HPS OmpP2膜外结构环诱导PAMs IL-8 mRNA转录的研究

Loops of OmpP2 Porin from Haemophilus parasuis InducesIL-8 mRNA Expression in PAMs

OmpP2是副猪嗜血杆菌的致病因子,也是其重要的免疫原性蛋白。为探明OmpP2蛋白8个膜外结构环(Loop)在HPS感染过程中的促炎作用,本试验以猪肺泡巨噬细胞(Porcine Alveolar Macrophages,PAMs)为细胞模型,以HPS血清5型标准菌株OmpP2蛋白、HPS血清5型标准菌株全菌体及牛血清白蛋白(BSA)作为参照,通过Real-Time PCR的方法研究OmpP2 8个膜外结构环诱导PAMs产生炎性应答的作用。结果显示,与空白对照组相比,HPS血清5型标准菌株OmpP2蛋白膜外结构环均能诱导PAMs细胞使其促炎细胞因子IL-8的转录水平出现不同程度的上调,表明它们在HPS感染过程中参与了宿主肺泡巨噬细胞的促炎性免疫应答;在8个膜外结构环中,Loop7(2^(-△△Ct)≈8,P<0.05)表现出非常显著的刺激活性,远远高于其他膜外结构环组,并与HPS血清5型标准菌株OmpP2蛋白组的诱导上调幅度(2^(-△△Ct)≈12(P<0.05))最接近,表明Loop7在OmpP2蛋白诱导PAMsIL-8促炎性免疫应答过程中扮演着重要角色,结果暗示Loop7很可能是OmpP2致宿主肺巨噬细胞促炎性功能活跃区。本试验为进一步了解OmpP2蛋白功能及了解HPS诱导宿主细胞炎症反应分子机制提供实验基础。

Outer membrane protein P2（OmpP2） is the pathogenic factor and the important immunogenicity protein of Haemophilus parasuis（HPS）.In order to study the role of loops of HPS 0mpP2 porin in the inflammatory process,porcine alveolar macrophages（PAMs） was used as a cell model in the present study.Interleukin 8 IL-8 induced expression by Loop1-Loop 8 of H.parasuis type 5 standard strains OmpP2 in PAMs was analyzed by real-time PCR.Our result showed that,all loops of H.parasuis type 5standard strains OmpP2 porin could trigger cytokine mRNA expression in PAMs in vitro.It demonstrated that loops of HPS type 5standard strains OmpP2 was involved in the inflammatory reaction of the host cell.Among loops,Loop7（2^-ΔΔCt≈8,P〈0.05） showedsignificant stimulation,much higher than other loops and closing to the OmpP2（2^ΔΔCt≈11,P〈0.05）of HPS standard type 5 strains.These results suggest that Loop7 is likely to be an active function region of the OmpP2 porin.This study provides further understanding of OmpP2 porin function.