半胱氨酸蛋白酶抑制剂C对局灶性脑缺血再灌注大鼠大脑皮质caspase-9、脑红蛋白表达和过氧化损伤的影响*

目的：探讨不同浓度半胱氨酸蛋白酶抑制剂C（CysC）对大鼠损伤大脑皮质组织caspase-9 和脑红蛋白（NGB）的表达、氧化损伤和脑组织形态学的影响。方法：大鼠按照随机数字表分为假手术组（Sham 组）、缺血再灌注组（I/R 组）、CysC 低、中、高浓度组。线栓法制备右侧局灶性脑缺血再灌注大鼠模型，缺血2h 再灌注24 h 后行H-E染色，观察脑组织形态学变化；免疫组织化学法检测caspase-9 的阳性细胞数；免疫组织荧光法检测皮质神经元NGB阳性细胞数；免疫印迹检测脑皮质组织中caspase-9 蛋白的表达。羟胺法检测超氧化物歧化酶（SOD）活力，化学比色法检测谷胱甘肽过氧化物酶（GSH-PX）活力，硫代巴比妥酸法检测丙二醛（MDA）含量。结果：与I/R 组相比，CysC 低、CysC 中浓度组caspase-9 表达较I/R 组明显减少；而CysC 高浓度组caspase-9 的表达明显升高，与I/R 无明显差别。CysC 低、CysC 中浓度组caspase-9 阳性细胞数较I/R 组明显减少，NGB阳性细胞数较I/R组显著增多，SOD和GSH-PX活力升高，MDA含量下降；而CysC 高浓度组caspase-9 阳性细胞数增多，NGB阳性细胞数减少，SOD和GSH-PX活力降低，MDA含量升高；CysC 高浓度组与I/R 组相比，差异无统计学意义。结论：CysC 低、中浓度可能通过上调NGB的表达，下调caspase-9 的表达，减轻脑缺血再灌注后的自由基损伤。而浓度过高则作用相反。

Effects of cystatin C pretreatment on expression of caspase-9,neuroglobin and peroxide in damage cerebral cortex of rats with cerebral ischemia and reperfusion

Objective To study the effect of different concentrations of cystatin C （CysC）pretreatment on theexpression of caspase-9 and neurolobin （NGB）， oxidative stress and pathomorphology in rats with focal cerebralischemia reperfusion injury. Methods Male SD rats were randomly divided into sham operation group（ Sham group），cerebral ischemia reperfusion injury group（I/R group） and cystatin C different concentration groups（the groupsof low， middle and high respectively）. The model of cerebral ischemia reperfusion injury was established by suturemethod. After ischemia for 2 h and reperfusion for 24 h， the pathologic change of the brain was observed by H-Estaining under the microscope. The number of caspase-9 and NGB positive cells were detected by immunohistochemicalSP method and immunofluorescence technology. Western blotting was used to detect the expression of caspase-9protein. Hydroxylamine method was used to detect activity of SOD， chemical colorimetry method was used to measureactivity of GSH-PX， and TBA method was used to detect the content of MDA. Results Compared with I/R group，the expression of caspase-9 had different degrees of decrease in CysC low and middle group， while the expression ofcaspase-9 obviously increased in CysC high group. The change of caspase-9 positive cells was consistent with the resultof Western blotting. The number of NGB positive cells in CysC low and middle group was significantly higher thanthat in I/R group， the activity of SOD and GSH-PX wasall enhanced but the content of MDA was reduced. Thenumber of NGB positive cells in CysC high group wassignificantly lower. The activity of SOD and GSH-PXwas weakened， and the content of MDA was increased. There was no significant difference between CysC high groupand I/R group. Conclusion CysC low and middle concentration groups may reduce free radical damage after cerebralischemia reperfusion by increasing NGB expression and reducing caspase-9 expression. Overconcentration has theopposite effect.