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中华肺部疾病杂志(电子版)

中华肺部疾病杂志(电子版) ›› 2025, Vol. 18 ›› Issue (05) : 679 -684. doi: 10.3877/cma.j.issn.1674-6902.2025.05.003

论著

PALM3对铜绿假单胞菌致血管内皮细胞损伤作用的机制研究
吴秀琳1, 郭亮2, 蔡俊3, 何志强4, 王悦4, 杨天仪4, 唐敏4, 李明霞5, 杨智勇6, 易斌6, 熊玮1, 廖江荣7,†(), 吴学玲8,†()   
  1. 1400038　重庆，陆军（第三）军医大学第一附属医院老年医学与特勤医学科
    2400037　重庆，陆军（第三）军医大学大学第二附属医院呼吸及危重症医学科
    3400061　重庆，武警重庆总队医院检验科
    4400042　重庆，重庆医科大学检验医学院临床检验系
    5400038　重庆，陆军（第三）军医大学第一附属医院干细胞与再生医学科
    6400038　重庆，陆军（第三）军医大学第一附属医院麻醉科
    7563099　贵州，贵州航天医院呼吸及危重症医学科
    8201112　上海，上海交通大学医学院附属仁济医院呼吸科
  • 收稿日期:2025-06-23 出版日期:2025-10-25
  • 通信作者: 廖江荣, 吴学玲
  • 基金资助:
    重庆市自然科学基金(cstc2019jcyj-msxmX0746); 遵义市呼吸疾病快速精准诊断创新人才团队

Study on the mechanism of PALM3 expression on human umbilical vein endothelial cells induced by Pseudomonas aeruginosa

Xiulin Wu1, Liang Guo2, Jun Cai3, Zhiqiang He4, Yue Wang4, Tianyi Yang4, Min Tang4, Mingxia Li5, Zhiyong Yang6, Bin Yi6, Wei Xiong1, Jiangrong Liao7,†(), Xueling Wu8,†()   

  1. 1Department of Geriatrics and Special Services, the First Affiliated Hospital of Army Medical University, Chongqing 400038, China
    2Department of Respiratory and Critical Care Medicine, the Second Affiliated Hospital of Army Medical University, Chongqing 400037, China
    3Laboratory Department of Chongqing Armed Police Corps Hospital, Chongqing 400061, China
    4Department of Clinical Laboratory Science, School of Laboratory Medicine, Chongqing Medical University, Chongqing 400042, China
    5Department of Stem Cell and Regenerative Medicine, the First Affiliated Hospital of Army Medical University, Chongqing 400038, China
    6Department of Anesthesiology, the First Affiliated Hospital of Army Medical University, Chongqing 400038, China
    7Department of Respiratory and Critical Care Medicine, Guizhou Aerospace Hospital, Chongqing 563099, China
    8Department of Respiratory Medicine, Renji Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Chongqing 201112, China
  • Received:2025-06-23 Published:2025-10-25
  • Corresponding author: Jiangrong Liao, Xueling Wu
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引用本文：

吴秀琳, 郭亮, 蔡俊, 何志强, 王悦, 杨天仪, 唐敏, 李明霞, 杨智勇, 易斌, 熊玮, 廖江荣, 吴学玲. PALM3对铜绿假单胞菌致血管内皮细胞损伤作用的机制研究[J/OL]. 中华肺部疾病杂志(电子版), 2025, 18(05): 679-684.

Xiulin Wu, Liang Guo, Jun Cai, Zhiqiang He, Yue Wang, Tianyi Yang, Min Tang, Mingxia Li, Zhiyong Yang, Bin Yi, Wei Xiong, Jiangrong Liao, Xueling Wu. Study on the mechanism of PALM3 expression on human umbilical vein endothelial cells induced by Pseudomonas aeruginosa[J/OL]. Chinese Journal of Lung Diseases(Electronic Edition), 2025, 18(05): 679-684.

目的

探讨PALM3对铜绿假单胞菌(Pseudomonas aeruginosa, PA)致血管内皮细胞损伤的作用机制。

方法

将人脐血静脉血管内皮细胞(human umbilical vein endothelial cells, HUVEC)随机分为Control组、Control＋PA组、siRNA-Control＋PA组、siRNA-PALM3＋PA组。Control组为HUVEC细胞＋生理盐水，Control＋PA组HUVEC细胞暴露于荧光标记5×108 CFU/ml PA菌液，siRNA-Control＋PA组、siRNA-PALM3＋PA组分别使用siRNA-Control、siRNA-PALM3质粒转染细胞，转染48 h后加入PA；检测细胞上清液中血管内皮纤维连接蛋白(fibronectin)、血管内皮生长因子C(vascular endothelial growth factor C, VEGF-C)、血管生成素-2(angiopoietin-2, ANGPT2)；检测细胞中PALM3、核因子NF-κB表达，采用MTT比较细胞生存率，通过流氏细胞术分析细胞凋亡。

结果

与Control组相比，Control＋PA组和siRNA-Control＋PA组细胞中PALM3蛋白表达升高(P<0.05)；与Control＋PA组和siRNA-Control＋PA组相比，siRNA-PALM3＋PA组细胞中PALM3蛋白表达量降低，NF-κB蛋白表达量降低；siRNA-PALM3＋PA组纤维连接蛋白(235.7±17.5)、VEGF-C(0.72±0.05)、ANGPT2(807.8±24.6)高于Control＋PA组纤维连接蛋白(109.9±5.8)、VEGF-C (0.49±0.06)、ANGPT2(464.16±26.27)和siRNA-Control＋PA组纤维连接蛋白(188.9±8.6)、VEGF-C(0.49±0.02)、ANGPT2(510.57±27.6)(P<0.05); siRNA-PALM3＋PA组细胞存活率(1.102±0.125)，较Control＋PA组(0.870 ± 0.052)和siRNA-Control＋PA(0.887±0.054)增高(P<0.05)；Control＋PA组和siRNA-Control＋PA组6 h后细胞存活率分别为(2.760±0.744)、(2.777±0.604)，较Control组(1.662±0.150)增高，与Control＋PA组和siRNA-Control＋PA组比较，siRNA-PALM3＋PA组细胞存活率(3.195±1.342)升高。

结论

下调PALM3对铜绿假单胞菌致肺血管内皮细胞损伤炎症反应有保护作用，与血管通透性呈负相关。

关键词: 急性肺损伤, PALM3, 铜绿假单胞菌, 血管内皮细胞, 血管通透性相关因子
Objective

To explore the mechanism of PALM3 on vascular endothelial cell injury induced by Pseudomonas aeruginosa (PA).

Methods

Human umbilical vein endothelial cells (HUVEC) were randomly divided into the control group, the control+ PA group, the siRNA-control+ PA group, and the siRNA-PALM3+ PA group. Control group: HUVEC cells + normal saline; Control+ PA group: HUVEC cells exposed to fluorescently labeled 5×108 CFU/ml PA bacterial solution .The siRNA-Control+ PA group and the siRNA-PALM3+ PA group transfected cells with siRNA-Control and siRNA-PALM3 plasmids respectively, and PA was added 48 hours after transfection. It was detected that vascular endothelial fibronectin, vascular endothelial growth factor c (VEGF-C), and angiopoietin-2 (ANGPT2) in the cell supernatant; The expressions of PALM3 and nuclear factor NF-κB in cells were detected. The cell survival rate was compared by MTT, and cell apoptosis was analyzed by flow cytometry.

Result

Compared with the Control group, the expression of PALM3 protein in the cells of the Control+ PA group and the siRNA-Control+ PA group increased (P<0.05); Compared with the Control+ PA group and the siRNA-Control+ PA group, the expression level of PALM3 protein in the cells of the siRNA-PALM3+ PA group decreased, and the expression level of NF-κB protein decreased. The fibronectin (235.7±17.5), VEGF-C(0.72±0.05), and ANGPT2 (807.8±24.6) in the siRNA-PALM3+ PA group were higher than those in the Control+ PA group (109.9±5.8) and VEGF-C(0.49±0.06), ANGPT2 (464.16±26.27), and fibronectin (188.9±8.6), VEGF-C (0.49±0.02), ANGPT2 (510.57±27.6) in the siRNA-Control+ PA group (P<0.05); The cell survival rate of the siRNA-PALM3+ PA group (1.102±0.125) was higher than that of the Control+ PA group (0.870±0.052) and the siRNA-Control+ PA group (0.887±0.054) (P<0.05); The cell survival rates of the Control+ PA group and the siRNA-Control+ PA group after 6 hours were (2.760 ± 0.744) and (2.777 ± 0.604), respectively, which were higher than that of the Control group (1.662±0.150). Compared with the Control+ PA group and the siRNA-Control+ PA group, the cell survival rate in the siRNA-PALM3+ PA group (3.195±1.342) increased.

Conclusion

Down-regulation of PALM3 has a protective effect on the inflammatory response of pulmonary vascular endothelial cell injury caused by Pseudomonas aeruginosa, and is negatively correlated with vascular permeability.

Key words: Acute lung injury, PALM3, Pseudomonas aeruginosa, Vascular endothelial cells, Proinflammatory cytokines
图1 Western biot检测血管内皮细胞中PALM3表达注：Control＋PA组为铜绿假单胞菌组；SiRNA-Control＋PA组为铜绿假单胞菌＋腺病毒转染阴性对照组；SiRNA-PALM3＋PA组为铜绿假单胞菌＋PALM3腺病毒转染组；β-action为β-肌动蛋白
表1 每组HUVEC血管内皮细胞通透性指标比较(±s)
组　别 纤维连接蛋白 VEGF-C ANGPT2
Control 125.6±4.33 0.28±0.03 570.3±29.4
Control＋PA 109.9±5.8a 0.49±0.06a 464.16±26.27a
SiRNA-Control＋PA 188.9±8.6ab 0.49±0.02a 510.57±27.6ab
SiRNA-Plam3＋PA 235.7±17.5abc 0.72±0.05abc 807.8±24.6abc
F值 124.7 53.66 127.6
P值 0.0001 0.0001 0.0001
图2 Western biot检测血管内皮细胞中NF-κB蛋白表达注：Control＋PA组为铜绿假单胞菌组；SiRNA-Control＋PA组为铜绿假单胞菌＋腺病毒转染阴性对照组；SiRNA-PALM3＋PA组为铜绿假单胞菌＋PALM3腺病毒转染组；β-action为β-肌动蛋白；NF-κB为核因子κB
图3 流氏细胞术检测血管内皮细胞凋亡图注：Annexin V-FITC单阳细胞(第四象限)为早期凋亡细胞，Annexin V-FITC和PI双阳细胞(第二象限)为坏死或晚期凋亡细胞，PI单阳细胞为裸核细胞
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