Study On Protective Effect And Mechanism Of HDAC Inhibitor Entinostat(MS-275) On Acute Kidney Injury In Liver Failure

Part I Clinical characteristics and HDAC activity changes in acute kidney injury patients with HBV-related liver failureObjective:Acute kidney injury(AKI)is one of the common complications in patients with liver failure.Inflammatory mediators play crucial roles in the pathogenesis of AKI in HBV-related liver failure patients.Histone deacetylases(HDACs)involve in regulating proliferation and differentiation of normal renal tissue and the pathogenesis of kidney related diseases.The aim of this study was to investigate the clinical characteristics and roles of HDAC activity expression in AKI patients with HBV-related liver failure.Methods:Collected the clinical data and serum samples of AKI patients with HBV-related liver failure,AKI patients with HBV-related cirrhosis,non-AKI HBV-related cirrhosis patients,non-AKI HBV-related liver failure patients(four groups).Summarized and compared the clinical characteristics,detected and compared HDAC activity,levels of NGAL,KIM-1,liver function,renal function,coagulation function,blood routine testing,procalcitonin(PCT)level and HBV DNA replication levels among different groups.Results:The HDAC activities in AKI patients with HBV-related liver failure,AKI patients with HBV-related cirrhosis were significantly higher than that in non-AKI HBV-related liver failure patients and non-AKI HBV-related cirrhosis patients.HDAC activities in AKI patients with HBV-related liver failure were significantly higher than that in AKI patients with HBV-related cirrhosis.The levels of neutrophil cell count(Neu%)and PCT were higher in AKI patients with HBV-related liver failure,AKI patients with HBV-related cirrhosis than that in non-AKI HBV-related liver failure and non-AKI HBV-related cirrhosis patients,and opposite results in estimated glomerular filtration rate(eGFR)levels.The levels of ALT,TBA,TBIL,PCT were significantly higher in AKI patients with HBV-related liver failure than that in AKI patients with HBV-related cirrhosis.The expressions of NGAL,KIM-1 were significantly higher in AKI patients with HBV-related liver failure than that in AKI patients with HBV-related cirrhosis and non-AKI HBV-related liver failure patients.Conclusion:AKI patients with HBV-related liver failure,AKI patients with HBV-related cirrhosis usually accompany with the occurrence of infection.The injuries of renal tubules in AKI patients with HBV-related liver failure were much severer than that in AKI patients with HBV-related cirrhosis.The HDAC activity was aberrant and associated with the infection,liver and kidney injury severity.HDAC activity may be a predictive factor of AKI occurrence in HBV-related liver failure or cirrhosis patients,we can reduce the incidence of AKI and alleviate the kidney damage by inhibiting HDAC in AKI patients with HBV-related liver failure.Part II The protective effects of HDAC inhibitor Entinostat(MS-275)on kidney of endotoxin-induced acute liver failure with acute kidney injury miceObjective:Renal tubules injury and increased HDAC activity occur in AKI patients with HBV-related liver failure.Systemic inflammatory response plays an important role in the pathogenesis of AKI.This study built an endotoxin-induced acute liver failure with AKI mice model.Examined the occurrence of acute liver failure and AKI in model mice,intervened the model mice with selected classic I HDAC inhibitor,Entinostat(MS-275),to investigate the effects of MS-275 on kidney of endotoxin-induced acute liver failure with AKI mice.Methods:The endotoxin-induced acute liver failure with AKI mice models were induced by lipopolysaccharide(LPS).C57BL/6 mice were randomly divided into 3 groups:control group,model group,and MS-275 group.MS-275 was administered in a dose of 2 mg/kg/day for 5 consecutive days before models were built,while the control and model group were treated with normal saline at same amount.Observed 72-hour survival rate of mice after the models were built and collected the blood,liver and kidney of mice in 24 hours after the models were built.Detected renal function,histopathology changes of liver and kidney;measured expressions of HDAC1,HDAC3,histone H3,H4 acetylation levels and HDAC activity in kidney;detected release of inflammatory factors,expression and nuclear translocation of NF-κB p65,ROS production and markers of renal tubules injury in kidney.Results:The 72-hour survival rate in model mice was decreased to 37.5%,MS-275 enhanced the survival rate to 62.5%.Compared with the control group,histopathology of liver in most of the model mice presented as edema and necrosis of hepatocyte,disordered hepatic lobular structure and infiltration of inflammatory cells.Most of the mice in model group suffered from renal tubules injury with higher NGAL,KIM-1 levels and impaired renal function,even some of them had renal histopathology changes with swollen tubular epithelial cells,indistinct brush border and infiltration of inflammatory cells.So this model can be used to study the acute liver failure with AKI.MS-275 inhibited the expressions of HDAC1,HDAC3,decreased the HD AC activity and promoted histone H3,H4 acetylation levels in kidney;MS-275 alleviated the damage of kidney tubules,impaired kidney function,kidney and liver pathology injury;MS-275 suppressed the release of inflammatory factors,ROS production and the nuclear translocation of NF-κB p65 in kidney of endotoxin-induced acute liver failure with AKI mice.Conclusion:MS-275 has protective effects on kidney of endotoxin-induced acute liver failure with AKI mice through inhibiting inflammatory response and reducing ROS production.Part III The effects of HDAC inhibitor Entinostat(MS-275)on apoptosis and inflammation of endotoxin-induced HK-2 cellsObjective:The apoptosis of renal tubular epithelial cells occurs in endotoxin induced AKI.MS-275 can inhibit the inflammatory response and renal tubules injury in endotoxin-induced acute liver failure with AKI mice.In this study,we used MS-275 to intervene LPS-induced HK-2 cells to investigate the effects of MS-275 on apoptosis and inflammation of LPS-induced HK-2 cells.Methods:Cells were divided in control,model(LPS)and MS-275 group.The AKI renal tubules injury model was conducted by LPS stimulating HK-2 cells.MS-275 was administrated at 2 hours before models were built.The apoptosis of HK-2 cells were detected;the expressions of inflammatory factors,ROS production,antioxidants Nrf2,GSH,SOD activity,anti-apoptosis protein,Bcl-2 were detected;molecules in the apoptosis pathway of mitochondrion and endoplasmic reticulum(ER),Bax,caspase-3,CHOP and GRP78 in HK-2 cells were detected.Results:MS-275 inhibited the apoptosis of LPS-induced HK-2 cells;MS-275 increased the antioxidants Nrf2,SOD activity and GSH levels,enhanced the expression of anti-apoptosis protein,Bcl-2 levels;MS-275 inhibited release of inflammatory factors and activation of NF-κB p65;MS-275 suppressed Bax,caspase-3,CHOP and GRP78 levels in LPS-induced HK-2 cells.Conclusion:MS-275 has protective effects on LPS-induced HK-2 cells through inhibiting the inflammation,enhancing the production of antioxidants,decreasing oxidative stress and ER stress induced ap optosis.