agr群体感应系统调控MRSA毒力因子介导的巨噬细胞免疫应答效应的分子机理

Methicillin-resistant Staphylococcus aureus (MRSA) is the major cause of community-acquired and hospital acquired infection, the strategy to decrease the virulence via targeting the quorum sensing system is able to lower the pathogenicity and resistance. Recent study showed that virulence factor of MRSA could produce the lethal damage through activation of macrophage inflammasome. However, the latter induced pyroptosis has beneficial effect on clearance of intracellular bacteria and anti-infection function. Our previous results demonstrated that inhibitor of agr could decrease the release of virulence factor significantly, inhibit the activation of macrophage inflammasome, relief the damage of organs and increase the survival rate in infected mice. Based on these evidence, we proposed that moderate activation of inflammasome induced macrophage pyroptosis would be helpful to clear the pathogen, and present the anti-infection activity, while over-activation of inflammasome will induce macrophage necrosis and severe tissue damage. This project will study the regulation mechanism of agr quorum sensing system on inflammasome activation mediated by MRSA virulence factor in macrophage, and the mechnism of immune response of macrophage to the virulence factor, as well as clearance of pathogenic bacterium. It will shed a light on the regulatory response mechanism between the MRSA QS system and body macrophage, and explore a new way for MRSA infection treatment.

耐甲氧西林金葡菌（MRSA）是医院和社区感染的主要致病菌，干扰MRSA agr群体感应（QS）的抗毒力策略，可显著减弱其致病力、降低其耐药性。新近研究表明，MRSA毒力因子可激活巨噬细胞炎症小体产生致死性损伤；而炎症小体活化诱导的巨噬细胞焦亡却有利于机体清除胞内菌，发挥抗感染作用。我们前期研究证明，在MRSA败血症模型中，agr抑制剂能够明显降低毒力因子分泌、抑制巨噬细胞炎症小体活化、减轻脏器损伤、提高小鼠生存率。因此我们推测，炎症小体适度激活所诱导的巨噬细胞焦亡，有利于宿主清除致病菌，发挥抗感染作用；而炎症小体过度激活则诱发巨噬细胞坏死，造成组织严重损伤。本课题将展开MRSA QS系统调控毒力因子介导的炎症小体活化、巨噬细胞对毒力因子的免疫应答及致病菌清除机制研究。阐明上述问题，将有助于揭示MRSA QS系统与宿主巨噬细胞间的调节应答机制，为有效防治MRSA感染探索新途径。

耐甲氧西林金葡菌（MRSA）群体感应（QS）agr系统所调控的毒力因子可被宿主免疫细胞特异性识别，并激活巨噬细胞炎症小体组装、活化，激活的炎症小体对机体产生保护还是损伤作用颇受争议。本研究通过应用agr缺陷菌和细菌群体感受器抑制剂RIP阻断葡萄球菌群体感受系统，评价和确认阻断葡萄球菌agr系统后的体内抗菌效果，探索阻断agr群体感受系统对细菌致病毒力和宿主免疫细胞应答效应的影响，揭示MRSA QS系统与宿主巨噬和中性粒细胞间的调节应答机制，为有效防治MRSA感染探索新途径。我们的研究结果表明阻断葡萄球菌agr群体感受系统能有效抑制MRSA RNAIII、PSMα、PSMβ、α-溶血素等重要毒力因子的表达和释放，显著降低脓毒症和肺炎模型小鼠的死亡率和脏器荷菌量；通过氯膦酸脂质体清除小鼠体内巨噬细胞，确证阻断agr群体感应系统对小鼠感染的保护作用与巨噬细胞相关；发现PSMα3可通过FPR2-AKT-NF-κB炎性通路诱导巨噬细胞从M0或M2抗炎状态向M1促炎方向极化；阻断agr群体感应系统可抑制巨噬细胞NLRP3炎症小体的激活，促进巨噬细胞清除MRSA，发挥抗感染作用。另外，通过腹腔注射环磷酰胺清除小鼠体内中性粒细胞，证明阻断agr群体感应系统对小鼠感染的保护作用与中性粒细胞密切相关；通过体外细菌与中性粒细胞共培养模型以及psmα突变菌株，发现MRSA 的毒力因子PSM可诱导中性粒细胞发生坏死性凋亡；通过MLKL抑制剂NSA，坏死性凋亡抑制剂Nec，TNFα抗体和FPR2抑制剂WRW4显著抑制PSMα1介导的中性粒细胞裂解，证明PSMα诱导的中性粒细胞坏死性凋亡依赖于FPR2介导的自分泌TNFα。上述结果将为抑制群体感受系统抗菌策略及抗毒力新型抗菌药物研究提供新思路，为耐药菌感染治疗探索新途径。