The Regulatory Mechanism Of Histone Modifying Enzymes In Diabetic Complications

The prevalence of type2diabetes mellitus continuously increases all across the world, and it becomes one of major threats to human health. Type2diabetes mellitus is accompanied with many complications, including non-alcoholic fatty liver desease (NAFLD) and diabetic nephropathy (DN).NAFLD is characterized by over accumulation of lipid in the liver in the absence of history of significant alcohol use. This clinical syndrome is one of common complications of type2diabetes. Insulin resistance is one of major causes of type2diabetes. Insulin resistance is defined as the pathological condition in which tissues are insensitive to insulin, and fail to response to insulin regulation. Now, it has been demonstrated that insulin resistance is also one of major causes of NAFLD. In this thesis, we found the decreased H3K9me2and G9a levels in the livers of db/db mice, a typical type2diabetic mouse model. In HepG2and LO2cells, two cultured hepatic cell lines, knocking down G9a decreased the expression levels of insulin receptor (IR)、 p-AKT and p-GSK3β,which led to dysfunction of insulin signaling. Meanwhile, overexpression of G9a in cultured hepatic cells enhanced the levels of IRα and p-AKT, suggesting the activation of insulin signaling. Further investigation revealed that, the impairment of insulin signaling caused by palmitate and glucosamine, two insulin resistance inducers, were rescued by overexpression of G9a in cultured hepatic cells. Through the analysis of microarray, we discovered that HMGA1and ARRB2, two genes that previously have been shown to play important roles in the pathogenesis of insulin resistance, were decreased in the G9a-deficient hepatic cell lines. Meanwhile, G9a overexpression enhanced the protein levels of HMGA1and ARRB2in cultured hepatic cell lines. In addition, the luciferase assay also demonstrated that the overexpression of G9a enhanced the transcription levels of HMGA1and ARRB2. Further, overexpressions of HMGA1or ARRB2in the G9a-deficient cells could rescue the G9a deficient-induced downregulation of insulin signaling. More importantly, the injection of AAV2/8virus harbored shG9a in normal mice impaired the hepatic insulin signaling. While, the injection of adenovirus-mediated G9a plasmid in db/db mice improved the impaired hepatic insulin signaling, and reduced the high blood glucose level.Inflammation is one of the important mechanisms that contributes to the development of several renal diseases. Chronic and acute inflammation lead to renal damage, or renal failure. However, the mechanisms responsible for the up-regulation of these inflammatory factors are still unclear. In this thesis, we used two kinds of renal disease models:diabetic nephropathy (kidneys from of db/db mice) and lipopolysaccharide (LPS)-induced acute kidney injury (AKI). In these two models, inflammatory factors were enhanced in kidneys. Meanwhile, the levels of histone acetylations and PCAF, a histone acetyltransferase, were also significantly increased. In HK-2cells, a human proximal tubule epithelial cell line, PCAF knockdown led to the decrease of H3K18ac, but not H3K9ac. Furthermore, knockown of PCAF led the decreases of inflammatory factors including VCAM-1、ICAM-1、MCP-1and p50subunit of NF-κB (p50) in the mRNA and protein levels. Consistent with these results, the expressions levels of these inflammation-related factors were enhanced by overexpression of PCAF in HK-2cells. In addition, the CHIP assays demonstrated that PCAF-deficiency reduced the recuitment of H3K18ac on promoters of ICAM-1and MCP-1in the presence or absence of palmitate. Using LPS-induced AKI mice, we further demonstrated that LPS promoted the recruitment of H3K18ac on the promoters of VCAM-1, ICAM-1and MCP-1.In summary, we studied the roles of two distinct histone modifying enzymes in two different diabetic complications. The results demonstrated that G9a regulated insulin signaling pathway through HMGA1and ARRB2in the liver. In kidney, PCAF regulated the expressions of inflammatory factors through H3K18ac, to trigger the inflammatory response. Thus, my thesis demonstrates that epigenetic regulation plays important roles in the pathogenesis of diabetic complications, and provides some possible targets for future treatments.