纤维胶凝蛋白激活的补体凝集素途径抵抗弓形虫感染及机制研究

Toxoplasmosis, caused by Toxoplasma gondii, is widely spread in warm-blooded animals and human beings. The drugs that currently used to treat toxoplasmosis can only kill the tachyzoites but not able to cure toxoplasmosis completely.Therefore, there is an urgent need to find new anti-Toxoplasma drugs or drug targets.Previous works had shown that the ficolin could activate MBL pathway and regulate the pathogen phagocytosis of macrophages by recognizing the glycoproteins or sugar side chains on the surface of pathogens (viruses, bacteria and fungi). It was widely accepted that macrophages played an important role in Toxoplasma gondii infection defense, but the mechanism of the complement system activation in defending Toxoplasma gondii infection has not been clarified. The protective effect for ficolin molecules in anti-toxoplasma infection and the sugar chains or glycoproteins on Toxoplasma gondii cell surface that can be recognized by ficolin molecules are still unclear.This study intends to evaluate the protective effect of ficolin on Toxoplasma gondii infected-hosts/host-cells in the mice/cell model by expressing both human’s and mice’s ficolin genes using the prokaryotic and eukaryotic expression vectors respectively, and then evaluate the possibility of developing ficolin as a new anti-Toxoplasma drug. In addition, this study will also provide us a new idea in developing new preventive or therapeutic strategies for other Apicomplexa parasites.

弓形虫病由刚地弓形虫引起，是一种在世界范围内广泛流行的人兽共患寄生虫病，一线抗弓形虫药仅仅能够压制弓形虫血症并不能够彻底治疗弓形虫病；故迫切需要寻找新的抗弓形虫药物或药物靶点。纤维胶凝蛋白(ficolin)能够通过识别病原（病毒、细菌和真菌）表面的糖链或糖蛋白激活MBL途径、调理巨噬细胞对病原的吞噬作用，有作为抗病原药物潜力；巨噬细胞在抵抗弓形虫感染中的重要作用已获公认，但激活补体系统抗弓形虫感染机制尚未厘清，且ficolin在抗弓形虫感染中的保护作用和识别弓形虫细胞上膜糖链或糖蛋白的种类也没有阐明。本研究构建人和小鼠的ficolin原核/真核载体表达获得重组ficolin蛋白，进行体内/体外试验，研究ficolin蛋白保护宿主或细胞抵御弓形虫感染作用，寻找弓形虫细胞上ficolin所识别膜糖链或糖蛋白，为开发新的抗弓形虫药物提供立论依据，同时也能够为其他顶器门病原生物的防控提供新的思路。

项目在体外/体内模型中的研究，发现ficolin A 蛋白保护宿主或宿主细胞抵御弓形虫感染作用中的直接证据，阐明ficolin A 蛋白作为固有免疫中重要分子与弓形虫之间在感染发生时相互作用的机制。主要结论如下：（1）验证了ficolin A能够调节巨噬细胞对弓形虫的吞噬作用，并探讨了不同糖链分子对ficolin A结合弓形虫的影响。（2）探讨了ficolin A结合弓形虫主要环境条件的最适范围，并证实了ficolin A能够识别弓形虫膜表面抗原，进而同弓形虫结合，其中SAG1和SAG2可能是比较重要的膜表面抗原。（3）利用动物实验进一步证实了，ficolin A可通过提升Th1型细胞免疫水平、进而显著地提高巨噬细胞对弓形虫的吞噬作用实现对宿主的部分保护力；并且，C3分子在宿主细胞膜表面的沉积能够启动MBL 补体激活途径，抵抗弓形虫的感染。为基于ficolin A的干预弓形虫感染的策略提供实验依据，也为开发新的抗弓形虫药物提供新的科学思路。