重组梅毒螺旋体蛋白Tp47通过诱导uPA增加血管内皮细胞通透性  

Recombinant Treponema pallidum protein Tp47 increases permeability of vascular endothelial cells through inducing uPA

作　　者：于宝丹[1] 陈聪 柯吴坚 吕萍 YU Bao-dan;CHEN Cong;KE Wu-jian;LYU Ping(Department of Laboratory Medicine, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120;Department of Dermatology, Dermatology Hospital, Southern Medical University, Guangzhou 510095,China)

机构地区：[1]广州医科大学附属第一医院检验科,广东广州510120 [2]南方医科大学皮肤病医院皮肤科,广东广州510095

出　　处：《基础医学与临床》2022年第1期120-125,共6页Basic and Clinical Medicine

基　　金：广东省公益研究与能力建设资金(2017A020215152)。

摘　　要：目的探讨重组梅毒螺旋体蛋白Tp47(rTpp47)对单核-巨噬细胞系THP-1合成尿激酶型纤溶酶原激活物(uPA)的调控及其对人脐静脉/血管内皮细胞(HUVECs)通透性的影响。方法用rTpp47刺激THP-1细胞24 h后,分别收集细胞培养上清和THP-1细胞,用ELISA和Western blot检测THP-1细胞表达的uPA含量;用THP-1细胞培养上清刺激人单层血管内皮细胞后,使用FITC-葡聚糖评价单层内皮细胞通透性的变化,用Western blot检测uPA对HUVECs细胞紧密连接蛋白claudin-5表达的影响以及PKC信号通路是否参与rTpp47诱导的THP-1细胞表达uPA。结果重组蛋白rTpp47刺激THP-1细胞合成和分泌的uPA显著高于对照组(P<0.05,P<0.001);用rTpp47与THP-1细胞共培养24 h后收集的细胞培养上清刺激单层血管内皮细胞12和24 h,实验组血管内皮细胞相对通透性显著高于对照组(P<0.05,P<0.0001);uPA活性抑制剂阿米洛利(amiloride)抑制了rTpp47刺激THP-1细胞分泌uPA(P<0.0001),改善了rTpp47对HUVECs的claudin-5蛋白表达的抑制作用(P<0.05),抑制了rTpp47诱导的THP-1细胞表达蛋白激酶C(PKC)的磷酸化(P<0.05)。结论重组蛋白rTpp47可能通过激活PKC信号通路刺激THP-1细胞合成和分泌的uPA,uPA作用于血管内皮细胞,可能导致血管内皮细胞通透性升高。Objective To investigate the effect of recombinant Treponema pallidum protein Tp47(rTpp47)on the expression of uPA by THP-1 macrophages and on the permeability of human umbilical vein/vascular endothelial cells(HUVECs).Methods THP-1 cells were stimulated with rTpp47 for 24 hours and then the culture supernatant and THP-1 cells were collected respectively.The expression of urokinase-type plasminogen activator(uPA)in THP-1 cells was detected by ELISA and Western blot.After stimulation of HUVECs monolayers by THP-1 cell culture supernatant,FITC-dextran was used to evaluate the permeability of HUVECs.Western blot method wasused to observe the effect of uPA on the expression of tight junction protein claudin-5 of HUVECs and the protein kinase C(PKC)signaling pathway of THP-1 cells stimulated by rTpp47.Results The synthesis and secretion of uPA of THP-1 cells stimulated by rTpp47 were significantly higher than those in the control group(P<0.05,P<0.001).When HUVECs were stimulated by the THP-1 cell supernatant for 12 hours and 24 hours,the relative permeability of vascular endothelial cells in the experimental group was significantly higher than the control group(P<0.05 for 12 hours and P<0.0001 for 24 hours).Amiloride,the synthetic inhibitor of uPA,significantly inhibited the uPA expression and secretion of THP-1 cells stimulated by rTpp47(P<0.0001),alleviated the inhibition effect of rTpp47 on claudin-5 expression of HUVECs(P<0.05),and significantly suppressed PKC phosphorylation(P<0.05)in THP-1 cells induced by rTpp47.Conclusions The rTpp47 can stimulate the synthesis and secretion of uPA in THP-1 cells through a potential mechanism of activating PKC signaling pathway and uPA may increase the permeability of vascular endothelial cells.

关 键 词：梅毒螺旋体 重组蛋白Tp47(rTpp47) 尿激酶型纤溶酶原激活物(uPA) 巨噬细胞 人脐静脉/血管内皮细胞(HUVECs) 

分 类 号：R75[医药卫生—皮肤病学与性病学]