金黄葡萄球菌杀白细胞素(PVL)对THP-1巨噬细胞IL-8、IL-6表达的影响

摘要

目的 研究金黄葡萄球菌杀白细胞素( panton-valentine leucocidin,PVL)对THP-1巨噬细胞Toll样受体4(TLR4)/核因子κB(NF-κB)信号通路及IL-8、IL-6的表达影响,探讨PVL相关肺组织损伤的致病机制.方法 实验前用100nmol/L佛波酯(PMA)孵育THP-1细胞48 h,使其诱导分化成巨噬细胞,同体外诱导PVL重组表达载体表达纯化的LukS-PVL和LukF-PVL组分共同孵育,采用ELISA法检测细胞上清IL-8、IL-6蛋白的表达,RT-PCR法检测TLR4、IL-8和IL-6 mRNA水平表达,免疫组化和Western blot检测NF-κB的移位及其表达.结果 rPVL能促进THP-1巨噬细胞分泌IL-6和IL-8,而且具有明显的浓度和时间依赖性.同时,rPVL能上调TLR4表达,活化NF-κB蛋白.结论 PVL相关肺组织损伤机制可能与PVL激活TLR4/NF-κB信号通路,启动炎性因子大量释放有关.%Objective To investigate the influence of panton-valentine leucocidin (PVL) on expression of Toll like receptor 4(TLR4)uclear factor-kappa B(NF-κB) signals and IL-8,IL-6 in THP-1 macrophages,and to study the mechanism of PVL-related lung tissue damage.Methods THP-1 cells were cultured in the presence of 100 nmol/L phorbol-12-myristate 3-acetate (PMA) for 48 h to induce monocytemacrophage differentiation.rPVL-F and rPVL-S were induced and expressed from the recombinant plasmid,respectively purified with chromatographic column. After that,THP-1 macrophages were incubated with rPVL,and then ELISA was performed to test expression of IL-8 and L-6 in supernatants fluid; RT-PCR was performed to detect expression of IL-8,L-6 and TLR4 ; NF-κB was analyzed by Western blot and immunohistochemistry method.Results PVL was able to induce expression of IL-8 and IL-6 in THP-1 macrophages in time-and concentration-dependent manners.PVL could also significantly promote the activation of TLR4/NF-κB signals.Conclusion PVL can activate the expression of TLR4/NF-κB signals,and increased the high expression of inflammatory cytokines.Maybe it's the mechanism of action of PVL exerts the function of lung tissue damage.