梅毒螺旋体脂蛋白刺激巨噬细胞产生细胞因子及脂蛋白耐受的机制研究

摘要

目的：通过对梅毒螺旋体（treponema pallidum，TP）脂肽激活巨噬细胞产生细胞因子以及脂肽所诱导的免疫耐受对信号通路的影响进行研究，为进一步完善TP感染人体的免疫学过程，解释TP感染人体后引起的血清固定现象提供理论依据。方法不同浓度的TP脂肽刺激经PMA诱导THP-1细胞转化的巨噬细胞，利用酶联免疫吸附试验（ELISA）检测其分泌细胞因子的水平，并通过阻断CD14受体后，检测巨噬细胞对合成脂肽的反应能力以及合成脂肽耐受刺激后诱导巨噬细胞产生免疫耐受的能力。采用Western blot法检测细胞内的信号转导分子。结果3种合成脂肽诱导巨噬细胞分泌白细胞介素(IL)-β及IL-8的能力随着脂肽浓度升高而递增。CD14受体阻断后，IL-1β及IL-8分泌水平均明显减少。合成脂肽经耐受后刺激的IL-1β及IL-8分泌水平明显低于直接刺激的细胞因子的水平。合成脂肽在耐受刺激下，其巨噬细胞内的信号转导分子toll样受体2（toll like receptor 2， TLR2）和核转录因子kappa B (NF-κB) P65的蛋白表达显著减少。结论3种合成脂肽均能诱导巨噬细胞产生IL-1β及IL-8。合成脂肽通过激活巨噬细胞膜表面CD14受体分子，活化下游信号通路，从而诱导细胞因子产生。合成脂肽能够诱导巨噬细胞模型产生免疫耐受，其可能机制为通过TLR2激活下游NF-κB信号通路，减少炎性细胞因子产生。%Objective To explore the possible mechanism by which treponema pallidum (TP) lipopeptides stimulates macrophages to produce cytokines and to investigate the immunotolerance induced by the lipopeptides in the NF-κB signaling pathway, so as to proivide a reference for improving the understanding of the immunological process of human infected by TP and interpreting the serofast phenomenon caused by TP infection. Methods The concentration of cytokines secreted by THP-1 cells under the stimulation of TP lipopeptides were tested by using ELISA. After blocking the CD14 receptor, the reaction of macrophages against lipopeptides was analyzed and the ability of the lipopeptides to induce immunotolerance of macrophages was evaluated. The expressions of intracellular signal transduction molecules were measured by western blot. Results The abilities of three types of lipopeptides to induce macrophages cells to secrete the cytokines IL-βand IL-8 progressively increased with the lipopeptide levels. The levels of both IL-1βand IL-8 signiifcantly decreased after blocking the CD14 receptor. The levels of both IL-1βand IL-8 produced by the immunotolerance induced by lipopeptides were considerably less than those resulting from direct stimulation. The protein expression levels of the signal transduction molecules TLR2 and NF-κB P65 signiifcantly decreased after the immunotolerance induced by lipopeptides. Conclusion Macrophages cells can be induced by three types of lipopeptides to produce IL-1βand IL-8. The downstream signaling pathway is activated by the lipopeptides through CD14 receptor on the membrane of macrophages to induce cytokine production. The lipopeptides may stimulate macrophages to create immunotolerance by activating the downstream NF-κB signaling pathway via TLR2 to reduce cytokines production. The immunotolerance induced by TP lipopeptides may be an important cause of the syphilis serofast phenomenon. Further study and veriifcation are still required.