Effect And Mechanism Of Empagliflozin On Contrast Media-associated Acute Kidney Injury

Objective: To investigate the effect and mechanism of empagliflozin on contrast-associated acute kidney injury.Methods: This study was divided into two parts: the experiment on SD rats and the experiment on mice and cells.Male SD rats were randomly divided into control group,empagliflozin group,contrast group,contrast plus empagliflozin group.The control group was given normal saline(1.5mg/kg)for7 consecutive days.Empagliflozin group was given empagliflozin(1.5mg/kg)for 7 consecutive days.The contrast group was deprived of water for 16 hours,and CA-AKI model was established.Contrast plus empagliflozin group were given empagliflozin(1.5mg/kg)intragastric administration for 7 consecutive days.After 16 h of water prohibition,the remaining treatment was the same as that of contrast group.At the same time,the patients were fed enough food and water,and were euthanized 12 hours after injection of iohexol to obtain both kidneys and blood.Serum renal function changes were detected by ELISA and HE staining was used to evaluate the pathological injury of the kidney.Western Blotting and immunohistochemistry were used to detect the expression of apoptosis-related protein,and TUNEL staining was used to evaluate the apoptosis of kidney cells.To further determine whether UCP2 was involved in the inhibition of oxidative stress and apoptosis by empagliflozin.Male C57BL/6J mice were divided into control group,contrast group,contrast plus normal saline group,contrast plus empagliflozin group.The contrast group was established CA-AKI model.Contrast plus normal saline groups and contrast plus empagliflozin groups were given normal saline(1.5mg/kg,)and empagliflozin(1.5mg/kg)were given intragastric administration for 7 days,and water was forbidden for 18 hours,and the rest treatment was the same as the contrast group.Mice in the control group were not treated with dehydration,furosemide,iohexol and empagliflozin.Blood and kidneys were collected after12 hours of iohexol injection.Renal function and glomerular pathological injury were evaluated by ELISA and HE staining.Renal apoptosis was evaluated by TUNEL and IHC.Mitochondrial structure was evaluated by electron microscopy.Human renal tubular epithelial cells(HK-2)were divided into control group,contrast group,contrast plus DMSO group,contrast plus empagliflozin group.The contrast group was treated with high glucose medium and iohexol(80mg I/ml),contrast plus DMSO group were treated with high glucose medium +DMSO,contrast plus empagliflozin group were pretreated with empagliflozin,and then treated with high glucose medium and iohexol(80mg I/ml).Cell vitality testing by CCK8 after 2 hours.The mitochondrial membrane potential and oxidative stress response were evaluated by JC-1 and ROS fluorescence staining.Western Blotting was used to detect the expression of proteins related to mitochondrial fusion and division.In the UCP2 knockout mice experiment: UCP2-/-mice were randomly divided into control group,contrast group,contrast plus normal saline group,contrast plus empagliflozin group,and the residual treatment was the same as that of C57 mice experiment.Results: Renal function,glomerular injury and cell apoptosis were aggravation in contrast model.The renal function,glomerular pathological injury and cell apoptosis were significantly relieved in Empagliflozin preconditioning group.The pretreatment of HK-2 by empagliflozin can repair the mitochondrial membrane potential of HK-2 induced by contrast agent and inhibit oxidative stress.In UCP2-/-mice pretreated with empagliflozin,kidney damage caused by contrast agent could not be alleviated,and mitochondrial structural and functional changes could not be repaired.Conclusion: Empagliflozin inhibits oxidative stress and apoptosis through UCP2 pathway,thereby alleviating the occurrence and development of contrast-associated acute kidney injury.