梅毒螺旋体纤连蛋白结合蛋白Tp0136前炎症活性的研究

Proinflammatory Activity of Recombinant Fibronectin-Binding Proteins Tp0136 of Treponema Pallidum

目的初步研究梅毒螺旋体纤连蛋白结合蛋白Tp0136潜在的前炎症活性。方法构建重组质粒p ET-28a/Tp0136并诱导其表达目的蛋白;将不同浓度(1、2、5、10μg/m L)的r Tp0136分别刺激THP-1源性巨噬细胞,ELISA法分别检测其产生前炎症细胞因子IL-1β、IL-6和TNF-α的水平;并同时用二硫代氨基甲酸吡咯烷(PDTC)预先处理巨噬细胞,检测相应细胞因子分泌量。结果成功构建了p ET-28a/Tp0136重组质粒,并诱导表达和纯化出r Tp0136;重组蛋白能诱导巨噬细胞产生IL-1β、IL-6和TNF-α,并且呈浓度和时间依赖性;重组蛋白刺激经PDTC预处理后的巨噬细胞,其IL-1β、IL-6和TNF-α的分泌水平分别降至32%、35%和27%。结论r Tp0136可诱导巨噬细胞产生前炎症细胞因子,该过程可能受NF-κB调控。

Objective To explore potential proinflarmnatory activity of the Treponema pallidum fibronectin-binding protein （FnBP） Tp0136. Methods The recombinant pET-28a/Tp0136 was constructed and induced to express the targeting protein in E.coli BL21.In vitro various concentrations of protein was used to stimulate human macrophages derived from THP-1 cell line at different time.ELISA was performed to detect the level of proinflammatory cytokines IL-1β,IL-6 and TNF-α;Recombinant protein was used to stimulate the macrophages pre-treated with PDTC,and ELISA was performed to detect the releasing IL-1β,IL-6 and TNF-α respectively. Results The recombinant plasmid was constructed and the corre- sponding recombinant protein was obtained successfully;rTpO136 could induce production of IL-1β, IL-6 and TNF-α in dose- and time-dependent manners significantly.Secretion of IL-1β, IL-6 and TNF-α decreased to 32%, 35% and 27% respectively when macrophages was pre-treated with PDTC. Conclusion The data suggested that the FnBP TpO136 could activate the THP-derived human macrophages to release the proinflammatory cytokines,which might be regulated via NF-κB.