蛋白激酶C和线粒体ATP敏感性钾通道在七氟醚预处理抗肠缺血／再灌注损伤中的作用

The role of protein kinase C and mito ATP-sensetive potassium channel in sevoflurane preconditioning against intestinalischemia/reperfusion injury in rats

目的探讨蛋白激酶c（protein kinaseC，PKC）和线粒体ATP敏感性钾通道（mito ATP-sensetive potassium channel，mKm）在0．5MAC七氟醚（1．15％）预处理减轻大鼠肠缺血，再灌注损伤（ischemia／reperfusion injury，I／RI）中的作用。方法45只大鼠按随机数字表法分成5组（每组9只）：①假手术对照组（Sham组）；②肠缺血，再灌注组（II／R组）；③0．5MAC七氟醚预理组（APC组）；④PKC非特异性阻滞剂Chelerythrine组（CHE组）：在0．5MAC七氟醚预处理前15min腹腔注射CHE5mg／kg；⑤mK。阻滞剂5-Hydroxydecanoic组（5HD组）：在0．5MAC七氟醚预处理前15min腹腔注射5-HD10mg／kg。再灌注120min时剪取小肠组织3份，分别行组织切片HE染色，在光镜下观察其结构病理变化程度及改良Chiu’s评分，免疫组织化学法检测含半胱氨酸的天冬氨酸蛋白水解酶3（cysteinyl aspartate specific proteinase-3，Caspase-3）活化表达，并用Westernblot方法检测肠组织B细胞淋巴瘤／白血病-2（Bcelllymphoma／lewkmia-2，Bcl-2）和Caspase-3活化蛋白表达的变化。结果APC组的Chiu＇s病理评分值4．0±1.3，显著性低于II／R组6．7±1．0、CHE组6．8±1．1及5HD组5．8±1．2（p〈0．01），而后3组差异无统计学意义（p-~o．05o与Sham组比较，其他各组的Bcl-2及Caspase-3表达量显著升高（P〈0．01，0．05）；与CHE组、5HD组及II／R组比较，APC组的Bcl-2增加而Caspase-3减少（P锄．01）；CHE组和5HD组与II／R组比较差异无统计学意义（P〉0．05）。结论0．5MAC七氟醚预处理减轻II／R组后肠组织的病理损害，提高Bcl-2和降低Caspase-3蛋白的表达，而阻滞剂CHE和5HD能消除此效应。0．5MAC七氟醚预处理的保护机制与PKC和mKATP通道的激活有关。

Objective To investigate the role of protein kinase C （PKC） and mito ATP-sensetive potassium channel（mKATP） channels in 0.5 MAC sevoflurane （1.15%） pretreatment preventing intestinal ischemia/reperfusion injury（I/RI）in rats. Methods 45 rats were randomly divided into five groups （n=9）： （1） Sham-operated group （Sham group）; （2） The intestinal ischemia/reperfusion group （II/R group）; （3） 0.5 MAC sevoflurane preconditioning group （APC group）; （4） PKC nonspecific blocker Chelerythrine group （CHE group）： intraperitoneal injection of CHE 5 mg/kg at 15 min before 0.5 MAC sevoflurane pretreatment; （5） mKATP channel blocker 5-Hydroxydecanoic group （5HD group）： intraperitoneal injection of 5HD 10 mg/kg at 15 min before 0.5 MAC sevoflurane pretreatment. Three pieces of intestinal tissues were used as following at 120 min after reperfusion： the pathological changes of intestinal tissues structure were observed by HE staining tissue sections and scored by improved Chiuls in the microscope. The expression of cysteinyl aspartate specific proteinase-3 （Caspase-3） was observed by immunohistochemistry. The expressions of B cell lymphoma/lewkmia-2 （Bcl-2） and Caspase-3 in intestinal tissues were detected by Western blotting. Results Chiu＇s pathological score in APC group （4.0±1.3） was significantly lower than that of II/R group （6.7±1.1）, CHE （6.8±1.1） and 5HD group （5.8±1.2） （P〈0.01）. The latter three groups showed no significant difference （P〉0.05）. Compared with sham group, Bcl-2 and Caspase-3 expression levels in the other four groups increased significantly （P〈0.01）. Compared with APC group, Bcl-2 expression levels inCHE group, 5HD group and II/R group decreased significantly（P〈0.05）. Compared with sham group, Caspase-3 expression levels in the other four groups increased significantly （P〈0.01）. The increased level of APC group is significantly less than CItE group, the 5HD group tI/R group （P〈0.01）, the latter three groups showed no significant difference （P〉0.05）. Conclusions 0.5 MAC sevoflurane pretreatment could protect against the intestinal II/R through activation of PKC and mKATp channel.