| The prevalence of type 2 diabetes(T2D)in China is increasingly serious,with more than100 million people estimated to have T2 D,resulting in a substantial healthcare burden to our country.Physical activity is a critical element in the early prevention and management of T2 D.It has been observed that a single session of exercise(acute exercise)can attenuate hyperglycemia and enhance insulin sensitivity.Although considerable effort has been made to reveal comprehensive changes stimulated by exercise,its relevant mechanisms are still unclear in T2 D.Compared with T2 D patients in Europe and America,the mean body mass index in Chinese patients with T2 D is much lower,and non-obese T2 D is the primary type.Also,postprandial hyperglycemia is more pronounced in Chinese patients with early T2 D than most other ethnic groups.The Goto-Kakizaki(GK)rat,a spontaneous T2 D rodent model,was generated by repeated inbreeding of Wistar rats selected at the upper limit of the normal distribution for glucose tolerance(postprandial hyperglycemia).It is suitable for studying pathogenic mechanisms and therapeutic approaches of human T2 D patients in China.Skeletal muscle is one of the predominant sites involved in glucose disposal,accounting for 60-80% of postprandial glucose uptake,and plays a critical role in maintaining glycemic homeostasis.The hypothalamus also plays a vital role in modulating energy homeostasis by integrating and adapting to peripheral signals,including nutrients and hormones,and regulating the metabolism of peripheral tissues,including skeletal muscle.To reveal the molecular mechanisms of exercise to improve non-obese and postprandial hyperglycemia-induced T2 D in China,we investigated the effects of acute exercise(15 m/min for 60 min)on skeletal muscle and hypothalamic responses in GK rats.We measured animal physiological indicators and assessed transcriptional profiles in skeletal muscle and hypothalamus from both 8-week-old GK and Wistar rats with and without exercise by RNA sequencing in the current study.Based on gene expression profiling,we conducted comparative transcriptomic analysis among GK,Wistar,exercise Wistar,and exercise GK rats.By integrating our transcriptomic data(results of GO and KEGG functional enrichment analysis,protein-protein interaction network analysis,cluster analysis,transcriptional regulatory network and co-expressed network)and previous studies of gene knockout or overexpression,we identified key differentially expressed genes and hypothalamus-skeletal muscle co-expression gene pairs associated with exercise to ameliorate hyperglycemia and improve insulin sensitivity.We also studied the effect of the Klf3 gene selected from the analysis of transcriptome on glucose uptake and insulin action in rat L6 myotubes.The main results are as follows.(1)At 8 weeks of age,Wistar rats were heavier than GK rats.A significantly higher level of random plasma glucose was observed in GK rats than in Wistar rats,and the insulin level was also elevated.After acute exercise,plasma glucose and insulin levels were reduced in exercise GK rats compared to that of GK rats,indicating that acute exercise could improve hyperglycemia and insulin sensitivity.(2)Through pairwise comparisons of skeletal muscle transcriptome,we identified 819,291,and 598 differentially expressed genes between GK and Wistar,exercise GK and GK,and exercise Wistar and Wistar,respectively.By integrating our data and previous studies,we found the downregulated Pfkfb3 and upregulated Ppp1r3 c,Slc27a1,Fasn,and Tbc1d1 genes related to energy metabolism and glucose transport in the diabetic GK rats compared to Wistar rats.We predicted that these dysregulated genes might be associated with hyperglycemia and skeletal muscle insulin resistance.After acute exercise,the downregulated Fasn and upregulated Tbc1d1,Hk2,Lpin1,Ppargc1 a,Sorbs1,and Hmox1 genes related to increasing energy metabolism,ameliorating insulin resistance and enhancing glucose uptake might be critical regulators to lower plasma glucose and improve insulin sensitivity in the exercise GK rats.(3)Through pairwise comparisons of hypothalamus transcriptome,we identified 726,197,and 98 differentially expressed genes between GK and Wistar,exercise GK and GK,and exercise Wistar and Wistar,respectively.Based on previous studies,we found the downregulated Nr1d1 gene and upregulated Plin2,Alox15,and Nr1i2 genes related to lipid metabolism dysfunctions,which might be implicated in the disorders of the hypothalamus in the diabetic GK rats.These dysregulated genes might contribute to hyperglycemia or insulin resistance in GK rats.In the exercise GK rats,the expression of Plin2 was significantly decreased,and that of Cd36 and Lpl genes enhancing lipid sensing were significantly upregulated,which might improve lipid metabolism in the hypothalamus,consequently ameliorating T2 D.Moreover,we found that the expression of Wfs1 and Cck related to the nervous system was increased,which might also contribute to the beneficial effects of acute exercise on T2 D.(4)Analyzing the correlation coefficients between differentially expressed genes in the hypothalamus and skeletal muscle,we identified 493 and 255 hypothalamus-skeletal muscle co-expressed gene pairs in exercise GK rats and exercise Wistar rats,respectively.Among coexpressed gene network in exercise GK rats,we found differential expression of hypothalamus genes Dio2 and Crym related to thyroid hormone(TH)signaling and T2 D,and differential expression of skeletal muscle genes Hmox1,Lpin1 and Tbc1d1 associated with improving insulin sensitivity and promoting glucose transport.Therefore,exercise might regulate the coexpressed gene pairs Dio2-Hmox1,Dio2-Lpin1,Crym-Hmox1 and Crym-Tbc1d1,change the hypothalamic TH signaling pathway,increase skeletal muscle insulin sensitivity,promote glucose uptake,and thus improve hyperglycemia in GK rats.(5)In L6 myotubes,knockdown of the Klf3 gene promoted basal and insulin-stimulated glucose uptake,while overexpression of the Klf3 gene resulted in the opposite.Through pairwise comparisons of L6 cell transcriptome,we identified 2256 and 1988 differentially expressed genes in the knockdown and overexpression of Klf3 groups,respectively.In insulin signaling,the expression of genes Slc2a4,Akt2,Insr,Sorbs1,Ptprf and Fasn was significantly changed in the Klf3 gene knockdown and overexpression groups.In addition,the expression of gene Gys1 related to glucose metabolism was significantly upregulated in the Klf3 gene knockdown group.In the Klf3 gene overexpression group,we found the Hk2 gene was downregulated,and the Ppp1r3 c gene was upregulated.Combined with previous studies,it is speculated that the changes in the expression of these genes might be related to the regulations of insulin action and glucose uptake of the Klf3 gene.In L6 myotubes,downregulation of Klf3 promoted the expression of GLUT4 and AKT proteins,as well as the translocation of GLUT4 to the cell membrane in the basal situation,and enhanced insulin action,characterized by increased insulin-stimulated GLUT4 translocation and AKT,TBC1D1 and TBC1D4 phosphorylation,while overexpression resulted in the opposite.These results suggest that knocking down the Klf3 gene increases glucose uptake and insulin sensitivity by enhancing insulin signal transduction.In summary,it is the first time to investigate post-acute exercise transcriptomic responses of skeletal muscle and hypothalamus in diabetic GK rats and Wistar rats.By integrating our transcriptomic analysis and the results of gene knockout or overexpression studies reported before,we identified key differentially expressed genes and hypothalamus-skeletal muscle coexpression gene pairs associated with exercise lowering hyperglycemia in GK rats.In addition,this study reported for the first time that knockdown of the Klf3 gene enhanced insulin sensitivity and increased glucose uptake in L6 myotubes,and the exertion of this gene in skeletal muscle might has important implications for the study of acute exercise to improve T2 D.The research reveals some molecular mechanisms of exercise to improve insulin sensitivity and hyperglycemia in GK rats,providing theoretical evidence for the prevention and treatment of non-obese T2 D in China. |
PDF Full Text Request