蛋白激酶C在机械牵张致肺上皮细胞损伤中的作用  

Role of protein kinase C in ventilation-induced lung injury

作　　者：刘孟洁[1] 谷长平[1] 王月兰[1] LIU Meng-jie;GU Chang-ping;WANG Yuelan(Department of Anesthesiology and Perioperative Medicine,Qianfoshan Hospital of Shandong Province,the First Hospital Affiliated with Shandong First Medical University,Jinan,Shandong 250014,China)

机构地区：[1]山东第一医科大学第一附属医院(山东省千佛山医院)麻醉与围术期医学科,山东济南250014

出　　处：《实用医药杂志》2021年第5期441-444,共4页Practical Journal of Medicine & Pharmacy

基　　金：国家自然科学基金面上项目(81770076);山东第一医科大学学术提升计划(2019QL015、2019RC012);山东省医药卫生科技发展计划项目(2019WS506)。

摘　　要：目的探讨蛋白激酶C(PKC)在机械牵张致肺上皮细胞损伤中的作用。方法体外培养小鼠肺上皮细胞MLE-12,传代至稳定后分别进行8%和20%应变率的机械牵张,频率0.5 Hz,牵张时间分别为0 h,1 h,2 h,4 h,采用Western blot法测定肺组织occludin蛋白的表达。之后再将MLE-12细胞随机分为3组:牵张组(C组)、二甲基亚砜对照组(D组)和PKC抑制剂BIM组(B组)。牵张前1 h使用PKC抑制剂BisindolylmaleimideⅠ5μmol/L(用二甲基亚砜溶解)处理细胞,采用20%的强度牵张0 h,1 h,2 h,4 h,频率0.5 Hz。采用Western blot法测定肺组织occludin蛋白的表达、显微镜下观察细胞生长状态。结果Occludin蛋白在MLE-12细胞上表达明显,occludin蛋白表达随着牵张强度和时间的增加而减少,20%的强度、牵张4 h occludin蛋白表达明显下降;与C组比较,D组occludin蛋白表达差异无统计学意义(P>0.05),B组机械牵张4 h时occludin蛋白表达上调(P<0.05)。同时,显微镜下显示:与C组比较,D组细胞损伤较明显,B组细胞损伤程度明显减轻。结论蛋白激酶C参与机械牵张致肺上皮细胞损伤。Objective To explore the effect of protein kinase C in damage to alveolar epithelial cells caused by mechanical stretch.Methods MLE-12 cells cultured in vitro and cyclic stretch was conducted at 8%and 20%change in basement membrane surface area applied in a cyclic manner.After the end of stretching for 0h,1h,2h,and 4h,the cells were lysed,and occludin expression was determined by Western blot.Then MLE-12 cells were randomly divided into 3 groups using a random number table:cyclic stretch group(group C),dimethyl sulfoxide control group(group D),and protein kinase C inhibitor BIM group(group B).In D and B groups,dimethyl sulfoxide 30pl/ml and BIM 5μmol/L were added to the culture medium,respectively,and the cells were then cultured for 60min.The cells underwent cyclic stretch for 4h with frequency of 0.5Hz and amplitude of 20%in the three groups.Then MLE-12 cells were collected for determine the expression of occludin using Western blot.Immunofluorescence was used to observe the cell growth status.Results Occludin protein was significantly expressed in MLE-12 cells,and its expression decreased with the increase of stretch strength and time,while occludin protein expression decreased significantly with 20%strength for 4h.Compared with C group,no statistically significant difference in occludin protein expression in group D(P>0.05),while the expression of occludin protein was upregulated in group B(P<0.05).Microscopical showed that the cell damage of group D was more obvious,and the degree of cell damage of group B was significantly reduced compared with C group.Conclusion It considered that the activation of PKC is involved in damage to alveolar epithelial cells caused by mechanical stretch.

关 键 词：蛋白激酶C 连接蛋白类 OCCLUDIN 上皮细胞 

分 类 号：R392.2[医药卫生—免疫学]