Research On Regulation Of Muscle Fiber Transformation Through Adiponectin Pathway By Reseratrol

Resveratrol can improve muscle pH,flesh,hydration,muscle tenderness and effectively ameliorate meat quality.As the only negative regulation hormones in fat cell factors,adiponectin(AdipoQ)plays an important role in skeletal muscle oxidation and metabolism.In order to explore the molecular mechanism of resveratrol in regulating the transformation of muscle fibers through adiponectin signaling pathway,The study made a thorough inquiry on the transformation of muscle fiber type through the combination of adiponectin and its receptor in vivo and vitro with the method of over-expression and RNA interference(RNAi).The main results obtained are as follows.1.According to the sequence published in GenBank,a pair of primers were designed to clone the coding sequence(CDS)of AdipoQ gene of Luchuan pig,which was used for bioinformatically analysis.The results indicated that the CDS length of AdipoQ of Luchuan pig was 732 bp,encoding 242 amino acids,and its molecular weight was 26400.7 Da.Nucleotide sequence analysis showed that the homology of AdipoQ gene was higher among different species.The protein structure of Luchuan pig AdipoQ has a strong hydrophilicity by the prediction of protein structure.Threre was highly random coil hydrophilia and alpha helix,a low level of beta sheet,where contained a signal peptide,which was a secreted protein.Meanwhile,a lentiviral expression vector,pLV-AdipoQ,was constructed for further study.2.The pLV-AdipoQ recombinant lentiviral vector plasmids were transfected into C2C12 cells and the AdipoQ gene successfully expressed.The results on the cell differentiation in Recombinant gAcrp30 treated group and pLV-AdipoQ overexpression group showed that the expression levels of AdipoRl?AdipoR2?AMPK?PGC-1??MyHCl?MyHC2a were significantly increased compared with the control group.The above results indicated that AdipoQ gene has the function in regulating muscle fiber type transformation.3.In the process of differentiation in C2C12 cells,comparing with control group,the results of three groups which treated with resveratrol,recombinant rat adiponectin gAcrp30,and gAcrp30 combining with resveratrol,showed that the two groups treated with resveratrol and gAcr:p30 could significantly up-regulated the expression of AdipoQ?AdipoRl?AdipoR2?AMPK?PGC-1???MyHC1 and down-regulated that of MyHC2b gene.In the group treated with gAcrp30 combining with resveratrol showed that there was a synergistic effect in the regulation of muscle fiber type transformation.The expression of AMPK and PGC-1?was down-regulated with AdipoRl gene disturbed.When the expression level of AdipoR2 was significantly increased in C2C12 cells which treated with resveratrol,however,there were no obvious differences in those of AMPK and PGC-1? gene.With AdipoR2gen expression was downregulated,the expression level of AdipoRl?AMPK and PGC-1?genes were not affected,however,those showed an upward trend in C2C12 cells which treated with resveratrol.Meanwhile,resveratrol could increase the expression level of AdipoRl,AdipoR2,AMPK,PGC-1?genes after AdipoRl and AdipoR2 genes were both down-regulated by interference.Comprehensive cell level validation results suggested that resveratrol could adjust the type of muscle fiber through the binding of AdipoQ with its AdipoRl in C2C12 cells and stimulating AMPK and PGC-la activity.4.Compared to the control group,there was no significant difference in body weight,feed intake and biochemical indexes of mice which were fed with resveratrol for 12 weeks.Meanwhile,the expression of AdipoQ in liver,adipose tissue and serum was significantly higher than that in control group.In the extensor digitorum longus and soleus muscle of experimental group mice fed with resveratrol in experimental group,the expression of AdipoQ,AdipoRl,AdipoR2,AMPK,PGC-1?,MyHC1,MyHC2a and MyHC2x were significantly higher than those in control group,however,the expression of MyHC2b was lower than that of control group.In summary,resveratrol increased adipoRl gene expression through adiponectin signaling pathway,activated AMPK and the expression of PGC-1? gene in the downstream and then regulated the muscle fiber transforming to the oxidative type.