytes in the regulation of HFD-induced whole-body metabolic changes.
Project description:Morbidly obese patients who accomplish substantial weight loss often display a long-term decline in their resting metabolism, causing even relatively restrained caloric intake to trigger a relapse to the obese state. Paradoxically, we observed that morbidly obese mice receiving chemotherapy for cancer experienced spontaneous weight reduction despite unabated ingestion of their high fat diet (HFD). This response to chemotherapy could also be achieved in morbidly obese mice without cancer. Optimally dosed methotrexate (MTX) or cyclophosphamide (CY) enabled the mice to completely and safely normalize their body weight despite continued consumption of obesogenic quantities of HFD. Weight reduction was not attributable to decreased HFD intake, enhanced energy expenditure or malabsorption. MTX or CY dosing significantly depleted both adipose tissue and preadipocyte progenitors. Remarkably, however, despite continued high fat feeding, a compensatory increase in hepatocyte lipid storage was not observed, but rather the opposite. Gene microarray liver analyses demonstrated that HFD mice receiving MTX or CY experienced significantly inhibited lipogenesis and lipid storage, whereas Enho (energy homeostasis) gene expression was significantly upregulated. Further metabolic studies employing a human hepatocellular line revealed that MTX treatment preserved robust oxidative phosphorylation, but also promoted mitochondrial uncoupling with a surge in proton leak. This is the first report that certain optimally dosed chemotherapeutic agents can induce weight loss in morbidly obese mice without reduced dietary intake, apparently by depleting stores of adipocytes and their progenitors, curtailment of lipogenesis, and inconspicuous disposal of incoming dietary lipid via a steady state partial uncoupling of mitochondrial oxidative phosphorylation.