| Objective:To investigate the effect of Normobaric hyperoxia on renal injury induced by ischemia-reperfusion in rats and its related mechanism.Methods:Twenty-four adult male SD rats(Purchased from Liaoning Changsheng Biotechnology Co.,Ltd.)were enrolled and their left kidney was excised to establish a solitary kidney model.After one week,they were randomly assigned to 4 groups,6rats in each group,namely sham-operated group(Group S),control group(Group C)experimental group(Group E)and Zn PP+Group E.In Group S,only the right renal pedicle was bluntly separated,but no ischemic treatment was given.Both Group C,Group E and Zn PP+Group E suffered from right renal ischemia for 40 minutes by clamping renal pedicle.After 24 hours,the four groups were all placed in a sealed oxygen chamber.Rats in Groups S and Group C inhaled normal concentration oxygen(21%),while rats in Group E and Zn PP+Group E inhaled high concentration oxygen(50%-55%),once a day for 2 hours each time for 7 days.The rats in the Zn PP+Group E were intraperitoneally injected with zinc protoporphyrin(Zn PP),a specific inhibitor of HO-1,before inhaling high concentrations of oxygen.On the 8th day after the operation,by measuring the weight of the rat,dynamically monitoring the renal function of the rat,and the degree of pathological damage in the rat kidney group was evaluated by HE staining and the apoptosis of rat kidney tissue was evaluated by Tunel method to determine the protective effect of Normobaric hyperoxia on rat renal ischemia-reperfusion injury.The expression of Nrf2 and HO-1 was studied by RT-PCR,Western blotting and immunohistochemical analysis.And detecting the oxidative stress markers in the kidney tissue.In order to further determine whether HO-1 induced by Nrf2 is involved in the anti-oxidative stress damage,We established the Zn PP+E group by intraperitoneal injection of HO-1 specific inhibitor zinc protoporphyrin(Zn PP)(45umol/Kg)after rat kidney ischemia-reperfusion injury and before inhaling normobaric hyperoxia,the postoperative treatment measures were the same as those in group E.On the 8th day after operation,the m RNA and protein expression levels of the target gene in the Zn PP+E group were analyzed by RT-PCR and Western blotting;HE staining and Tunel detection method were used to analyze the degree of renal tissue pathological damage and the changes of renal tubular epithelial cell apoptosis.Results:Four was no significant difference in BUN and Cr levels in the three groups before ischemia(P>0.05).On the 8th day after operation,there was no significant change in BUN and Cr in group S rats;The BUN and Cr of rats in group C and E gradually increased;while the BUN and Cr of rats in group E were significantly lower than those in group C(P<0.05).On the 8th day after operation,the rats in Group S weighed(224.33±5.43 g),and Group C(209.5±4.09 g)and Group E(221±2.53 g)were lighter,but the rats in Group E weighed more than those in Group C(P<0.05).The rats in Group S MDA contented(5.53±0.38umol/L),and Group C(15.20±0.84umol/L)and Group E(8.33±0.33 umol/L)were lighter,but MDA content of Group E was significantly lower than that of Group C(p < 0.05).The rats in Group S SOD contented(14.20±0.52nmol/L),and Group C(5.72±0.39nmol/L)and Group E(10.71±0.65 nmol/L)were lower,but the rats in Group E SOD contents more than those in Group C(p < 0.05).Compared with group S,Keap1 m RNA transcription level and protein expression in group C and group E decreased,and compared with group C,group E decreased the most significantly(P<0.05);However,compared with group S,Nrf2 and HO-1 m RNA transcription level,protein expression and A value were increased in group C and E(P<0.05),but the increase in group E was the most significant(P<0.05);Microscopically,the renal tubular injury score of rats in Group S was 7.5±2.25,while that in Group C(74.33±5.16)and Group E(40.83±5.04)were significantly more,but that in Group E was less than that in Group C(P < 0.05).The Apoptosis index(AI)of renal tubular epithelial cells in renal cortex of Group S was(0.74±0.20),which was less than that of Group C(12.18±1.82)and Group E(3.96±0.99).While AI of Group E was significantly lower than that of Group C(p <0.05).After applying the HO-1 specific inhibitor Zn PP,Compared with group E,renal tubular injury score and renal tubular epithelial cell apoptosis level increased in group Zn PP+E;Keap1 gene m RNA and protein expression levels increase,Nrf2 and HO-1m RNA and protein expression levels decrease,and the protective effect of normobaric hyperoxia is reversed.Conclusions:Normobaric hyperoxia can alleviate renal injury induced by ischemia-reperfusion,and its mechanism may be related to activation of the Nrf2/HO-1 signaling pathway. |
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