脑缺血再灌注损伤模型大鼠炎性反应相关信号研究

The research of the inflammation related signal in the cerebral ischemia reperfusion rat model

目的探讨脑缺血再灌注损伤模型大鼠的炎性反应相关信号链的变化,分析免疫系统炎性损伤与应激反应在中枢神经系统损伤过程中的作用及相互关系。方法采用改良Zea Longa方法制备Spraque-Dawley(SD)大鼠脑缺血再灌注损伤模型。将SD大鼠随机分为空白对照组、假手术组及模型组,采用免疫组化检测技术分析脑组织缺血局部炎性反应相关信号白细胞介素1β(IL-1β)、白细胞介素6(IL-6)、肿瘤坏死因子α(TNF-α)、基质金属蛋白酶9(MMP-9)、细胞间黏附分子1(ICAM-1)、单核细胞趋化蛋白1(MCP-1)和转化生长因子β(TGF-β)在12、24、48、72、96和144h的表达情况,采用ELISA方法检测相应时间点外周血中IL-1β、IL-6、TNF-α、热休克蛋白70(HSP70)和促肾上腺皮质激素(ACTH)的表达情况。结果模型组大鼠与假手术组及空白组大鼠比较,脑缺血损伤局部的炎性反应损伤相关信号IL-1β、IL-6、MMP-9、ICAM-1和TGF-β均表现为功能活动相关的增高表达,各指标相关信号随时间变化的趋势基本相似,呈经典的双峰形模式,24h～48h达到其表达量的第1个峰值,96h～144h达到其表达量的第2个峰值,未观察到TNF-α和MCP-1有相同的表达趋势;外周血中的炎性反应相关信号ACTH、IL-1β、HSP70和TNF-α亦表现为功能活动相关的增高性表达,模型组与假手术组及空白组大鼠比较其表达有统计学差异(P<0.05),各指标相关信号随时间变化的曲线峰形呈经典的单峰形模式,未观察到IL-6有相同的表达趋势。结论在脑缺血再灌注损伤情况下,中枢神经系统损伤局部及外周血中炎性反应相关信号呈活化状态,细胞因子参与了中枢神经系统局部的损伤及损伤修复过程和全身的网络功能调节,在中枢神经系统疾病发生发展过程中发挥关键作用。

Objective To explore the change of the inflammation related signals in the cerebral ischemia reperfusion rat model, and then analyze the effect of the inflammation injury and the stress response to the disease of central nervous system and their interaction. Methods Improved Zea Longa method was used to prepare cerebral isehemia reperfusion model of Spraque-Dawley （SD） rats. Immunohistochemical technique was used to analyse the expression of the inflammatory related signals [interleukin-1β （IL-1β）, interleukin-6 （IL 6）, tumor necrosis factor-~ （TNF-c~）, matrix metalloproteinases-9 （MMP-9）, intercellular adhesion molecule-1 （ICAM 1）, monocyte chemoattractant protein 1 （MCP-1）, transforming growth factor-13 （TGF-α）] in rats injured brain areas at 12 h, 24 h, 48 h, 72 h, 96 h and 144 h after ischemia/reperfusion, and ELISA was used to detect the levels of relevant signals [IL-1β, TNFα, IL-6, Heat Shock Proteins 70 （HSPT0）, adrenocorticotropic hormone （ACTH）] in the rats serum at the same time. Results Compared with the other two groups, in the injured brain regions of the model rats, the injury related signals including IL-1β, IL-6, MMP-9, ICAM-1 and TGF-β showed consistent elevated levels and expressed classical two peaks. They expressed the first peak 24 h to 48 h after ischemia/reperfusion, and then slightly decreased; the second peak appeared 96 h to 144 h after ischemia/reperfusion. The expression trends of TNF-α and MCP-1 were not consistent with others. The patterns of the elevated rats＇ serum signals including IL-1β, TNF-α, HSPT0 and ACTH showed one peak. There were significant differences between the model group and the control （P〈0. 05）. The expression trends ofIL-6 was not consistent with others. Conclusions During the cerebral ischemia reperfusion, the injury related signals both in CNS and serum expressed higher levels than the control, which indicated that cytokines were involved in the process of injury and repair and the neuro-endocrine-immune regulation network in the central nervous system iniury, they play a critical role in the development of central nervous system disease.