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

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<0.000 1);uPA活性抑制剂阿米洛利(amiloride)抑制了rTpp47刺激THP-1细胞分泌...

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

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.