MicroRNA-129-5p inhibits C2C12 myogenesis and represses slow fiber gene expression in vitro.

The miR-129 family is widely reported as tumor repressors, although their roles in skeletal muscle have not been fully investigated. Here, the function and mechanism of miR-129-5p in skeletal muscle, a member of the miR-129 family, were explored using C2C12 cell line. Our study showed that miR-129-5p was widely detected in mouse tissues, with the highest expression in skeletal muscle. Gain- and loss-of-function study showed that miR-129-5p could negatively regulate myogenic differentiation, indicated by reduced ratio of MyHC-positive myofibers and repressed expression of myogenic genes, such as MyoD, MyoG, and MyHC. Furthermore, miR-129-5p was more enriched in fast extensor digitorum longus (EDL) than in slow soleus (SOL). Enhanced miR-129-5p could significantly reduce the expression of mitochondrial cox family, together with that of MyHC I, and knockdown of miR-129-5p conversely increased the expression of cox genes and MyHC I. Mechanistically, miR-129-5p directly targeted the 3'-UTR of Mef2a, which was suppressed by miR-129-5p agomir at both mRNA and protein levels in C2C12 cells. Moreover, overexpression of Mef2a could rescue the inhibitory effects of miR-129-5p on the expression of myogenic factors and MyHC I. Collectively, our data revealed that miR-129-5p is a negative regulator of myogenic differentiation and slow fiber gene expression, thus affecting body metabolic homeostasis.

Elevated miR-10a-5p facilitates cell cycle and restrains adipogenic differentiation via targeting Map2k6 and Fasn, respectively.

miRNAs are a small class of noncoding RNAs that perform biological functions by regulating the stability or translation of target genes in various biological processes. This study illustrated the role of miR-10a-5p, which is relatively enriched in adipose tissues, using primary mouse preadipocytes as model. With elevated miR-10a-5p expression, the proliferative ability of mouse preadipocytes was significantly enhanced, indicated by increased EdU+ cells and G1/S transition, accompanied by upregulated Cyclin B, Cyclin D and PCNA and downregulated p21 and p27. Meanwhile, the adipogenic differentiation was significantly attenuated by elevated miR-10a-5p, supported by Oil Red O staining and suppressed PPARγ and aP2 expression. Furthermore, Map2k6 and Fasn were predicted to be the target genes of miR-10a-5p in silico, and dual luciferase reporter assay confirmed the direct targeting effects. Western blot analysis results showed that miR-10a-5p specially reduced Map2k6 expression at the proliferative stage without affecting Fasn expression, while significantly restrained Fasn expression with unchanged Map2k6 expression during adipogenic differentiation. Taken together, these results revealed a potential role of miR-10a-5p in adipogenesis and in the treatment of obesity.

Triptolide enhances lipolysis of adipocytes by enhancing ATGL transcription via upregulation of p53.

Lipolysis is an essential physiological activity of adipocytes. The Patatin Like Phospholipase Domain Containing 2 (PNPLA2) gene encodes the enzyme adipose triglyceride lipase (ATGL) responsible for triglyceride hydrolysis, the first step in lipolysis. In this study, we investigated the potential of triptolide (TP), a natural plant extract, to induce weight loss by examining its effect on ATGL expression. We found that long- and short-term TP administration reduced body weight and fat weight and increased heat production in brown adipose tissue in wild-type C57BL/6 mice. In 3T3-L1 fibroblasts and porcine adipocytes, TP treatment reduced the number of lipid droplets as determined by Oil Red O and BODIPY staining, with concomitant increases in free fatty acid and triglyceride levels in the culture medium. Combined treatment with TP and p53 inhibitor reversed these lipolytic effects. We next amplified the ATGL promoter region and identified conserved p53 binding sites in the sequence by in silico analysis. The results of the dual-luciferase reporter assay using a construct containing the ATGL promoter harboring the p53 binding site showed that p53 induces ATGL promoter activity and consequently, ATGL transcription. These results demonstrate that TP has therapeutic value as an anti-obesity agent and acts by promoting lipolysis via upregulation of p53 and ATGL transcription.

Butyrate Alleviates High-Fat-Induced Metabolic Disorders Partially through Increasing Systematic Glutamine.

Obesity has emerged as a worldwide epidemic. Both butyrate and glutamine counteract obesity-related metabolic disorders; however, whether and how they synergistically cooperate with each other remains a mystery. In the study, a high-fat diet (HFD, 60% calories from fat) was used to develop a model of obesity-related metabolic disorder and compared with administrated saline and sodium butyrate (SB, 300 mg/kg body weight) daily by gavage. Compared with HFD counterparts, oral administration of SB in mice exhibited significantly reduced body weight and fat mass and decreased hepatic triglyceride content. The targeted mass spectrum revealed that SB restored serum contents of glutamine, which were significantly decreased by HFD. Furthermore, SB significantly elevated the expression of glutamine synthetase (GS, encoded by GLUL) in the liver, accompanied by more enrichment of H3K27ac modifications within its promoter. In summary, the study verified the contribution of elevated glutamine to the beneficial effects of butyrate on metabolic disorders induced by a high-fat diet, providing a novel pathway for understanding how butyrate benefits metabolic homeostasis.