肺炎衣原体假定蛋白Cpn0423通过NOD2信号途径诱导炎症反应的作用机制

Contribution of NOD2 signal pathway to Chlamydia pneumonia Cpn0423-induced inflammatory response

目的了解肺炎衣原体(Chlamydia pneumoniae,Cpn)假定蛋白Cpn0423的生物学活性及其诱导宿主细胞炎症反应的作用途径。方法生物信息学软件初步分析Cpn0423的生物学特性,激光共聚焦显微技术检测核苷酸结合寡聚化结构域样受体2(nucleotide-binding oligomerization domain-like receptor 2,NOD2)在骨髓源性的巨噬细胞(bone marrow-derived macrophages,BMDMs)中的亚细胞定位。采用NOD2-siRNA干扰试验检测其抑制BMDMs细胞中NOD2的表达水平,ELISA法检测Cpn0423诱导BMDMs细胞分泌的巨噬细胞炎性蛋白2(MIP-2)和IL-6,PCR法检测Cpn阳性患者支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)中Cpn0423 DNA水平。结果Cpn0423与衣原体Ⅲ型分泌系统效应蛋白同源性为85%~93%。NOD2-siRNA能有效抑制BMDMs细胞中NOD2 mRNA的表达水平。NOD2-siRNA干扰后,Cpn0423诱导BMDMs细胞分泌的细胞因子MIP-2[(920.5±99.1)pg/ml vs(130.1±11.5)pg/ml,P<0.001]和IL-6[(266.2±58.4)pg/ml vs(165.7±21.5)pg/ml,P<0.001]水平显著降低。83.3%(10/12)Cpn阳性患者BALF中Cpn0423 DNA为阳性,对照组未检出Cpn0423 DNA。结论Cpn0423通过NOD2途径诱导宿主细胞免疫应答,与Cpn引起机体慢性炎症损伤密切相关。

Objective To understand and determine the biological properties of Chlamydia pneumonia(Cpn)hypothetical protein Cpn0423 and the mechanisms of which involved in Cpn0423-induced inflammatory response.Methods The biological properties of Cpn0423 gene were analyzed using bioinformatic software.The subcellular localization of nucleotide-binding oligomerization domain-like receptor 2(NOD2)in bone marrow-derived macrophages(BMDMs)was detected by confocal microscope.NOD2-siRNA was used to inhibit the expression of NOD2 at mRNA level.Cpn0423-induced macrophage inflammatory protein 2(MIP-2)and IL-6 production in BMDMs were detected by ELISA.PCR was performed to detect Cpn0423 DNA in bronchoalveolar lavage fluid(BALF)of Cpn-positive patients.Results The homology between Cpn0423 and other typeⅢsecretion system effector proteins of Chlamydia ranged from 85%to 93%.NOD2-siRNA could effectively inhibit the expression of NOD2 at mRNA level in BMDMs(P<0.001).Moreover,Cpn0423-induced production of MIP-2[(920.5±99.1)pg/ml vs(130.1±11.5)pg/ml,P<0.001]and IL-6[(266.2±58.4)pg/ml vs(165.7±21.5)pg/ml,P<0.001]in BMDMs were decreased following NOD2-siRNA pre-treatment.Cpn0423 DNA was detected in the BAlF of 83.3%(10/12)of Cpn-positive cases,but not in Cpn-negative cases.Conclusions Cpn0423 induced inflammatory response in host cells through NOD2 pathway,which was closely related to the chronic inflammatory injury caused by Cpn.