The Role And Mechanism Of HDAC9 In Diabetic Model Susceptible To Contrast-induced Acute Kidney Injury

Background:Diabetic kidney diseases(DKD)have been recognized as a serious complication of diabetes and a major cause of end-stage renal disease,causing serious health problems and a huge economic burden worldwide.Diabetes is an independent risk factor for Contrast-induced acute kidney injury(CIAKI).The incidence of CIAKI is significantly increased in patients with kidney injury,especially in patients with diabetes-related kidney diseases.The accumulation of reactive oxygen species(ROS)in renal tubular epithelial cells leads to oxidative stress and thus induces cell apoptosis.Hyperglycemia is an important factor to promote CIAKI.On the one hand,it can increase the production of ROS then leading to the occurrence of oxidative stress;on the other hand,hyperglycemic conditions can enhance renal vasoconstriction and cause inadequate renal oxygen supply.CIAKI can also exacerbate the pathological process of diabetes mellitus and diabetic kidney disease,resulting in a sustained decline in renal function.Histone deacetylase 9(HDAC9)belongs to the class IIa family of histone deacetylases,an enzyme that freely shuttles between the nucleus and cytoplasm and promotes tissue-specific transcriptional regulation through interactions with histone and nonhistone substrates.HDAC9 has been shown to be involved in the pathogenesis of chronic diseases,including cardiovascular disease,osteoporosis,autoimmune diseases,cancer,obesity,insulin resistance and liver fibrosis.Previous studies have shown an important role for HDAC9 in renal diseases,but the role and specific mechanisms in diabetic mice susceptible to CIAKI are unclear.We hypothesize that HDAC9 may promote diabetic mice susceptibility to CIAKI by inducing excessive ROS production in renal tubular epithelial cells.Objective:(1)To research the role of HDAC9 in the sensitivity of renal injury induced by contrast media in diabetes mellitus/high glucose;(2)To investigate whether HDAC9 could regulate TXNIP/Trx1/PASK1/p38 MAPK pathway to participate in oxidative stress and apoptosis of renal tubular cells in the susceptibility of diabetes to CIAKI.Methods:(1)In vitro study:(1)HK-2 cells were intervened with high glucose(HG,50 m M)for 44 h,and then iohexol was added for 4h co-culture,cell ROS,cell activity and cleaved caspase-3 were measured.(2)HK-2 cells were treated with 50 m M HG for 48 h,HDAC9 inhibitor(BRD-4354)was preincubated for 24 h before iohexol was added,then iohexol was finally added for 4 h of co-incubation.(3)HK-2 cells were transfected with HDAC9-si RNA,then 50 m M HG was added for 48 h.The cells were incubated with iohexol for 4 h.Cell activity,HDAC9、TXNIP、 Trx-1、P-ASK1、 P38 MAPK and cleaved caspase-3 expression were measured.(2)In vivo study: mouse models of type 2 diabetes、contrastinduced acute kidney injury and diabetic-CIAKI were constructed;HDAC9 inhibitor(BRD-4354)was injected intraperitoneally in the mouse model of diabetic-CIAKI;HE staining was used to observe renal pathological changes,immunofluorescence was performed to observe reactive oxygen species(ROS,reactive oxygen species)and apoptosis in renal tissue.Detection of oxidative stress related indicators(SOD,GSHPX and MDA)expression,and the protein expressions of HDAC9,TXNIP,Trx-1,P-ASK1,P38 MAPK and cleaved caspase-3 were measured by western blot.Results:1.In the diabetic model,treatment with contrast media induced more expression of HDAC9 in renal tubular epithelial cells.2.BRD-4354 attenuated contrast media-induced apoptosis of renal tubular epithelial cells under high-glucose conditions.3.Knockdown HDAC9,HK-2 incubated with high glucose reduced apoptosis and ROS production induced by the addition of contrast media;Meanwhile,knockdown HDAC9 inhibited TXNIP/Trx1/PASK1/p38 MAPK pathway activation in HG+CM group of HK-24.HDAC9 expression was significantly higher in the kidneys of diabetic mice and Diabetic-CIAKI mice than in the control and CIAKI group;5.BRD-4354 alleviated tubular injury in Diabetic-CIAKI mice;it also attenuated oxidative stress and apoptosis in the kidney.Meanwhile,BRD-4354 inhibited TXNIP/Trx1/P-ASK1/p38 MAPK pathway activation in DM-CIAKI mice.Conclusion.HDAC9 may promote the susceptibility of diabetic model to CIAKI;and it is possibly involved in oxidative stress and apoptosis of renal tubular cells by regulating TXNIP/Trx1/P-ASK1/p38 MAPK pathway.