| Objective:1.The establishment of an adult rat model of renal ischemia-reperfusion injury.2.To investigate the role and mechanisms of Fas and Caspase-3 in the(adult rat) renal ischemia-reperfusion injury in apoptosis of renal tubular epithelial cell.3.To investigate the effects of medical(Ketamine) postconditionings with different doses on rat renal ischemia-reperfusion injury and its molecular mechanism.4.To investigate the effects of remote(ischemic) postconditionings and the ischemic postconditionings on the renal ischemia-reperfusion injury in rats and its molecular mechanism.Methods:Thirty-six male Wistar rats(10-week-old healthy adult rats) were randomly divided on average into six groups,each with six:sham operation(S) group,ischemia-reperfusion(I/R) group,the left hind limb ischemic postconditioning(P1) group,renal ischemic postconditioning(P2) group,ketamine postconditioning group1(K1),ketamine postconditioning group 2(K2).In the model of renal ischemia,rats were subjected to unilateral ischemia by clamping of the left renal artery for 45 minutes followed by 6 hours of reperfusion,using a nontraumatic vascular clip,and the right kidneys were removed just after the beginning of the reperfusion of left kidneys.In group S,rats were subjected to the same surgical procedure without clamping the left renal artery.Group P1:One minute before the reperfusion following 45 minutes ischemia in the left kidney,the left femoral arteries have been clipped for 5 minutes;Group P2:these left Kidneys were subjected to six cycles of 10 seconds of reperfusion followed by 10 seconds ischemia immediately after 45 min of ischemia.In group K1 and K2,2mg/Kg or 10mg/Kg Ketamine were injected via the tail vein 5 min before the reperfusion.The rats were killed after 6 h of reperfusion,and blood samples were collected from right atrium,for measurement of creatinine(Cr) and blood urea nitrogen(BUN).The renal expression of Fas, Caspase-3 stained by immunohistochemistry and the apoptotic cell stained by terminal transferase-mediated dUTP nick-end labeling(TUNEL) were detected through a microscope.Pathological changes of renal tissue were observed under light microscope and electron microscope.The positive cells of Fas,Caspase-3,and apoptosis in tubular epithelial cell were calculated for positive index(PI) and apoptosis index(AI).SPSS10.0 software was used to analyze the data,a=0.05 was considered the level of significance.Results:(1) Compared with the sham-operated group,the levels of serum BUN and Cr in other five groups were significantly increased after renal I/R injury (P＜0.01).The kidneys of model rats in the five groups displayed significant pathologic changes and the expression of Caspase-3 and Fas in kidney cells increased,a large number tubular epithelial cell degeneration,necrosis and apoptosis compared with the sham-operated groups(P＜0.01).(2) Fas and Caspase-3 were significantly increased in the ischemic group at 6 h of reperfusion compared with the sham-operated groups(P＜0.01).Histological sections of kidneys from ischemia/reperfusion rats showed extensive tubular necrosis,mainly located at the proximal tubules.(3) The pathological changes and apoptosis in both postconditioning groups and both treatment groups were improved and all of the renal functions were attenuated(P＜0.01).Compared with group I/R,the other four groups in serum BUN,Cr concentrations decreased(P＜0.01),and renal tissue Fas, Caspase-3 positive index and the apoptosis index(P＜0.01).(4) Compared with P1, P2 rat serum BUN,Cr concentrations decreased(P＜0.05),and renal tissue Fas, Caspase-3 positive index and the apoptosis index(P＜0.05).Compared with K1,K2 rat serum BUN,Cr concentrations decreased(P＜0.01),and renal tissue Fas, Caspase-3 positive index and the apoptosis index(P＜0.01).Conclusions:The apoptosis of tubular epithelial cell in the kidneys of I/R injury model rats result in renal dysfunction.Fas,as the representatives of apoptotic pathway,were activated,leading to renal tubular epithelial cell apoptosis.Both of the ketamine treatment can inhibit the apoptosis in tubular epithelial cells and attenuate the I/R injury.The actions of ketamine(10mg/Kg) treatment(K2) groups are more significant.Both of the ischemic postconditionings can inhibit the apoptosis in tubular epithelial cells and attenuate the I/R injury.The actions of renal ischemic postconditionings groups are more significant.
|
PDF Full Text Request