内皮细胞分裂周期蛋白42在急性肺损伤所致肺血管屏障受损中的机制研究  

Mechanistic study of endothelial cell division cycle protein 42 in pulmonary vascular barrier injury during acute lung injury

作　　者：徐雯 李家飞 王芳[1] 吴南林 秦立龙 XU Wen;LI Jiafei;WANG Fang;WU Nanlin;QIN Lilong(Department of Respiratory and Critical Care Medicine,the Affiliated Chuzhou Hospital of Anhui Medical University,the First People's Hospital of Chuzhou,Chuzhou 23900,China;Department of Pathology,the Affiliated Chuzhou Hospital of Anhui Medical University,the First People's Hospital of Chuzhou,Chuzhou 23900,China;Department of Respiratory and Critical Care Medicine,the First Affiliated Hospital of Wannan Medical College,Wuhu 241000,China)

机构地区：[1]安徽医科大学附属滁州医院(滁州市第一人民医院)呼吸与危重症医学科,安徽滁州239001 [2]安徽医科大学附属滁州医院(滁州市第一人民医院)病理科,安徽滁州239001 [3]皖南医学院第一附属医院呼吸与危重症医学科,安徽芜湖241000

出　　处：《中国病理生理杂志》2025年第4期704-713,共10页Chinese Journal of Pathophysiology

基　　金：皖南医学院中青年基金资助项目(No.WK2021F31);安徽省卫健委科研项目(No.2023xkj208)。

摘　　要：目的:探讨细胞分裂周期蛋白42(cell division cycle protein 42,Cdc42)在急性肺损伤(acute lung injury,ALI)所致肺血管屏障受损中的可能机制。方法:(1)通过GEO(Gene Expression Omnibus)数据库分析ALI中Cdc42和含IQ基序GTP酶激活蛋白1(IQ motif-containing GTPase-activating protein 1,IQGAP1)的表达。(2)收集2022年1月至2023年12月期间在呼吸与危重症医学科及重症医学科确诊为ALI的30例患者和30名健康体检者的血浆样本,同时收集ALI患者的支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)。动物实验选取18只雄性C57小鼠,随机分为对照(control,CON)组、脂多糖(lipopolysaccharide,LPS)组和LPS+ML141(Cdc42抑制剂)组,每组6只。各组小鼠在72 h后处死,收集BALF,计数BALF细胞数目,BCA法测定BALF中蛋白浓度,酶联免疫吸附实验检测人血浆及小鼠BALF中Cdc42及炎症因子白细胞介素6(interleukin-6,IL-6)、IL-1β和肿瘤坏死因子α(tumor necrosis factorα,TNF-α)水平,通过小鼠肺组织切片HE染色评估肺损伤程度。(3)内皮磁珠分选小鼠肺微血管内皮细胞(pulmonary microvascular endothelial cells,PMVECs),细胞实验分为CON组、LPS组和LPS+ML141组。采用血管成环实验检测各组PMVECs的血管成环能力,细胞Ca^(2+)成像技术测定PMVECs的Ca^(2+)水平,活性氧(reactive oxygen species,ROS)检测试剂盒检测PMVECs的ROS水平,并通过Western blot分析各组PMVECs中Cdc42、血管内皮钙黏蛋白(VE-cadherin)、细胞间黏附分子1(intercellular adhesion molecule-1,ICAM-1)、肌球蛋白轻链(myosin light chain,MLC)、磷酸化MLC(phosphorylated MLC,p-MLC)和IQGAP1的蛋白水平。结果:(1)GEO数据库分析显示,ALI模型中Cdc42的表达水平显著升高(P<0.01)。(2)ALI患者血浆中Cdc42及炎症因子(IL-6、IL-1β和TNF-α)水平显著升高(P<0.01),BALF中性粒细胞显著增多;治疗后上述指标显著下降(P<0.05)。ALI动物模型显示,BALF细胞计数、蛋白浓度、炎症因子及肺组织损伤评分均显著升AIM:This study aims to investigate the potential mechanism of cell division cycle protein 42(Cdc42)in acute lung injury(ALI).METHODS:(1)The levels of Cdc42 and IQ motif-containing GTPase-activating protein 1(IQGAP1)in ALI were analyzed using the Gene Expression Omnibus(GEO)database.(2)The plasma samples were collected from 30 patients diagnosed with ALI and 30 healthy controls between January 2022 and December 2023.The bronchoalveolar lavage fluid(BALF)from ALI patients was also collected.Eighteen male C57BL/6 mice were randomly divided into control(CON)group,lipopolysaccharide(LPS)group,and LPS+ML141(Cdc42 inhibitor)group,with 6 mice in each group.After 72 h,the mice were euthanized,and the BALF was collected for analysis,including cell enumeration and protein concentration determination using the bicinchoninic acid method.Enzyme-linked immunosorbent assay was used to measure the levels of Cdc42 and inflammatory cytokines[interleukin-6(IL-6),IL-1βand tumor necrosis factorα(TNF-α)]in human plasma and mouse BALF.Lung damage in mouse tissue sections was evaluated by HE staining.(3)Mouse pulmonary microvascular endothelial cells(PMVECs)were isolated by magnetic bead-based cell sorting and were divided into CON,LPS and LPS+ML141 groups.Vascular ring formation assay was conducted to assess the angiogenic potential of PMVECs,and calcium ion imaging technology was employed to measure calcium ion concentrations in PMVECs.The levels of reactive oxygen species(ROS)were assessed using a ROS detection kit.Western blot was utilized to analyze the protein levels of Cdc42,VE-cadherin,intercellular adhesion molecule-1(ICAM-1),myosin light chain(MLC),phosphorylated MLC(p-MLC)and IQGAP1 in PMVECs.RESULTS:(1)The GEO database analysis revealed significant up-regulation of Cdc42 expression in ALI model(P<0.01).(2)Clinical assessments showed markedly elevated plasma levels of Cdc42 and pro-inflammatory cytokines(IL-6,IL-1βand TNF-α)in ALI patients(P<0.01),with subsequent reductions after treatment(P<0.05).Neutrophil counts in the

关 键 词：急性肺损伤 细胞分裂周期蛋白42 钙黏蛋白 含IQ基元GTP酶激活蛋白1 血管生成 

分 类 号：R563[医药卫生—呼吸系统] R363.2[医药卫生—内科学] Q51[医药卫生—临床医学]