Study On Molecular Mechanism Of Hyperglycemia In T2D Animal Model GK Rats During The Early Stage

Type 2 diabetes(T2D)is a complex metabolic disease,which is caused by a variety of factors.The number of T2 D patients and incidence of T2 D are increasing worldwide,which threatens human health,seriously.Compraed with T2 D paients in Europen and American,the mean body mass index(BMI)of Chinese and Asian T2 D patients is lower.It has been reported that about 70% of T2 D patients in China are non-obese.Moreover,postprandial hyperglycemia is common among Chinese and even Asian T2 D patients.Therefore,more attention should be paid to the postprandial hyperglycemia of non-obese T2 D in China and Asia.Non-obese T2 D animal mode Goto-Kakizaki(GK)rat is a genetic rat model with multiple gene defects.Moreover,there are several similar characteristics between GK rats and non-obese T2 D patients,such as hyperglycemia,reduced insulin secretion of islet ? cells,insulin resistance of peripheral tissues and increased hepatic gluconeogenesis.Considering these characteristics,GK rats may be helpful in studying the mechanism of non-obese T2 D.Skeletal muscle is the main tissue for postprandial glucose uptake,and it is responsible for 70%-80% of postprandial glucose uptake.Therfore,skeletal muscle might play key roles in the developing of hyperglycemia.However,there is little study to investigate the regulation mechanism of skeletal muscle on postprandial hyperglycemia,especially postprandial hyperglycemia during early stage of T2 D.Based on transcriptome sequencing,this study aims to investigate the molecular mechanism of skeletal muscle m RNA and long non-coding RNA(lnc RNA)on hyperglycemia of GK rats duiring early stage of T2 D,and integrating the genome and transcriptome to investigate the genetic mechanism of early hyperglycemia in GK rats,with the expectation of filling the gaps in the regulatory mechanism of hyperglycemia in skeletal muscle during early stage of T2 D.The specific research contents are as follows:Firstly,we sequenced the transcriptome of skeletal muscle of non-obese T2 D animal mode GK rats and control Wistar rats at the age of 3 and 4 weeks to investigate the molecular mechanism of skeletal muscle m RNA on hyperglycemia of GK rats during the early stage of T2 D.The results from transcriptome data analysis and real-time PCR showed that the expression of Tbc1d4 was significalty lower in skeletal muscle of GK rats at the age of 3 and 4 weeks,Pdk4 was significalty higher in skeletal muscle of GK rats at the age of 3 and 4 weeks,genes Acadl,Acsl1 and Fabp4 were up-regulated in skeletal muscle of GK rats at the age of 4 weeks.Among these genes,protein encoded by gene Tbc1d4 was involved in insulin-stimulated glucose uptake.Pdk4 was implicated in glucose uptake and oxidation.While,proteins encoded by genes Acadl,Acsl1 and Fabp4 were implicated in fatty acids oxidation.Previous studies have shown overexpression or knockout of Tbc1d4,Pdk4,Acadl,Acsl1 and Fabp4 could change glucose uptake and fatty acid oxidation in rodents.Thus,these dysregulated genes Tbc1d4,Pdk4,Acadl,Acsl1 and Fabp4 might be raleted to the reduced glucose uptake and oxidation,increased fatty acid oxidation,resulting postprandial hyperglycemia in GK rats during early stage of T2 D.Secondly,we sequenced skeletal muscle transcriptomes including lnc RNAs and m RNAs,in GK and Wistar rats at the age of 3 and 4 weeks,to invistage the molecular mechanism of skeletal muscle lnc RNA on hyperglycemia of GK rats during the early stage of T2 D.After analysis of the differentially expressed m RNA and lnc RNA,we analyzed the correlation coefficients between differentially expressed lnc RNA and differentially expressed m RNA,prediction of targets m RNA for differentially expressed lnc RNA and protein-protein interactions.We found these differentially expressed genes Pdk4,Stc2,Il15,Fbxw7 and Ucp3 might be regulated by lnc RNAs and involved in glucose intolerance,increased fatty acid oxidation and hyperglycemia.The differentially expressed lnc RNA-m RNA pairs NONRATG017315.2-Pdk4,NONRATG003318.2-Stc2,NONRATG011882.2-Il15,NONRATG013497.2-Fbxw7 and MSTRG.1662-Ucp3 were with a high correlation coefficient.In addition,these lnc RNA-m RNA pairs NONRATG011882.2-Il15,NONRATG013497.2-Fbxw7 and MSTRG.1662-Ucp3 were differentially expressed lnc RNA-target m RNA pair.Therefore,these differentially expressed lnc RNA-m RNA pairs NONRATG017315.2-Pdk4,NONRATG003318.2-Stc2,NONRATG011882.2-Il15,NONRATG013497.2-Fbxw7,MSTRG.1662-Ucp3 in skeletal muscle of GK rats at the age of 3 and 4 weeks,might be involved in glucose intolerance,increased fatty acid oxidation and hyperglycemia.Lastly,we integrated the analysis of PASS(putative artificial selective sweeps),SAAC(specific amino acid changed)and the data of transcriptome sequencing.The results showed that genes Ide,Tgm2,Prkcq,Esr1,Zbtb16,Col5a3 and Tgm2 were both PASS region genes and special SAAC genes in GK rats;Egr2,Slc25a20,Ech1,Pcnt,Nrip1,Ephx2,Cmah and Ucp2 were special SAAC genes in GK rats.In addition,these genes were differentlay expressed in GK rats compared with control Wistar rats.Previous studies have shown that genes Tgm2,Ephx2,Pcnt,Ucp2 and Cmah were implicated in impaired insulin secretion.Genes Prkcq,Esr1,Zbtb16,Col5a3,Ide,Nrip1,Egr2 and Ech1 were reported to involved in insulin resistance,and genes Col5a3 and Nrip1 were reported to implicated in regulation of glucose uptake in skeletal muscle.Thus,these special mutations of above genes and their differentially expressed in GK rats might be related to impaired insulin secretion,insulin resistance,and reduced glucose uptake in skeletal muscle,as well as might be involved in the regulation of hyperglycemia in GK rats during early stage of T2 D.Conclusion: Integrating analysis of transcriptome dats of GK rats and control Wistar rats at the age of 3 and 4 weeks,and genome data of GK rats,combined with previous studies of genes knockout and knockin,this study amis to reveal hyperglycemia mechanism in GK rat during eary stage of T2 D.The study found the differentially exressed genes and differentially lnc RNA-m RNA pairs might be involved in the regulation of glucose intolerance,increased fatty acid oxidation,and hyperglycemia.As well as the study found that special genes mutitations in GK rats might be implicated in regulation of impaired insulin secretion,insulin resistance and decreased glucose uptake in skeletal muscle in GK rats.Our data might provide a theoretical evidence for the regulation of hyperglycemia in non-obese T2 D during early stage,and provide a potential candidate target for T2 D prevention and control in China and Asia.