Pgp3促进衣原体在小鼠生殖道内致病的机制研究

chlamydia trachomatis(C.t)urogenital tract infection is the major cause of female tubal infertility,abortion and ectopic pregnancy. Recent studies showed that chlamydia plasmid encoded protien pgp3 play an important role in promoting chlamydia propagating, ascending infection and oviduct pathology development. The mechanism remains unknown. Our previous work has demonstrated that antimicrobial peptide LL-37 can kill chlamydia in vitro and pgp3 inhibit LL-37’s killing ability by binding to LL-37 . Nevertheless, pgp3 also stimulate macrophages and dendritic cells to release pro-inflammatory cytokines which play an important role in oviduct pathology. The structure analysis revealed that the C-terminal of pgp3 is very similar with the binding domain of TNF-α which means that the C-terminal of pgp3 may directly bind to TNFR1 and activate the relative signaling pathway, inducing inflammation and apoptosis. Based on these findings, a project using molecular cloning technique, GST-pull down assay and native-gel to explore the possible pathogenic mechanism of pgp3. To further testify the mechanism, genetic recombination will be used to construct chlamydia muridarum strains with either pgp3C or pgp3M fragment deletion. CRAMP KO mice, MyD88 KO mice and TNF-α KO mice will be infected with different chlamydia mutant strains to testify the in vitro results. The aim of this project is trying to reveal the pathogenic mechanism of pgp3, providing a new strategy for controlling tubal infertility caused by chlamydia.

沙眼衣原体（chlamydia trachomatis, C.t）泌尿生殖道感染是女性输卵管性不孕的重要原因之一。最新研究发现衣原体质粒编码蛋白pgp3可促进衣原体繁殖、上行感染及输卵管病变形成，但其机制尚未明确。申请者发现抗菌多肽LL-37在体外可以杀伤衣原体，pgp3可结合LL-37并抑制其杀伤功能；pgp3还可刺激巨噬细胞及树突状细胞释放促炎症因子IL-6及MIP-2；Pgp3的C段结构与肿瘤坏死因子α（TNF-α）受体结合区域相似，该段可能直接结合TNF-α受体引发炎症反应。本项目拟通过分子克隆表达、GST-pull down、Native-gel等技术探索pgp3致病机制；利用基因重组技术构建pgp3 C段或M段被敲除的衣原体菌株，感染CRAMP、MyD88及TNF-α敲除小鼠加以验证。本项目的完成试图揭示pgp3在衣原体致病中的作用机制，为治疗C.t所致输卵管性不孕提供新思路。

沙眼衣原体（chlamydia trachomatis, C.t）泌尿生殖道感染是女性输卵管性不孕的重要原因之一。最新研究发现衣原体质粒编码蛋白pgp3可促进衣原体繁殖、上行感染及输卵管病变形成，但其机制尚未明确。本项目旨在阐明pgp3对输卵管的致病机制。项目组通过体外细胞培养及动物实验发现pgp3可以通过其中间段（67-132）结合MyD88调控的人类抗菌多肽从而促进衣原体在下生殖道的繁殖及上行感染；将原核表达的pgp3经卵巢囊内注射后可引起输卵管及卵巢的炎性反应，其机制是通过pgp3 C段（133-260）结合TNF-a来抑制宿主细胞的凋亡并促进上生殖道的炎性反应从而导致输卵管的病变。.考虑到pgp3是衣原体致病的重要毒力因子，靶向抑制pgp3可能会降低衣原体的致病性，为了验证这个推测，项目组制备了抗pgp3的单克隆抗体，经腹腔注射及阴道给药的方式干预鼠肺炎沙眼衣原体感染的小鼠，结果发现两组小鼠下生殖道的衣原体繁殖能力及对上生殖道的致病性均显著减弱，提示抗pgp3的单克隆抗体对衣原体感染有治疗作用。.本项目还探索了衣原体膜蛋白OmcB全长及C段的免疫原性，结果发现OmcB C段与佐剂CpG联合经腹腔注射免疫小鼠后，鼠肺炎沙眼衣原体在小鼠下生殖道的繁殖能力及对上生殖道的致病能力均显著减弱，提示OmcB C段具有良好的免疫保护作用，可作为 C.t的候选疫苗之一。.本项目的所有发现目前国内外均无相关报道，这为临床上治疗C.t感染及C.t疫苗的研发提供了理论依据。