The Study Of Effects And Mechanisms Of TET2 On Atherosclerotic Plaque In ApoE^-/- Mice

TET2 protein(ten-eleven-translocation 2) one of TET(ten-eleven-translocation) family members, is highly conserved in evolution and plays a key role in DNA demthylation via catalyzing the oxidation of 5-methylcytosine(5m C) into5-hydroxymethylcytosine(5hm C). Previous studies showed that autophagy dysfunction was contributed to atherosclerotic lesions. Meanwhile,abnormal DNA methylation profile was detected even in early atherosclerotic lesions. Our provious studies showed that autopghagic dysfunction and TET2 expression inhibition in high-fat fed Apo E-/- mice atherosclerotic lesions.Based on these results, we speculated that TET2 inhibited atherosclerotic lesions via improving autophagy. To test this hypothesis, the effect of overexpression TET2 on high-fat fed Apo E-/- mice atherosclerotic lesions and autophagy were detected. Then, the effect of overexpression TET2 on Human umbilical vein endothelial cells(HUVECs) and its potential mechanisms were explored.Objective: To investigate the effect of TET2 on atherosclerotic lesion and its potential mechanisms.Methods: After one week of adaptive feeding, forty male eight-week-old apo E-/- mice were randomly divided into two groups: overexpression TET2 group and control group(n = 20). The overexpression group was injected overexpression TET2 lentivirus 4×107 TU through tail vein every six weeks and the control group was treated with saline. After fed withhigh-fat diet for 12 weeks, all mice were sacrificed. The plasma lipid levels were detected by Automatic biochemical analyzer. The atherosclerotic lesions in aorta and aortic sinus were measured with Sudan? staining and HE staining. The lipid accumulation was observed with Oil Red O staining. Immunofluorescence was used to investigate the expression of TET2 and BECN1, microtubule-associated protein light chain 3?(LC3?), p62, 5m C and 5hm C. In cultured HUVECs, after treated with overexpression TET2 plasma and TET2 si RNA for 24 h,HUVECs wereincubuted with 50?g/m L ox-LDL for another 24 h. The levels of TET2, 5m C and 5hm C were detected with Immunofluorescence.The expression of Beclin 1 ? LC3 and P62 were detected withwestern blotting. The autophagy flux was detected with m RFP-GFP-LC3 adenovirus. Autophagy-related gene Beclin1 promoter methylation situation was investigated with BSP.Results: There was were no significant change of plasma lipid levels between the TET2 overexpression group and the control group. However,the aortic atherosclerotic lesion area and lipid accumulaion of the TET2 overexpression group were obviously decreased. In TET2 overexpression group, the content of Beclin1, LC3 ?, 5hm C were significantly increasedand the expression of p62 was significantly decreased. In cultured HUVECs, overexpression TET2 increased the contents of TET2,Beclin 1, LC3? and 5hm C, decreasing the contents of P62 and 5m C.Consisted with these results, TET2 sh RNA downregulated the contents of Beclin 1, LC3 ? and upregulated the content of P62.m RFP-GFP-LC3 virus transfection showed that, TET2 overexpression significantly enhanced the increased autophagy flux. BSP showed that the methylation of beclin1 promoter was significantly downregulation after treated with TET2 overexpression.Conclusion: TET2 inhibited high fat diet apo E-/-mice atherosclerotic lesions. The mechanism may achieved, at least partly, by regulated endothelial cell autophagy and autophagic flux.