HSPA12A is required for adipocyte differentiation and diet-induced obesity through a positive feedback regulation with PPAR?.
ABSTRACT: Obesity is one of the most serious public health problems. Peroxisome proliferator-activated receptor ? (PPAR?) plays the master role in adipocyte differentiation for obesity development. However, optimum anti-obesity drug has yet been developed, mandating more investigation to identify novel regulator in obesity pathogenesis. Heat shock protein 12A (HSPA12A) encodes a novel member of the HSP70 family. Here, we report that obese patients showed increased adipose HSPA12A expression, which was positively correlated with increase of body mass index. Intriguingly, knockout of HSPA12A (Hspa12a-/-) in mice attenuated high-fat diet (HFD)-induced weight gain, adiposity, hyperlipidemia, and hyperglycemia compared to their wild type (WT) littermates. Increased insulin sensitivity was observed in Hspa12a-/- mice compared to WT mice. The HFD-induced upregulation of PPAR? and its target adipogenic genes in white adipose tissues (WAT) of Hspa12a-/- mice were also attenuated. Loss- and gain-of-function studies revealed that the differentiation of primary adipocyte precursors, as well as the expression of PPAR? and target adipogenic genes during the differentiation, was suppressed by HSPA12A deficiency whereas promoted by HSPA12A overexpression. Importantly, PPAR? inhibition by GW9662 reversed the HSPA12A-mediated adipocyte differentiation. On the other hand, HSPA12A expression was downregulated by PPAR? inhibition but upregulated by PPAR? activation in primary adipocytes. A direct binding of PPAR? to the PPAR response element in the Hspa12a promoter region was confirmed by chromatin immunoprecipitation assay, and this binding was increased after differentiation of primary adipocytes. These findings indicate that HSPA12A is a novel regulator of adipocyte differentiation and diet-induced obesity through a positive feedback regulation with PPAR?. HSPA12A inhibition might represent a viable strategy for the management of obesity in humans.
Project description:OBJECTIVE:Obesity due to overnutrition causes adipose tissue dysfunction, which is a critical pathological step on the road to type 2 diabetes (T2D) and other metabolic disorders. In this study, we conducted an unbiased investigation into the fundamental molecular mechanisms by which adipocytes transition to an unhealthy state during obesity. METHODS:We used nuclear tagging and translating ribosome affinity purification (NuTRAP) reporter mice crossed with Adipoq-Cre mice to determine adipocyte-specific 1) transcriptional profiles (RNA-seq), 2) promoter and enhancer activity (H3K27ac ChIP-seq), 3) and PPAR? cistrome (ChIP-seq) profiles in mice fed chow or a high-fat diet (HFD) for 10 weeks. We also assessed the impact of the PPAR? agonist rosiglitazone (Rosi) on gene expression and cellular state of adipocytes from the HFD-fed mice. We integrated these data to determine the transcription factors underlying adipocyte responses to HFD and conducted functional studies using shRNA-mediated loss-of-function approaches in 3T3-L1 adipocytes. RESULTS:Adipocytes from the HFD-fed mice exhibited reduced expression of adipocyte markers and metabolic genes and enhanced expression of myofibroblast marker genes involved in cytoskeletal organization, accompanied by the formation of actin filament structures within the cell. PPAR? binding was globally reduced in adipocytes after HFD feeding, and Rosi restored the molecular and cellular phenotypes of adipocytes associated with HFD feeding. We identified the TGF?1 effector protein SMAD to be enriched at HFD-induced promoters and enhancers and associated with myofibroblast signature genes. TGF?1 treatment of mature 3T3-L1 adipocytes induced gene expression and cellular changes similar to those seen after HFD in vivo, and knockdown of Smad3 blunted the effects of TGF?1. CONCLUSIONS:Our data demonstrate that adipocytes fail to maintain cellular identity after HFD feeding, acquiring characteristics of a myofibroblast-like cell type through reduced PPAR? activity and elevated TGF?-SMAD signaling. This cellular identity crisis may be a fundamental mechanism that drives functional decline of adipose tissues during obesity.
Project description:This study confirms the anti-obesity effect of the ethyl acetate fraction of Distylium racemosum (DRE), a member of Hamamelidaceae, that naturally grows on Jeju Island, on adipocyte differentiation in 3T3-L1 cells. This study further demonstrated that DRE exhibits anti-obesity effects in C57BL/6 obese mice. The degree of adipocyte differentiation was determined using Oil red O stain; results indicated a decrease in fat globules, which was dependent on DRE concentration, when pre-adipocytes were treated with differentiation-inducing agents. In addition, this significantly reduced the expression of the adipogenic transcription factor and related genes. C57BL/6 obese mice treated with DRE showed a lower rate of body weight gain than the high-fat diet (HFD) group mice. Further, the level of serum triglyceride in the DRE treatment group was lower than that in the HFD group. The findings show that DRE are capable of suppressing adipocyte accumulation; therefore, DRE may represent a promising source of functional materials for the anti-obesity.
Project description:White adipose tissue (WAT) expansion in obesity occurs through enlargement of preexisting adipocytes (hypertrophy) and through formation of new adipocytes (adipogenesis). Adipogenesis results in WAT hyperplasia, smaller adipocytes and a metabolically more favourable form of obesity. How obesogenic WAT hyperplasia is induced remains, however, poorly understood. Here, we show that the mechanosensitive cationic channel Piezo1 mediates diet-induced adipogenesis. Mice lacking Piezo1 in mature adipocytes demonstrated defective differentiation of preadipocyte into mature adipocytes when fed a high fat diet (HFD) resulting in larger adipocytes, increased WAT inflammation and reduced insulin sensitivity. Opening of Piezo1 in mature adipocytes causes the release of the adipogenic fibroblast growth factor 1 (FGF1), which induces adipocyte precursor differentiation through activation of the FGF-receptor-1. These data identify a central feed-back mechanism by which mature adipocytes control adipogenesis during the development of obesity and suggest Piezo1-mediated adipocyte mechano-signalling as a mechanism to modulate obesity and its metabolic consequences.
Project description:Retinoblastoma (Rb1) has been described as an essential player in white adipocyte differentiation in mice. No studies have been reported thus far in human adipose tissue or human adipocytes. We aimed to investigate the possible role and regulation of RB1 in adipose tissue in obesity using human samples and animal and cell models. Adipose RB1 (mRNA, protein, and activity) was negatively associated with BMI and insulin resistance (HOMA-IR) while positively associated with the expression of adipogenic genes (PPAR? and IRS1) in both visceral and subcutaneous human adipose tissue. BMI increase was the main contributor to adipose RB1 downregulation. In rats, adipose Rb1 gene expression and activity decreased in parallel to dietary-induced weight gain and returned to baseline with weight loss. RB1 gene and protein expression and activity increased significantly during human adipocyte differentiation. In fully differentiated adipocytes, transient knockdown of Rb1 led to loss of the adipogenic phenotype. In conclusion, Rb1 seems to play a permissive role for human adipose tissue function, being downregulated in obesity and increased during differentiation of human adipocytes. Rb1 knockdown findings further implicate Rb1 as necessary for maintenance of adipogenic characteristics in fully differentiated adipocytes.
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:The preadipocyte-to-adipocyte differentiation (adipogenesis) is a key process in fat mass increase and thus it is regarded as a compelling target for preventing or treating obesity. Of adipogenic hormone receptors, peroxisome proliferator-activated receptor gamma (PPAR?) has crucial roles in adipogenesis and lipid accumulation within adipocytes. Here we demonstrate that the NEDD8 (neuronal precursor cell expressed, developmentally downregulated 8)-based post-translation modification (neddylation) of PPAR? is essential for adipogenesis. During adipogenesis, NEDD8 is robustly induced in preadipocytes and conjugates with PPAR?, leading to PPAR? stabilization. When the neddylation process was blocked by NEDD8-targeting siRNAs (or viral vectors) or an inhibitor MLN4924, adipocyte differentiation and fat tissue development were substantially impaired. We also demonstrate that MLN4924 effectively prevents the high-fat diet-induced obesity and glucose intolerance in mice. This study provides a better understanding of how the PPAR? signaling pathway starts and lasts during adipogenesis and a potential anti-obesity strategy that targets the neddylation of PPAR?.
Project description:Berries of Aronia melanocarpa (chokeberry) are known to be a rich source of biologically active polyphenols. In the present study, the effects of seven anti-adipogenic polyphenolic phytochemicals isolated from A. melanocarpa methanol extract on adipogenic transcription factors were investigated. Amygdalin and prunasin were found to inhibit 3T3-L1 adipocyte differentiation by suppressing the expressions of PPAR? (peroxisome proliferator-activated receptor ?), C/EBP? (CCAAT/enhancer binding protein ?), SREBP1c (sterol regulatory element binding protein 1c), FAS (fatty acid synthase), and aP2 (adipocyte fatty-acid?binding protein). A. melanocarpa extract-treated (100 or 200 mg/kg/day on body weight) high fat diet (HFD)-induced obese mice showed significant decreases in body weight, serum triglyceride (TG), and low-density lipoprotein cholesterol (LDLC) levels and improved insulin sensitivity as compared with HFD controls. This research shows A. melanocarpa extract is potentially beneficial for the suppression of HFD-induced obesity.
Project description:Peroxisome proliferator-activated receptor-? (PPAR?) is a dietary lipid sensor, whose activation results in hypolipidemic effects. In this study, we investigated whether PPAR? activation affects energy metabolism in white adipose tissue (WAT). Activation of PPAR? by its agonist (bezafibrate) markedly reduced adiposity in KK mice fed a high-fat diet. In 3T3-L1 adipocytes, addition of GW7647, a highly specific PPAR? agonist, during adipocyte differentiation enhanced glycerol-3-phosphate dehydrogenase activity, insulin-stimulated glucose uptake, and adipogenic gene expression. However, triglyceride accumulation was not increased by PPAR? activation. PPAR? activation induced expression of target genes involved in FA oxidation and stimulated FA oxidation. In WAT of KK mice treated with bezafibrate, both adipogenic and FA oxidation-related genes were significantly upregulated. These changes in mRNA expression were not observed in PPAR?-deficient mice. Bezafibrate treatment enhanced FA oxidation in isolated adipocytes, suppressing adipocyte hypertrophy. Chromatin immunoprecipitation (ChIP) assay revealed that PPAR? was recruited to promoter regions of both adipogenic and FA oxidation-related genes in the presence of GW7647 in 3T3-L1 adipocytes. These findings indicate that the activation of PPAR? affects energy metabolism in adipocytes, and PPAR? activation in WAT may contribute to the clinical effects of fibrate drugs.
Project description:Geldanamycin derivatives are benzoquinone ansamycin antibiotics that bind to Hsp90 and alter its function. The alteration of Hsp90 activity limits some cellular hormonal responses by inhibiting nuclear receptors activation. The nuclear receptors activity, such as PPAR?, the mineralocorticoid and glucocorticoid receptors (MR and GR) play a critical role in the conversion of preadipocytes to mature adipocytes. Given the importance of these nuclear receptors for adipogenesis, we investigated the effects of geldanamycin analogues (GA) on adipocyte differentiation and function. We found that early exposure of preadipocyte cells to GA inhibited their conversion into mature adipocytes by inhibiting the adipogenic transcriptional program and lipid droplets accumulation. Furthermore, GA altered the adipokines secretion profile of mature adipocyte. The anti-adipogenic effect of GA was also confirmed in mice fed a high fat diet. Biochemical analysis revealed that anti-adipogenic effects of geldanamycin analogues may result from the simultaneous inhibition of MR, GR and PPAR? activity. Taken together, our observations lead us to propose Hsp90 as a potent target for drug development in the control of obesity and its related metabolic complications.
Project description:<h4>Objective</h4>Adipogenesis plays an essential role in maintaining energy and hormonal balance. Cavin-2, one of the caveolae-related proteins, is abundant in adipocytes, the leading site of adipogenesis. However, the details of the roles of Cavin-2 in adipogenesis remain unknown. Here, we demonstrate the requirement of Cavin-2 for the expression and stability of IRβ in adequate adipocyte differentiation.<h4>Methods</h4>Cavin-2 knockout (Cavin-2 KO) and wild-type (WT) mice were fed with a high-fat diet (HFD) for 8 weeks. We evaluated body weight, food intake, and several tissues. Glucose homeostasis was assessed by glucose and insulin tolerance tests. Insulin signaling in epididymal white adipose tissue (eWAT) was determined by Akt phosphorylation. In vitro study, we evaluated adipocyte differentiation, adipogenesis-related genes, and insulin signaling to clarify the relationship between Cavin-2 and adipogenesis under the manipulation of Cavin-2 expression.<h4>Results</h4>Caveolae structure decreased in eWAT of Cavin-2 KO mice and Cavin-2 knockdown 3T3-L1 cells. Cavin-2 enhanced the stability of insulin receptor (IR) through direct association at the plasma membrane in adipocytes, resulting in accelerated insulin/IR/Akt signaling-induced adipogenic gene expression in insulin-containing solution-stimulated 3T3-L1 adipocytes. IR-mediated Akt activation also enhanced Cavin-2 and IR expression. Cavin-2 knockout mice showed insulin resistance with dyslipidemia and pathological hypertrophic adipocytes after a HFD.<h4>Conclusions</h4>Cavin-2 enhances IR stability through binding IR and regulates insulin signaling, promoting adequate adipocyte differentiation. Our findings highlight the pivotal role of Cavin-2 in adipogenesis and lipid metabolism, which may help to develop novel therapies for pathological obesity and adipogenic disorders.