大鼠肢体缺血再灌注所致脑损伤及其机制探讨

摘要

目的：观察肢体缺血-再灌注对脑的损伤作用，并探讨其可能的机制。方法： SD大鼠随机分为正常(N)、假手术(S)、后肢单纯缺血(I)及缺血-再灌注(I-R)各时间点组。通过夹闭腹主动脉末端4 h、开放2-24 h复制I、I-R组模型。光、电镜观察脑的病理变化；反转录多聚酶链反应及免疫组化染色法，观察脑组织iNOS表达的变化及过氧亚硝基阴离子(ONOO-)的硝基化产物硝基酪氨酸(NT)的生成与分布；比色法测定脑组织MDA含量及SOD活性。结果：①I-R组脑组织水肿，神经元受损较重，S、I组未见异常；②S、I、I-R组iNOS均有表达，I-R 6 h表达量最大。I-R组大脑皮质、海马等区可见弥散分布的NT阳性神经元；③I-R 6 h组MDA含量显著高于N、S、I组，SOD活性显著低于这些组(P＜0.05)；而N、S、I组之间无显著差别。结论：肢体I-R能引发脑损伤，脑内高表达的iNOS-NO-ONOO-可能是脑损伤的重要因素。%AIM:To investigate the pathologic changes in the brain and its underlying mechansims during ischemia-reperfusion of rat hindlimbs.METHODS:SD rats were divided into the normal(N), sham(S), 4 h ischemia without reperfusion(I), and 4 h ischemia-2, 6,12,18 or 24 h reperfusion (I-R) groups at random. Ischemia and ischemia-reperfusion were established with the occlusion or/and re-opening of the terminal of abdominal aorta, respectively. The pathologic changes in the brain tissue were morphologically observed. The expression of inducible nitric oxide synthase (iNOS) mRNA, and iNOS protein and the nitrotyrosine, a marker of peroxynitrite (ONOO-)，in the brain tissue were detected with RT-PCR and immunohistochemical technique, respectively. The brain superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were spectraphotometrically measured.RESULTS:Hydropic degeneration and severe injury to neurons were only showed in I-R group. Expressions of iNOS mRNA and protein were demonstrated in I-R, I and S groups, which were maximal in I-R 6 h group. iNOS positive neurons and microglias were more spread in I-R 6 h group than those in S and I groups. NT positive neurons were localized in the cerebral cortex and hippcampus of I-R 6 h group. The contents of MDA markedly increased, while the activity of SOD significantly decreased in I-R 6 h group compared to the N, S and I groups. There were no significant changes in MDA and SOD in N, S and I groups.CONCLUSION:Severe ischemia-reperfusion of rat hindlimbs could induce brain injury, and its mechanisms might be related to enhanced expression of iNOS-NO-ONOO- in the brain.