Proteomics Analysis And Study Of Skeletal Muscle Tissue In Chinese Hamsters With Spontaneous Type 2 Diabetes Mellitus

Objective:The identification of differentially expressed proteins in Chinese hamsters skeletal muscle tissue of type 2 diabetes mellitus(T2DM)by quantitative proteomics of TMT labeling is expected to find potential targets related to the pathogenesis of T2 DM.Methods:Diabetic Chinese hamsters(FBG ≥ 6.0 mmo L/L)and control hamsters(FBG ≤4.5 mmo L/L)were selected according to fasting blood glucose value(FBG),skeletal muscle tissue was selected for HE staining and proteomics identification,and functional enrichment analysis of differentially expressed protein was performed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG).Candidate differentially expressed proteins were validated by Western blot and q RT-PCR,and the content of reactive oxygen species(ROS)in skeletal muscle tissue and serum was determined by ELISA.5-oxoprolinase(OPLAH)gene knockdown by interfering small RNA(si RNA)in human skeleta L muscle cells(HSKMC Cells),ROS content was detected by DCFH-DA probe,GSH content by GSH kit,key protein expression of PI3 K / Akt / GLUT4 pathway by Western blot,and glucose uptake by 2-NBDG probe.Results:The HE pathology results of Chinese hamster skeletal muscle tissue showed that the skeletal muscle structure was normal,and the muscle fiber cell nuclei were neatly arranged in the control group,and a large number of muscle fiber cell nuclei and increased cell spacing in the diabetic group.Proteomics in diabetic Chinese mouse skeletal muscle tissue of 38 differential expressed protein(14 up,24 down),GO function enrichment analysis results showed the differential expressed protein function and inorganic ion transmembrane transport,gene expression regulation,transferase activity and pentyl,voltage gated ion channel activity and voltage gated channel activity.The results of KEGG pathway enrichment analysis showed that the differentially expressed proteins were mainly enriched in the six pathways: ABC transporters,purine metabolism,cytochrome P450 to isobiotic hormone metabolism,chemical carcinogenicity,glutathione metabolism and metabolic pathways.Five differentially expressed proteins(SIRT5,ALDH1L1,EPHX1,OPLAH,and GST)were confirmed at the protein level and the m RNA level,and the results were consistent with the proteomic results.The ROS content in skeletal muscle tissue and serum of diabetic gopher was significantly higher than in the control group.Knockdown of OPLAH in HSKMC cells induced increased ROS content,decreased GSH content,decreased phosphorylation of PI3 K and Akt,decreased GLUT4 expression,and decreased glucose uptake.Conclusion:In this study,the differentially expressed proteins identified in Chinese hamster skeletal muscle using TMT labeling may be associated with T2 DM,and all the five differentially expressed proteins validated were associated with oxidative stress.Knockdown of OPLAH may lead to insulin resistance(IR)and glucose uptake disorders in HSKMC cells through oxidative stress,so OPLAH may be a new therapeutic target for T2 DM.