TLR4抑制剂TAK-242预处理改善急性缺血性脑卒中诱导的肠黏膜屏障损伤

Pretreatment with TLR4 inhibitor TAK-242 improved intestinal mucosal barrier injury induced by acute ischemic stroke

目的:探讨Toll样受体4(TLR4)抑制剂TAK-242预处理对大鼠急性缺血性脑卒中(AIS)所致肠黏膜屏障损伤的影响。方法:将45只雄性SD大鼠随机分成假手术组(sham)、模型组(model)和AK-242预处理组(TAK-242),每组15只,采用改良Longa线栓法制作AIS大鼠模型。于造模后24 h评估各组大鼠神经功能缺损情况;TTC法检测各组大鼠脑梗死面积;ELISA检测血清中二胺氧化酶(DAO)、D-乳酸(D-Lac)、内毒素(ET)、TNF-α、IL-1β和IL-6的含量;HE染色观察肠黏膜组织病理学变化;异硫氰酸荧光素-葡聚糖(FITC-Dextran)示踪法检测肠黏膜通透性;Western blot检测小肠组织紧密连接蛋白Claudin-1和ZO-1以及TLR4信号通路蛋白TLR4、MyD88和NF-κB p65等蛋白的表达水平。结果:与sham组比较,model组大鼠神经功能缺损评分、脑梗死面积、肠黏膜损伤程度及通透性均显著提高(P<0.05),血清DAO、D-Lac、ET、TNF-α、IL-1β及IL-6含量均显著升高(P<0.05),而小肠组织中Claudin-1和ZO-1蛋白表达水平均显著降低(P<0.05),TLR4、MyD88和NF-κB p65蛋白表达水平均显著升高(P<0.05);与model组比较,TAK-242组大鼠神经功能缺损评分、脑梗死面积、肠黏膜损伤程度及通透性均显著改善(P<0.05),血清DAO、D-Lac、ET、TNF-α、IL-1β及IL-6含量均显著下降(P<0.05),且小肠组织中Claudin-1和ZO-1蛋白表达水平均显著升高(P<0.05),MyD88和NF-κB p65蛋白表达水平均显著降低(P<0.05),而TLR4蛋白水平无明显变化(P>0.05)。结论:TLR4抑制剂TAK-242可改善AIS诱导的肠黏膜屏障损伤,其机制与抑制TLR4/NF-κB信号通路介导的炎症反应有关。

Objective:To investigate the effect of TLR4 inhibitor TAK-242 pretreatment on intestinal mucosal barrier injury in rats with acute ischemic stroke(AIS).Methods:45 male SD rats were randomly divided into sham group,model group,TAK-242 group,15 in each group.The rat model of AIS was established by modified Longa thread method.24 hours after modeling,the neurological deficits of rats was evaluated.The area of cerebral infarction was detected by TTC,the contents of DAO,D-Lac,ET,TNF-α,IL-1βand IL-6 in serum were detected by ELISA,the histopathological changes of intestinal mucosa were observed by HE staining,the intestinal mucosal permeability was measured by FITC-Dextran,the expression levels of Claudin-1 and ZO-1,as well as the TLR4 signaling pathway proteins TLR4,MyD88 and NF-κB p65 in intestinal tissues were detected by Western blot.Results:Compared with sham group,the neurological function score,cerebral infarction area,intestinal mucosa injury degree and permeability of the model group were significantly increased(P<0.05).The contents of DAO,D-Lac,ET,TNF-α,IL-1βand IL-6 in serum were significantly increased(P<0.05).The expression levels of Claudin-1 and ZO-1 in intestinal tissues were significantly decreased(P<0.05),while TLR4,MyD88 and NF-κB p65 expressions were significantly increased(P<0.05).Compared with model group,the neurological function score,cerebral infarction area,intestinal mucosa injury degree and permeability of the TAK-242 group were significantly decreased(P<0.05).The content of DAO,D-Lac,ET,TNF-α,IL-1βand IL-6 in serum were significantly decreased(P<0.05).The expression levels of Claudin-1 and ZO-1 in intestinal tissues were significantly increased(P<0.05),while MyD88 and NF-κB p65 were significantly decreased(P<0.05),and no significant change in TLR4 protein level(P>0.05).Conclusion:TLR4 inhibitor TAK-242 can improve intestinal mucosal barrier injury induced by AIS,and its mechanism is related to the inhibition of TLR4/NF-κB signaling pathway-mediated inflammation.