脂多糖应答分子LRG参与内毒素预处理诱导脑缺血耐受机制的研究

The mechanisms of cerebral ischemia tolerance by LRG response molecule involved in endotoxin preconditioning

目的探讨脂多糖应答分子LRG参与内毒素预处理诱导脑缺血耐受机制。方法体外实验:小鼠脑皮质神经元细胞、体外神经元脑缺血再灌注模型,LPS不同处理后,Westernblot检测细胞中LRG的表达。体内实验:采用大脑中动脉线栓法(MCAO)建立小鼠脑缺血再灌注模型,分为LPS预处理+脑缺血再灌注模型组、脑缺血再灌注模型组和假手术对照组,Western blot检测小鼠脑组织LRG的表达。利用RNA干扰技术,构建LRG沉默型MCAO模型。术后24 h,Garcia法评价神经功能得分,再次检测LRG表达水平,并检测血清中炎性因子(IL-1β,IL-6,TNF-α)的表达水平。结果在神经元和小鼠脑缺血再灌注实验中,与对照组相比,LPS刺激组中LRG表达水平上调;与LPS直接刺激组相比,LPS预处理组中LRG表达水平下调,差异均有统计学意义(P<0.05)。和MCAO模型组比较,LRG基因沉默组小鼠中LRG的表达水平下调,术后24 h神经功能评分较低,神经功能缺损少,差异均有统计学意义(P<0.05)。结论脂多糖应答分子LRG可能参与内毒素诱导的缺血预处理刺激缺血耐受的发生。

Objective To explore the mechanisms of cerebral ischemia tolerance by lipopolvsaccharide response gene(LRG)response molecule involved in endotoxin preconditioning.Methods The study included in vitro experiments and in vivo animal model experiments.In in vitro experiments,cerebral cortex neurons were separated and used to establish in vitro ischemia-reperfusion model and then,expression level of LRG in cells was measured by Western blot after treatment with lipopolysaccharide(LPS).In animal experiments,middle cerebral artery occlusion(MCAO) mice model was established and then,the animals were divided into LPS pretreatment+cerebral ischemia reperfusion model group,cerebral ischemia reperfusion model group and sham operation group.The LRG expression level was detected by Western blot.The LRG silent MCAO model was also established in LRG silenced mice by using RNA interference technology.After 24 hours,neurological function score evaluated by Garcia method,and LRG expression levels and serum inflammatory factors such as IL-1β,IL-6 and TNF-α were measured.Results In neurons and cerebral Ischemia reperfusion experiments,LRG expression was significantly up-regulated in the LPS stimulation groups when compared to control group.LRG expression was significantly down-regulated in LPS pretreatment group when compared to LPS direct treatment(P <0.05).In compassion of the MCAO model group,LRG expression level in brain tissue of the LRG gene silencing group was significantly down-regulated(P <0.05).After24 hours of operation,neurological function score and neurological deficits were decreased(P <0.05).Conclusion LRG may involve in endotoxin-induced ischemic preconditioning to induce ischemic tolerance.