缺血后调适对大鼠骨胳肌缺血再灌注损伤的作用及其机制研究

The effect and mechanism of ischemic post-conditioning on skeletal muscle ischemia-reperfusion injury in rats

目的:探讨缺血后调适对大鼠骨胳肌缺血再灌注损伤(IRI)的作用及其机制。方法:随机将18只大鼠分为3组:假手术组(Sham组)、缺血再灌注组(IR组)、缺血后调适组(IPoC组),每组6只。各组均分离股动脉(右侧),IR组及IPoC组均施以4h缺血,缺血后IR组直接进行再灌注,IPoC组则行4个循环30s再灌注/30s缺血操作后再行灌注,Sham组仅分离股动脉无缺血再灌注操作。运用pH仪对缺血期、调适期、再灌注期进行测量;并于再灌注2h后取大鼠胫前肌行western blotting检测RISK通路中相关蛋白(Akt、Erk1/2及其磷酸化激活物p-Akt、p-Erk1/2)的表达情况;运用分光光度计监测Ca^(2+)诱发的线粒体通透性转换孔(MPTP)开放情况;测定24h后腓肠肌湿/干重(W/D)比值。结果:(1)IPoC组在调适初期2.67min内能维持组织内酸性环境,pH均值为(6.81±0.133)。(2)与Sham组比较,IPoC组的p-Akt/t-Akt、p-Erk1/2/t-Erk1/2比值均明显增加(均P<0.01),IR组的p-Akt/t-Akt、p-Erk1/2/t-Erk1/2比值则明显降低(均P<0.01)。(3)在2 min、4 min时,IPoC组MPTP开放数低于其他两组(均P<0.05)。(4)IPoC组W/D比值明显低于IR组(P<0.05),而Sham组与IPoC组W/D比值比较差异无统计学意义(P>0.05)。结论:缺血后调适能有效减轻大鼠骨骼肌缺血再灌注所致的肌肉水肿,其机制可能是通过维持再灌注初期组织酸环境,激活RISK通路而对再灌注骨骼肌起到保护作用。

Objective：To investigate the effect and mechanism of ischemic post-conditioning （IPoC） on skeletal muscle isehemia-reperfusion （SMIR） injury in rats. Methods： 18 rats were randomly divided into 3 groups （ n ：6 in each group）： sham group, SMIR group and IPoC group. The rats in the SMIR group were subjected to 4 hours ischemia of the right hind limb, followed by reperfusion. Rats in IPoC group were subjected to right hind limb occasion for 4 hours, followed by four cycles of 60-s intervals of ischemia and reperfusion starting at 30 s after the initial reperfusion. The expressions of RISK signaling pathway re- lated proteins such as Akt, Erkl/2, phosphorylated （p）-Akt and p-Erkl/2 were detected by western blotting. The opening of mitochondrial permeability transition pore （MPTP） was observed by UV spectrophotometer. The ratio of wet/dry weight （W/D） of gastrocnemius was measured after 24 h. Results： （1） IPoC could maintained the acidic environment in the tissues within 2.67 rain, and the pH value was （6.81± 0. 133） （ P 〈0.01）. （2） The ratios of p-Akt/t-Akt and p-Erkl/2/t-Erkl/2 in IPoC group were increased, while were decreased in the SMIR group compared with the sham group （ P 〈0. 01）. （3） At 2 and 4 min, the opening of MPTP in IPoC group was increased than that in the other two groups （ P 〈0.05）. （4） The W/D ratio of IPoC group was significantly lower than that of SMIR group （ P 〈0.05）, but there was no significant difference between sham group and IPoC group （ P 〉0.05）. Conclusion： IPoC could effectively alleviate SMIR injury in rats, and the mechanisms might be related to maintaining acid environment in initial reperfusion and activating the RISK signaling pathway.