Upregulation Of RBM3 Affects OGT-mediated Glycolysis In The Gastrocnemius Muscle Of Cold Exposed Mice

The organism basal metabolism and immune function was significantly changed at ambient hypothermia.Loss of UCP1 gene in domestic pigs resulted in lack of functional BAT in response to hypothermia environmental.The data showed that low temperatures in piglet nurseries could lead to viral or bacterial diarrhea.Furthermore,the pig breeding industry in piglet heating energy system expenditure was huge annually.Thus,studying the muscle cold response mechanism of important heat-producing organs in piglets at low temperature was of great significance for improving the low temperature resistance of piglets and reducing the energy expenditure in pig breeding industry.A preliminary study by the research team found that the significant changes in the glycolysis metabolism of the gastrocnemius muscle were synchronous with the changes in the expression of the cold-response protein RBM3 and the stress receptor protein OGT during the stable period of blood glucose in piglets under low temperature.In this study,mice were used as animal models to study changes in response to relevant signaling pathways and biochemical pathways.In this study,the changes of blood glucose and insulin in cold-exposed fasted 2 h and 4 h mice were measured.The results showed that blood glucose remained stable but insulin fluctuated,compared with the normal temperature control group.The content of glycogen and ATP in the gastrocnemius muscle of mice cold stimulation for 4 h was significantly decreased,compared with the normal temperature control group.Meanwhile,PI3K/AKT glycolytic,glucose transport and glycogen synthesis pathway-related proteins were detected.The results showed that the phosphorylation of AKT/GSK3?/GS,AKT/P-PFKFB2,p65 was significantly increased in coldexposed 4 h mice,and the expression levels of OGT,RBM3,Glut4 and Bcl-2/Bax proteins were significantly up-regulated.However,the expression of Cleaved-Caspase3 in the gastrocnemius muscle of mice cold-exposed for 4 h was significantly down-regulated.These results potentially demonstrates that the gastrocnemius muscle responds to cold exposure by regulating OGT,RBM3,Glut4,Bcl-2/Bax protein expression and PI3K/AKT glycolysis,glucose transport,and glycogen synthesis by 4 h of cold exposure.In this study,the AKT inhibitor of wortmannin was used to verify whether it affects the expression of cold-response-related protein expression in the gastrocnemius muscle after 4 h cold exposure.The results showed that the IRS-1 phosphorylation was no significant changed between the control group,AKT inhibitor group and DMSO group after 4 h cold exposure;while the content of Cleaved-Caspase 3 was significantly increased in contrast to Bcl-2/Bax expression,and the protein expression levels of OGT,RBM3,Glut4 and the phosphorylation of AKT/GSK3?/GS,AKT/P-PFKFB2,and AKT/p65 were significantly decreased.To further investigate the presence of crosstalk between RBM3,p65,AKT and OGT,the lentiviral vector was used to overexpress RBM3 protein in C2C12 mouse myoblasts.The results showed that the overexpression RBM3 group significantly promoted AKT phosphorylation at room temperature and 4 °C,compared with the wild-type and empty plasmid controls.Meanwhile,the results from conditional knockout of mouse skeletal muscle OGT showed that AKT,GSK3?,PFKFB2 phosphorylation and CleavedCaspase 3 expression were significantly increased,but the phosphorylation of p65,GS,RBM3 and Bcl-2/Bax expression were significantly decreased in OGT knockout mice compared to wild type mice.At the same time,the accumulation in glycolysis products of FDP and PA were detected in the gastrocnemius muscle of OGT knockout mice.These results indicate that the presence of OGTmediated RBM3 expression promotes AKT phosphorylation and OGT mediates AKT glycolysis flux.Based on the above results,researcher found that cold-exposed mouse gastrocnemius OGT mediates up-regulation of RBM3.cold-exposed mice gastrocnemius up regulates RBM3 promote AKT phosphorylation and participates in cytoprotection and glycolysis.cold-exposed mouse gastrocnemius OGT-mediated insulin glycolysis Flux.