临床铜绿假单胞菌TBCF10839抗吞噬机制研究

   铜绿假单胞菌是广泛分布在环境中的条件致病菌，并非典型的胞内生存菌，而铜绿假单胞菌慢性感染分离株TBCF10839却具有在吞噬细胞内复制的能力，其在细胞内逃逸吞噬效应的相关机制目前尚不明确。本项目旨在比较性地分析铜绿假单胞菌模式菌株PAO1和TBCF10839在吞噬细胞内的生长特征及其对吞噬细胞功能的影响，探究铜绿假单胞菌在吞噬细胞内的生存机制，为治疗铜绿假单胞菌的慢性感染提供理论依据。本研究中，我们发现部分铜绿假单胞菌PAO1在巨噬细胞内呈现生长状态。蛋白质组学分析结果显示，PAO1进入巨噬细胞后，与亚精胺代谢途径相关蛋白表达量显著提高。因此我们构建了亚精胺代谢途径缺失突变株，发现阻断亚精胺分解代谢途径可降低巨噬细胞内PAO1的细菌数量及PAO1对巨噬细胞产生的毒性。由于铜绿假单胞菌Ⅲ型分泌系统可被亚精胺摄取途径激活并在感染中发挥重要作用，我们进一步比对了PAO1与TBCF10839的转录组，以及被上述两菌株感染后的巨噬细胞转录组。结果显示，TBCF10839中Ⅲ型分泌系统相关基因的表达水平显著低于PAO1。相较于经PAO1感染的巨噬细胞，经TBCF10839感染的巨噬细胞下调了大量参与免疫激活相关的基因。此外，体外感染实验结果显示，TBCF10839在巨噬细胞内的细菌数量显著低于PAO1以及Ⅲ型分泌系统过表达突变株TBCF10839 ΔexsD；TBCF10839在巨噬细胞内逃逸吞噬小体的能力弱于PAO1以及TBCF10839 ΔexsD。因此，本研究结果证实了铜绿假单胞菌在进入吞噬细胞后能通过激活亚精胺分解代谢途径来增强保内生存能力，亚精胺代谢途径可能通过激活Ⅲ型分泌系统来增强铜绿假单胞菌逃逸吞噬小体，本研究结果也暗示慢性感染中分离的铜绿假单胞菌临床菌株TBCF10839可能通过下调Ⅲ型分泌系统相关基因的表达来降低宿主免疫细胞的激活从而逃逸吞噬。

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