通腑泻肺颗粒调控MLCK-pMLC通路影响急性肺损伤小鼠肺屏障功能的研究  

Effects of Tongfu Xiefei granules on lung barrier function in acute lung injury mice through regulating MLCK-pMLC signaling pathway

作　　者：钟曦 彭文宏[1] 刘红燕[1] 王思月[1] 王刚 丁燕 冯霞[1] ZHONG Xi;PENG Wenhong;LIU Hongyan;WANG Siyue;WANG Gang;DING Yan;FENG Xia(Chongqing Hospital of Traditional Chinese Medicine,Chongqing 400021,China)

机构地区：[1]重庆市中医院

出　　处：《现代中西医结合杂志》2020年第2期129-134,共6页Modern Journal of Integrated Traditional Chinese and Western Medicine

基　　金：重庆市卫计委中医药科技项目(ZY201702)

摘　　要：目的探讨通腑泻肺颗粒对急性肺损伤小鼠肺屏障功能的作用及可能机制。方法将SPF级雌性BALB/c小鼠随机分为空白组、对照组、模型组、治疗组和激动组,每组25只。模型组、治疗组和激动组采用脂多糖(LPS)诱导小鼠急性肺损伤模型;治疗组于注射LPS 30 min后灌胃通腑泻肺颗粒;激动组注射肌球蛋白轻链激酶(MLCK)激动剂P物质(SP)15 min后注射LPS,30 min后灌胃通腑泻肺颗粒。造模16 h后,颈动脉放血处死小鼠,光学显微镜下观察各组小鼠肺组织病理切片;测定小鼠肺组织湿/干重比;Evans Blue法检测肺血管通透性;BCA法检测支气管肺泡灌洗液中总蛋白浓度,并计数肺泡灌洗液中白细胞总数;qRT-PCR法检测肺组织中带状闭合蛋白-1(ZO-1)、血管内皮细胞钙依赖性黏附素(VE-cadherin)mRNA表达水平,Western Blot法检测肺组织中ZO-1、VE-cadherin、MLCK、磷酸化肌球蛋白轻链(pMLC)蛋白表达水平。结果治疗组小鼠肺部炎症反应减轻,且肺血管通透性、肺组织湿/干重比、肺泡灌洗液中总蛋白水平及白细胞数量均显著低于模型组及激动组(P均<0.05);治疗组小鼠肺组织中ZO-1、VE-cadherin mRNA及蛋白表达水平均明显高于模型组和激动组(P均<0.05),MLCK、pMLC蛋白表达水平均明显低于模型组和激动组(P均<0.05)。结论通腑泻肺颗粒可以显著改善LPS诱导的小鼠急性肺损伤的肺部屏障功能,机制可能为其可抑制MLCK-pMLC通路活性,维持肺血管内皮通透性,从而减轻肺部炎性渗出及肺水肿。Objective It is to investigate the effect of Tongfu Xiefei granules on lung barrier function in acute lung injury mice and its possible mechanism.Methods SPF female BALB/c mice were randomly divided into blank group,control group,model group,treatment group,and activating group,with 25 mice in each group.The model group,treatment group and activating group were induced by lipopolysaccharide(LPS)to establish models of acute lung injury.The treatment group was given Tongfu Xiefei granules by gavage in 30 minutes after injection of LPS.The activating group was injected with myosin light chain kinase(MLCK)Agonist Substance P(SP),then was injected with LPS in 15 min later and given Tongfu Xiefei granules by gavage in 30 minutes later.In 16 hours after modeling,all the mice were sacrificed by carotid exsanguination,and the lung tissue pathological sections of each group were observed under a light microscope;the lung wet/dry weight ratio of the mice was measured;pulmonary vascular permeability was measured by Evans Blue method;the concentration of total protein in bronchoalveolar lavage fluid by BCA method,and the total number of white blood cells in alveolar lavage fluid was counted;qRT-PCR method was used to detect the mRNA expression of zona occludenn-1(ZO-1)and vascular endothelial-cadherin(VE-cadherin)in lung tissue,Western Blot method was used to detect the expression levels of ZO-1,VE-cadherin,MLCK,and phosphorylated myosin light chain(pMLC)protein in lung tissue.Results The lung inflammation of the mice in the treatment group was alleviated,and the pulmonary vascular permeability,lung tissue wet/dry weight ratio,level of total protein and leukocyte counts in the alveolar lavage fluid were significantly lower than those in the model group and the activating group(P<0.05);the expression levels of protein and mRNA of ZO-1,VE-cadherin in lung tissue of the treatment group were significantly higher than those of the model group and the activating group(P<0.05),and the expression levels of MLCK and pMLC proteins were

关 键 词：通腑泻肺颗粒 急性肺损伤 内皮屏障功能 肌球蛋白轻链激酶 

分 类 号：R-332[医药卫生]