宿主CD44分子在弓形虫感染中的作用机制研究

Infection of Toxoplasma gondii is the result of interaction between parasite and host protein, and it is a hot spot in the T. gondii research. Recent studies have shown that CD44 has important physiological and pathological functions in the infection of various kinds of pathogens, including lymphocyte homing, cell proliferation and adhesion, inflammation and signal transduction. Infectcion with T. gondii can elevate the expression level of CD44 in parasite-infected macrophages, T cells , B cells, NK cells, thus affecting the adhesion, migration, and immune regulation of parasite-infected cells. However, the mechanism of that CD44 interacted with T.gondii is unclear. In this study, we compared the differences between the CD44-deficient mice and wild-type mice in the infection of Toxoplasma gondii in vivo and cellular level, containing changes in survival rate, clinlcal symptom, pathological changes,the number of parasites,reproduction and related cytokine downstream. And then,we compared the differences in protein expression during the infection of T.gondii in CD44-deficient mice and wild-type mice by protein two-dimensional electrophoresis and co-immunoprecipitation in vivo and cellular level. These studies can help us understand the mechanism of interaction between CD44 and T.gondii, and also provide a basis for immune defense mechanisms against Toxoplasma gondii infection in host.

弓形虫感染是虫体和宿主互作的过程，宿主在其中的作用及机理成为弓形虫研究的焦点。宿主CD44在多种病原体感染中具有重要的生物学功能，如淋巴细胞归巢、细胞黏附增殖、炎症反应、信号转导等。弓形虫的感染也引起巨细胞、T细胞、B细胞、白细胞以及NK细胞大量表达CD44，并影响染虫细胞的黏附、迁移及免疫调节。但和CD44在其它病原体感染中的生物学功能相比，CD44在弓形虫感染中的作用机制还了解的非常有限。本研究拟用CD44基因缺失型C57BL-6鼠作模型，在活体和细胞水平上研究缺失型小鼠对弓形虫（PRU株）感染的差异，包括生存率、临床症状、病理变化、虫体数目、繁殖速度及下游免疫相关细胞因子的变化，并通过双向电泳、免疫共沉淀等技术在活体和细胞水平上比较弓形虫感染正常和CD44缺失型鼠C57BL-6引起的宿主蛋白质表达的差异，阐明CD44在弓形虫感染中的作用机制，深化宿主抗弓形虫感染的免疫防御机制。

慢性临床弓形虫感染是引起严重脑炎的主要疾病。 CD44是一个细胞粘附分子家族的成员，在弓形虫感染中起重要作用。然而，蛋白质组学的改变慢性弓形虫感染期间CD44的报道很少。因此，本项目进行了基于RNA-seq的比较转录组分析和基于iTRAQ的蛋白质组学研究，以筛选出慢性弓形虫感染期间CD44相关的基因与蛋白。结果表明总共35,908，54,428，51,473和22,387差异基因通过转录组注释在KOG，Swissprot，GO和KEGG数据库中；总共鉴定和定量出了2612种蛋白质。根据结果，CD44-小鼠，野生型小鼠，感染弓形虫的野生型小鼠和CD44-刚地弓形虫感染的小鼠之间的免疫相关基因和蛋白发生了显着变化比较，与C57小鼠相比，在CD44-小鼠中免疫球蛋白，主要组织相容性复合体（MHC）II类抗原，趋化因子配体和干扰素（IFN）诱导型GTPase家族等基因显著下调。另外，Ca2 +和代谢相关蛋白在CD44-小鼠感染弓形虫脑组织中上调，这支持了CD44介导产生宿主Ca2+和IFN-γ，并且弓形虫优先侵入表达高水平CD44的细胞的假说。本研究结果证实并能更全面地了阐明CD44在慢性感染弓形虫的宿主中的作用，为CD44在慢性弓形虫病中的功能研究提供了新思路。