肺炎衣原体Cpn0423通过Nod2/Rip2途径介导小鼠肺部炎症损伤及其作用机制研究

Chlamydia pneumoniae (C. pneumoniae) is an important respiratory pathogen. As reported recently, Nod2 receptor was involved in the recognition of C. pneumoniae infection and stimulation of innate immune response, however, whose mechanism is still unclearly. We have recently found that 1) C. pneumoniae Cpn0423 is a protein secreted from C. pneumoniae, which is located in the cytoplasm of host cell after C. pneumoniae infection and possess the ability to interact with the Nod2 receptor; 2) Cpn0423 could induce Nod2 overexpression and MIP-2 production in BMDM, suggesting Cpn0423 may be a potential ligand of Nod2 during Cpn infection. According to these previous findings, we hypothesize that Cpn0423 interacts with host Nod2 receptor, activates Nod2/Rip2 signal pathway, and induces pulmonary inflammation. To testify our hypothesis, lazer scan confocal microscope,fluorescence resonance energy transfer and signal pathway blocking will be used to analyze the effects of Nod2/Rip2 signal pathway on Cpn0423-induced immune response. Next, Nod2-/- and Rip2-/- animal models will be used to identify the the effects of Nod2/Rip2 signal pathway on Cpn0423-induced pulmonary inflammation. This study will not only provide theoretical and experimental evidence on further revealing the pathogenesis of Chlamydia pneumoniae, but also shed light on the prevention and clinical treatment on the disease related C. pneumonia infection.

肺炎衣原体（Cpn）是一种重要的呼吸道病原体，新近发现Nod2参与Cpn感染的识别，但其作用机制不明。我们前期研究证实肺炎衣原体Cpn0423是一种定位于宿主细胞胞浆的分泌型蛋白，可诱导宿主细胞中Nod2活化及MIP-2等炎症因子分泌。据此，我们推测Cpn0423是Nod2的潜在配体，提出Cpn0423通过激活Nod2/Rip2路径介导小鼠肺部炎症损伤这一科学假说。本项目拟从细胞水平采用激光共聚集、荧光共振能量转移、信号通路阻断等手段，观察Nod2/Rip2途径对Cpn0423诱导宿主细胞产生炎症反应的影响；从动物水平上采用Nod2、Rip2基因敲除鼠模型，观察Nod2/Rip2途径对Cpn0423介导小鼠肺部炎症损伤的影响，最终阐明Cpn0423通过激活Nod2/Rip2路径及其介导小鼠肺部炎症损伤的作用机制，为拓展Cpn致病机制及肺炎衣原体感染性相关疾病的防治提供依据。

本项目按照申请要求对肺炎衣原体Cpn0423通过Nod2/Rip2途径介导小鼠肺部炎症损伤及其作用机制进行了初步探讨。首先，本项目通过采用激光共聚集、信号通路阻断等多种手段研究发现，Cpn0423与衣原体III型分泌系统效应蛋白同源性为85%~93%；NOD2-siRNA能有效地抑制BMDM细胞中NOD2 mRNA的表达水平；Cpn0423诱导BMDM细胞分泌的细胞因子MIP-2和IL-6，NOD2-siRNA干扰后，MIP-2分泌水平显著降低（P<0.001)；鼻饲Cpn0423可以诱导小鼠肺部炎症损伤，免疫细胞募集，实验组小鼠肺泡灌洗中IL-6、TNF-α水平也显著高于对照组（P<0.001）；83.3%（10/12）Cpn阳性患者BALF中Cpn0423 DNA为阳性。此外，本研究对衣原体与HPV共感染的分子特征也进行了初步研究，不孕女性衣原体感染率为3.5%（23/666），体检中心及妇科门诊就诊女性患者衣原体感染率分别为3.8%（38/1006）、5.2%（175/3334），年龄和HPV感染是衣原体感染的危险因素（OR=0.496，P<0.001; OR=1.710，P<0.001），衣原体感染是女性宫颈低级别病变的危险因素（OR=3.25，P=0.018）。以上结果表明，Nod2/Rip2途径参与了肺炎衣原体Cpn0423介导的小鼠肺部炎症损伤。