CD151通过囊泡内化再循环调控血管通透性的机制  

The mechanism of CD151 regulating vascular permeability through vesicle internalization and recycling

作　　者：范诗浪 蒋卢映 章子璇 季萌萌 左后娟 刘竞搏[2] Fan Shilang;Jiang Luying;Zhang Zixuan;Ji Mengmeng;Zuo Houjuan;Liu Jingbo(Dept of Cardiology,Tianyou Hospital Affiliated to Wuhan University of Science and Technology,Wuhan 430064;Dept of Child Health,Wuhan Children′s Hospital(Wuhan Maternal and Child Health Hospital),Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430016;Tongji Medical College of HUST,Wuhan 430014;Dept of Cardiology,Tongji Hospital Tongji Medical College of HUST,Wuhan 430014)

机构地区：[1]武汉科技大学附属天佑医院心血管内科,武汉430064 [2]华中科技大学同济医学院附属武汉儿童医院(武汉市妇幼保健院)儿童保健科,武汉430016 [3]华中科技大学同济医学院,武汉430014 [4]华中科技大学同济医学院附属同济医院心血管内科,武汉430014

出　　处：《安徽医科大学学报》2025年第2期218-225,233,共9页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81873535);湖北省自然科学基金(编号:2020CFB573)。

摘　　要：目的探究CD151通过调控囊泡内化再循环对血管通透性的影响及机制。方法野生型小鼠和CD151敲除小鼠分别分为WT-con组、WT-模型组、KO-con组和KO-模型组,每组6只。WT-模型组和KO-模型组采用腹腔注射脂多糖(LPS)制备脓毒症急性肺损伤(ALI)模型,WT-con组和KO-con组腹腔注射磷酸盐缓冲液(PBS)作为对照。造模后24 h,通过Miles实验测定肺血管通透性;将沉默CD151表达的siRNA(si-CD151)和阴性对照si-NC转入EA.hy 926细胞,分别观察基础条件及血管内皮生长因子(VEGF-A)刺激条件下,内皮细胞层在不同时间点对FITC-dextran的通透性;CD151敲低组和对照组内皮细胞行转录组测序;免疫荧光检测各组细胞CD151分布及内化;Western bolt及RT-qPCR检测CD151敲低组及对照组VE-cadherin表达情况,免疫荧光检测各组细胞VE-cadherin分布及内化。结果Miles实验结果提示,WT-模型组小鼠肺组织中染料渗出相较WT-con组增加(P<0.01),KO-模型组小鼠肺组织中染料渗出相较WT-模型组增加(P<0.05)。内皮细胞层通透性检测结果提示,在VEGF-A刺激30、60和120 min后,CD151敲低组对FITC-dextran的通透性显著高于对照组(P<0.05)。转录组测序结果提示,内皮细胞中CD151与囊泡介导的转运存在密切关联。Western bolt和RT-qPCR结果提示,CD151敲低的内皮细胞中VE-cadherin在蛋白和mRNA水平表达较对照组显著下降(均P<0.01)。免疫荧光染色结果提示,在VEGF-A刺激后,与对照组比较,CD151表达下降显著破坏细胞-细胞连接处VE-cadherin的表达,降低了CD151和VE-cadherin在核周区域的共定位。结论CD151的缺失影响内皮细胞囊泡内化再循环,影响VE-cadherin的表达与内化活动,进而影响血管通透性。Objective To explore the effect and mechanism of CD151 on vascular permeability by regulating vesicle internalization and recycling.Methods Wild-type mice and CD151 knockout mice were divided into WT-con group,WT-model group,KO-con group and KO-model group,with 6 mice in each group.WT-model group and KO-model group were intraperitoneally injected with LPS to prepare sepsis ALI model,and WT-con group and KO-con group were intraperitoneally injected with phosphate buffer saline(PBS)as a control.24 h after modeling,pulmonary vascular permeability was measured by Miles test.The siRNA silencing CD151 expression(si-CD151)and negative control si-NC were transfected into EA.hy 926 cells.The permeability of endothelial cell layer to FITC-dextran at different time points was observed under basic conditions and vascular endothelial growth factor-A(VEGF-A)stimulation conditions.Transcriptome sequencing of endothelial cells in si-CD151 group and si-NC group;the distribution and internalization of CD151 in each group were measured using immunofluorescence.Western blot and real-time quantitative RT-qPCR were used to detect the expression of VE-cadherin in si-CD151 group and other groups.The distribution and internalization of VE-cadherin in each group were measured using immunofluorescence.Results Miles experiment results indicated that dye exudation in lung tissue of WT-model group was significantly higher than that of WT-con group(P<0.01).The dye exudation in the lung tissue of KO-model group increased compared with WT-model group(P<0.05).The results of endothelial cell layer permeability test showed that the permeability of FITC-dextran in si-CD151 group was significantly higher than that in control group after VEGF-A stimulation for 30,60 and 120 min(P<0.05).Transcriptome sequencing results suggested that CD151 in endothelial cells was closely related to vesicle-mediated transport.Compared with other groups,protein and mRNA levels of VE-cadherin in CD151 knockdown endothelial cells was significantly lower(all P<0.01).The im

关 键 词：急性肺损伤 CD151 内化再循环 内皮细胞 血管通透性 VE-CADHERIN 

分 类 号：R6311.2[医药卫生—外科学]