Protection Effect And Its Mechanism Of Myricetin In Renal Ischemia-reperfusion Injury

Objective:Acute kidney injury(AKI)is a common clinical syndrome with a high mortality rate,and renal ischemia-reperfusion(IR)injury is the main cause of acute kidney injury.The pathophysiology of acute kidney injury is complex and there is no effective treatment strategy for acute kidney injury.Therefore,it is crucial to explore new drugs to prevent and treat acute kidney injury.Myricetin(Myr)is widely found in many natural plants and has been shown in previous studies to reduce ischemia-reperfusion injury in the brain,heart and intestine.In this study,we investigated the protective effect of Myricetin on renal ischemia-reperfusion injury in mice,and conducted a preliminary in vivo and in vitro study to investigate the mechanisms involved.Methods:1.In vivo experiment: 15 healthy male C57BL/6 mice(8-10 weeks old)were randomly divided into three groups of five mice each: sham-operated group(Sham group),renal ischemia-reperfusion injury group(RIRI group),and renal ischemia-reperfusion injury + Myr pretreatment group(RIRI+Myr group).Renal ischemia-reperfusion injury(RIRI)+ Myr pretreatment group(RIRI+Myr group): Mice were injected intraperitoneally with 25 mg/kg of Myr 30 minutes before surgery,and then bilateral renal vessels were blocked to make bilateral kidneys ischemic for 30 minutes,after which the bilateral renal vessels were unclamped and reperfused for 24 hours.Renal ischemia-reperfusion injury group(RIRI group): mice were injected intraperitoneally with an equal amount of saline 30 minutes before surgery,followed by ischemia-reperfusion.Sham surgery group(Sham group): mice were injected with equal amount of saline intraperitoneally 30 minutes before surgery,only opening and closing of the peritoneum was done without clamping the renal hilum,and the incision was sutured after exposing the abdominal cavity for 30 min.(1)To detect the effects of Myr pretreatment on blood creatinine and blood urea nitrogen in mice,and HE staining to observe the changes in kidney histology.(2)The expression of phosphorylated receptor interaction protein 3(p-RIPK3)and phosphorylated mixed-lineage kinase structural domain-like protein(p-MLKL)in kidney tissues was detected by Western Blot and immunohistochemistry to assess the effect of Myr on the anti-necroptosis aspects of renal ischemia-reperfusion injury in mice.2.In vitro experiments.(1)The effect of myricetin on the activity of mouse renal tubular epithelial cells(m RTEC)was examined by CCK8 assay(2)Induction of necroptosis in m RTEC cells,pretreatment of m RTEC cells with Myr and detection of intracellular p-RIPK3 expression by Western Blot.Results:1.Myr pretreatment significantly reduced serum creatinine and blood urea nitrogen levels in mice after renal ischemia-reperfusion injury,compared with the RIRI group.2.Myr pretreatment significantly reduced renal histopathological injury and reduced tubular injury scores compared to the RIRI group.3.Myr pretreatment significantly reduced the expression of p-RIPK3 and p-MLKL in renal tissues compared to the RIRI group.4.Myr pretreatment significantly reduced the expression of intracellular p-RIPK3 compared to m RTEC cells in the necroptosis group Conclusion:1.Myricetin pretreatment had a significant protective effect against renal ischemiareperfusion injury in mice.2.Myricetin pretreatment attenuated renal ischemia-reperfusion injury by inhibiting the necroptosis pathway,which may be achieved by downregulating p-RIPK3,p-MLKL.