ERK1/2在缺血再灌注损伤肺细胞凋亡中的作用及缺血后处理的干预

Effects of ERK1/2 on pneumocyte apoptosis after lung ischemia/reperfusion injury and ischemic postconditioning intervention

目的:探究细胞外信号调节激酶1/2(ERK1/2)在缺血/再灌注(IR)损伤肺细胞凋亡中的作用及缺血后处理的干预。方法:雄性SD大鼠,随机分成5组(n=8),即对照组(C组)、肺缺血/再灌注组(IR组)、肺缺血/再灌注+缺血后处理组(IPO组)、缺血后处理+溶剂对照组(D组)、缺血后处理+U0126组(U组)。分别于再灌注2 h留取左肺组织,电镜观察肺组织超微结构改变;原位末端标记(TUNEL)法检测肺细胞凋亡情况并计算凋亡指数(AI);RT-PCR法、免疫组化法测定肺组织Bax、Bcl-2基因和蛋白的表达。结果:与C组相比,IR组肺组织AI、Bax基因及蛋白表达均显著升高(P<0.05),电镜下肺细胞均发生明显损伤;Bcl-2、Bcl-2/Bax基因及蛋白表达显著降低(P<0.05);IPO组、D组、U组与IR组相比,肺组织AI、Bax基因及蛋白表达均显著降低(P<0.05),电镜下肺细胞损伤情况有所改善;Bcl-2、Bcl-2/Bax显著升高;D组与IPO组比较各项指标差异均无统计学意义(均P>0.05),U组与IPO组相比,肺组织AI值显著升高(P<0.05),电镜下肺上皮细胞超微结构破坏较严重,胞内细胞器不完整;Bcl-2基因及蛋白表达明显降低,Bax基因及蛋白表达明显升高,Bcl-2/Bax显著降低。结论:IR抑制了MAPK家族中的ERK1/2激活,导致大鼠肺组织结构严重破坏,细胞大量凋亡;IPO可以通过激活ERK1/2通路,改善其结构破坏和细胞凋亡。

Objective： To investigate the role of ERK1/2 on pneumocyte apoptosis after lung ischemia/reperfusion injury and ischemic postconditioning （IPO） intervention. Methods： Forty adult male Sprague-Dawley rats were randomly divided into 5 groups based upon the intervention （n=8）： control group （C）, IR group （IR）, IR＋IPO group （IPO）, IPO＋solution countrol group （D）, IPO＋U0126 group （U）. Left lung tissue was isolated after the 2 hours of reperfusion, The ultrastructure of the left lung were observed under electron transmission microscopes. Apoptosis index （AI） of lung tissue was determined by terminal deoxynuleotidyl transferase mediated dUTP nick end and labeling （TUNEL） method. The mRNA expression and protein levels 0f Bcl-2 and Bax were measured by RT-PCR and quantitative immunohistochemistry （IHC）. Results： Compared with C group, AI and the expression of Bax of IR were significantly increased, the expression of Bcl-2 and Bcl-2/Bax were significantly decreased （P〈 0.05）, and ultrastructure abnormality was obviously found in lung tissue. Compared with IR group, all the indexes of IPO except for the expression of Bcl-2 and Bcl-2/Bax were obviously reduced, the expression of Bcl-2 and Bcl- 2/Bax. was increased （P〈0.05）. All the indexes between D and IPO were little or no significant （P〉0.05）. the expression of Bcl-2 and Bcl-2/Bax of U was significantly decreased and other indexes were increased than those of IPO （P〈0.05）. Conclusion： IPO may attenuate pneumocyte apoptosis in LIRI by activation of ERKI/2 MAPK, up-regulating expression of Bcl-2/Bax ratio.