脑缺血再灌注引起大鼠脑细胞水肿及脑组织超氧化物歧化酶活性的变化

背景脑缺血再灌注产生的自由基主要是黄嘌呤氧化酶,导致梗死灶容积细胞肿胀.目的观察脑缺血再灌注引起脑自由基清除酶超氧化物歧化酶活性的变化,探讨嘌呤氧化酶抑制剂别嘌呤醇对缺血再灌注脑组织细胞含水量的影响.设计完全随机对照实验.单位沈阳医学院附属中心医院神经内科,中国医科大学附属第一医院神经外科,辽宁省肢体伤残矫形医院.材料实验于2003-05/2004-04在沈阳医学院附属中心医院实验室完成.选择Wistar大鼠40只,随机分为4组,每组10只.缺血+别嘌呤醇组脑缺血6 h,灌胃给予100mg/kg别嘌呤醇;缺血+悬浊剂组脑缺血6 h,灌胃给予相同剂量的悬浊剂(恶喹酸溶液);缺血再灌注+别嘌呤醇组脑缺血6 h,再灌注2 h,灌胃给予100 mg/kg别嘌呤醇;缺血再灌注+悬浊剂组脑缺血6 h,再灌注2 h,灌胃给予相同剂量的悬浊剂.其中别嘌呤醇组于脑缺血前48 h,24h和1 h共3次分别以100 mg/kg剂量灌胃给予别嘌呤醇.悬浊剂组同样方法给予悬浊剂.方法缺血+别嘌呤醇组、缺血+悬浊剂组的大鼠于闭塞后6 h,缺血再灌注+别嘌呤醇组、缺血再灌注+悬浊剂组于灌注后2 h测量脑组织含水量.脑组织过氧化物歧化酶分布采用过氧化物歧化酶免疫染色观察.主要观察指标①脑组织过氧化物歧化酶分布.②各组大鼠脑组织含水量.结果40只大鼠全部进入结果分析.①脑组织过氧化物歧化酶分布缺血+悬浊剂组、缺血再灌注+悬浊剂组铜锌超氧化物歧化酶染色,缺血灶内可见全部明显的染色增强.缺血+悬浊剂组的锰过氧化物歧化酶染色、缺血灶内可见血管周围轮状染色增强.同时可见染色增强的血管壁和神经细胞.缺血再灌注+悬浊剂组缺血灶内染色呈现弥漫性、稍低下.缺血+别嘌呤醇组和缺血再灌注+别嘌呤醇组铜锌过氧化物歧化酶都没有变化、在缺血+别嘌呤醇组可见血管周围染色增强,缺血再灌注+别嘌呤醇组未见弥漫性改变.缺血+悬浊剂组、缺血再灌注+悬浊剂组小动脉内皮细胞核肿大、中层肌细胞膨大、血管膜扩大,脑组织血管周围显著呈海绵状.缺血+别嘌呤醇组、缺血再灌注+别嘌呤醇组这些变化减轻.②各组大鼠脑组织含水量缺血+别嘌呤醇组,缺血再灌注+别嘌呤醇组低于缺血+悬浊剂组和缺血再灌注+悬浊剂组(78.56±0.30)%,(79.08±0.33)%;(78.85±0.49)%,(79.86±0.49)%,(P＜0.05)].结论别嘌呤醇可通过对过氧化物歧化酶的抑制有效减轻脑缺血再灌注后的组织损伤.

Ischemia/reperfusion-induced brain edema and changes of superoxide dismutase activity in rat brain tissue

BACKGROUND: The free radicals induced by cerebral ischemia/reperfusion consist mainly of xanthine oxidase, which induces cell swelling in the infarcted area.OBJECTIVE: To observe the changes of cerebral ischemia/reperfusioninduced changes in the activity of cerebral superoxide dismutase (SOD), an enzyme responsible for free radical clearance, and investigate the effect of apurin, a inhibitor of purine oxidase, on cellular water content in the brain tissue with ischemia/reperfusion injury.DESIGN: Completely randomized controlled study.SETTING: Department of Neurology of the Central Hospital Affiliated to Shenyang Medical College, Department of Neurosurgery of the First Hospital Affiliated to China Medical University, and Liaoning Provincial Orthopedic Hospital for Limb Disabilities.MATERIALS: The experiment was conducted in the Laboratory of the Central Hospital Affiliated to Shenyang Medical College from May 2003 to April 2004. Forty Wistar rats were subjected to a 6-hour cerebral ischemia and randomized into 4 equal groups to receive intragastric administration of 100 mg/kg apurin (ischemia + apurin group), oxolinic acid suspension of the same dose (ischemia+ oxolinic acid group), 100 mg/kg apurin after a 2-hour reperfusion (Ischemia/reperfusion + apurin group), or oxolinic acid of the same dosage after the 2-hour reperfusion (ischemia/reperfusion + oxolinic acid goup), respectively. The rats in apurin group had intragastric administration of 100 mg/kg apurin 48, 24 and 1 hour before occlusion of the cervical internal carotid artery (CICA) to induce the ischemia, respectively. Oxolinic acid was given in the two oxolinic acid groups in the same manner.METHODS:Water content of brain tissue of rats was measured after 6 hours of CICA occlusion in the two ischemia groups and after the 2-hour perfusion in the two ischemia/reperfusion groups. Distribution of SOD in the brain tissue was observed with SOD immunostaining.MAIN OUTCOME MEASURES: Distribution of SOD and water content in the brain tissue of rats.RESULTS: In the two oxolinic acid groups, Cu-Zn SOD staining identified obviously increased staining intensity in the ischemic foci. Mn SOD staining in ischemia+oxolinic acid group resulted in increased circular staining surrounding the vessels in the ischemic foci, with also obvious staining of the vascular wall and neural cells. The ischemic foci of the ischemia/reperfusion + oxolinic acid group showed diffuse but lightly weaker staining. Cu-Zn SOD staining in the two apurin groups revealed no significant difference. In the two oxolinic acid groups, endothelial cell nuclear swelling of the arteriole, protrusion of the mid-layer myocytes, and expansion of the vascular membrane were observed, with the tissues surrounding the vessels appearing spongy. These changes were less severe in the two apurin groups. The water content in the brain tissue was (78.56±0.30) % in ischemia + apurin group and (78.85±0.49) % in ischemia/reperfusion + apurin group, significantly lower than that of (79.08±0.33) % in ischemia + oxolinic acid group and (79.86±0.49) % in ischemia/reperfusion + oxolinic acid group (P ＜ 0.05).CONCLUSION: Apurin can relieve tissue injury after cerebral ischemia/reperfusion by inhibition of SOD.