The Study Of Protective Effect And Mechanism Of Dexmedetomidine On Renal Ischemia-reperfusion Injury

Part I The protective effect of dexmedetomidine on renal ischemia and reperfusion injury in ratsObjective:To investigate the effect of dexmedetomidine on the apoptosis and oxidative stress of renal ischemia and reperfusion injury in rats.Methods:All of the rats studied were divided into three groups(eight rats in every group), randomly, sham-operated group, ischemia and reperfusion injury (IRI) group and dexmedetomidine(DEX) group.30 minutes before ischemia, 100ug/kg dose of DEX was injected into each peritoneal cavity of the DEX group rats.Meanwhile,the ischemia and reperfusion injury group only recevied an injection of same amount of saline.After reperfusion 24h, Blood Urea Nitrogen(BUN) and creatine(Cr) were detected. Also, HE staining, SOD, MDA, Caspase-3 immunohistochemistry were performed. Western blot was used to detect BAX, AIF and Bcl-2expression levels.Results:The results indicated that renal functions (BUN and Cr)were highly significant after IRI,compared with sham-operated group,and the statistical difference was significant(P<0.05). From HE staining, the injury was very obvious in IRI group. SOD expression was downregulated in IRI group compared with sham-operated group,and the statistical difference was significant (P<0.05). The level of MDA was upregulated in IRI group compared with sham-operated group,and the statistical difference was significant (P<0.05).From Caspase-3 immunohistochemistry,the apoptosis was very obvious in IRI group.The expression of apoptotic factors(BAX and AIF)were upregulated in IRI group,while,The expression of anti-apoptotic factors(BCL-2)was downregulated in IRI group. However, DEX could inhibit these changes caused by IRI.Compared with IRI,SOD expression was upregulated in DEX group,BUN,Cr,the level of MDA were downregulated in DEX group,the statistical differences were significant (P<0.05).The expression of apoptotic factors(BAX and AIF)and apoptotic protein (Caspase-3) were downregulated in DEX group,while,The expression of anti-apoptotic factors(BCL-2)was upregulated in DEX group.Conclusions:DEX preconditioning had protective effect on the apoptosis and oxidative stress of renal ischemia and reperfusion injury in rats. Part Ⅱ The protective effect of dexmedetomidine oninflammation caused by renal ischemia and reperfusion injury in ratsObjective:To investigate the effect of dexmedetomidine on the inflammation of renal ischemia and reperfusion injury in rats.Methods:All of the rats studied were divided into three groups(eight rats in each group), randomly, sham-operated group, ischemia and reperfusion injury (IRI) group and dexmedetomidine(DEX) group.30 minutes before ischemia, 100ug/kg dose of DEX was injected into each peritoneal cavity of the DEX group rats.Meanwhile,the ischemia and reperfusion injury group only recevied an injection of same amount of saline.After reperfusion 8 weeks, Blood Urea Nitrogen(BUN) and creatine(Cr) were detected. Also, HE staining was performed. Realtime PCR was used to detect the mRNA levels of TNF-α,IL-1β, ICAM-1. Western blot was used to detect HMGB1 and TLR4 expression levels.Results:The results indicated that renal function had no significant differences among groups. From HE staining, the injury were very obvious in IRI group. Also, the mRNA levels of TNF-α,IL-1β, ICAM-1 were up regulated in IRI group compared with sham-operated group, as well as the expression of HMGB1 and TLR4. Compared with the sham-operated group,the statistical differences of the mRNA levels of TNF-α,IL-1β,ICAM-1 were significant (P<0.05). However, DEX could inhibit these changes caused by IRI.The mRNA levels of TNF-α,IL-1β, ICAM-1 were up downrgulated in DEX group compared with IRI group, as well as the expression of HMGB1 and TLR4. Compared with the IRI group,the statistical differences of the mRNA levels of TNF-α,IL-1β, ICAM-1 were significant (P<0.05).Conclusions:DEX had protective effect on chronic inflammation after renal ischemia and reperfusion injury in rats.Part Ⅲ The protective effect of dexmedetomidine on proximal tubular cells mimicking ischemia and reperfusion injury in vitroObjectives:The majority renal IRI studies has been based on animal models. To investigate the DEX on the IRI in vitro, we set up an in vitro model of IRI using the rat proximal tubule cell line NRK-52E and applied different concentrations (0.01ng/ml,0.1ng/ml,1ng/ml) DEX on it.Methods:The cells were randomly divided into three groups, control group, ischemia reperfusion group, dexmedetomidine pretreatment group.After placing the cells in ischemic condition for 3h, then put in in normal condition to mimic reperfusion process for 24h. In the dexmedetomidine pretreatment group,different concentrations(0.01ng/ml,0.1ng/ml,1ng/ml)DEX had been applied on the cells for 1h,next steps as the ischemia reperfusion group.Flow cytometry was used to access apoptosis. The SOD and MDA levels were also detected. The expression of PARP-1 and AIF were analyzed.Results:Our data showed that apoptosis was obviously found in mimic IRI group, and DEX could alleviate the apoptosis ratio caused by mimicking IRI.The apoptosis ratios were 9.0%(control group).55.0%(ischemia reperfusion group),49.1%(0.01ng/ml dexmedetomidine pretreatment group),28.6%(0.1ng/ml dexmedetomidine pretreatment group),22.1%(1ng/ml dexmedetomidine pretreatment group). The SOD level was decreased in mimic IRI group compared with DEX group, and MDA level was opposite. The increased expression of PAPR-1 and AIF could be found in mimic IRI group. However, DEX could inhibit these protein expression.Conclusions:DEX could protect the NRK-52E cells from mimicking IRI through decreasing oxidative stress and downregulated apoptotic protein.