间日疟原虫GAMA蛋白在侵入Duffy阳性/阴性红细胞的功能研究及其免疫学分析

疟疾是危害人类最严重的寄生虫病，而间日疟分布最为广泛，且防控难度大。PvDBP结合红细胞表面分子DARC曾被认为是间日疟原虫侵入红细胞的唯一途径。但最近发现Duffy阴性血型人群（DARC缺失）同样可感染间日疟，这提示间日疟原虫入侵红细胞可能存在PvDBP-DARC的替代途径。本项目前期发现：恶性疟原虫入侵红细胞关键的红细胞结合分子PfGAMA与其同源分子PvGAMA结构相似，表达位置相同，且同样具有结合一般红细胞的能力，因此PvGAMA很可能作为PvDBP-DARC的替代途径，在间日疟入侵Duffy阴性红细胞过程中发挥重要作用。因此，本项目拟通过检测PvGAMA与Duffy阳性/阴性两种红细胞的结合能力，并筛选出与之结合的红细胞膜表面结合受体，从而验证其作为PvDBP-DARC的替代途径的可能性。本研究将揭示间日疟原虫感染Duffy阴性红细胞相关分子机制，也可拓宽间日疟疫苗研新思路。

Plasmodium vivax is widely geographical malaria, and threatens almost 40% of the world’s population. P. vivax resists blood-stage infection results from absence of the Duffy (Fy) blood group at the surface of red blood cells (RBCs). Interestingly, resent studies from different malaria-endemic regions showed that P. vivax could infect Duffy-negative blood type people and cause clinical disease, suggesting that P. vivax may be involved into alternative erythrocyte invasion pathway(s). To well understand mechanism of P. vivax invasion, we are targeting PvGAMA which is the homologues of an important RBCs binding protein PfGAMA. Hence, we segmented PvGAMA, and expressed them by using wheat germ cell-free expression system. We clarified these proteins localization in parasite by immunofluorescence assay. We will analyze PvGAMA antigenicity and immunogenicity of these proteins, such as human sera response against antigens, serum titers, the percent of CD4+/ CD8+ positive immune cells, and Th1/ Th2 major cytokines levels of immunized mouse. Furthermore, Duffy +/- erythrocyte binding ability of these proteins will be tested that need to be expressed on the surface of COS-7 cell or HEK 293T cell incubating with erythrocytes. After binding assay, we term to screen the receptor(s) of PvGAMA from RBCs membrane using micro-array. Then protein-erythrocyte blocking assay of P. vivax will be processed using protein-specific antibodies. These results may explain the Duffy +/- erythrocyte infection by P. vivax parasite, and advance the development of P. vivax malaria vaccine as well.