旋毛虫副肌球蛋白H-2d限制性Th表位的鉴定及Th-B表位肽嵌合疫苗免疫学效应研究

Trichinellosis is a popular food-borne zoonosis which is threatening human health and causes great loss of livestock production. Our group has reported a newfound promising vaccine canditate, paramyosin (Pmy) of Trichinella spiralis (Ts-Pmy), exhibits protective immunity effect in mice. It has been studied that host humoral immunity is important in the elimination of the parasite. Furthermore, it is necessary to indentify T helper (Th) epitope of Ts-Pmy to specify its immune response caused accordingly. Based on BALB/c mice model, we are planning to predict H-2d restricted Th epitopes of Ts-Pmy by bioinformatics approach and prepare those epitope peptides by solid-phase procedure. Their function will be verified through CD4+ lymphocyte proliferation assays and cytokine production induced by the peptides. New-discovered Th2 epitope(s) will then be constructed in tandem with the protective B epitope of Ts-Pmy which has been identified by our group previously. Based on that, a chimeric Th-B vaccine is able to be constructed. The humoral and cellular immune responses against this multiple antigen peptide (MAP) will be studied in BALB/c mice. Larvae burden reduction will be calculated to evaluate the protective immunity of this new antigen in Trichinella spiralis challenged animals. If the antigen is confirmed to work, it will establish the foundation for epitope vaccine development against trichinellosis which is under exploration for years.

旋毛虫病是一种常见的人畜共患寄生虫病，严重危害人类健康和畜牧业生产。旋毛虫副肌球蛋白（Ts-Pmy）是本课题组新发现的一个具有应用前景的抗旋毛虫病疫苗候选抗原分子，其重组蛋白对免疫小鼠具有一定保护性。宿主的体液免疫对旋毛虫的清除十分重要，因此鉴定Ts-Pmy 的Th表位并研究其特异性的免疫应答特征具有重要的意义。本课题拟以BALB/c小鼠为模型，首先采用生物信息学预测Ts-Pmy的H-2d限制性Th表位，体外固相合成表位肽，然后结合CD4+T淋巴细胞功能实验和细胞因子检测等方法确定Th2表位；然后将筛选出的Th2表位和本课题组前期鉴定的Ts-Pmy中一个具有保护性的B细胞表位串联构建Th-B表位肽嵌合疫苗；将此多抗原肽（MAP）疫苗免疫BALB/c小鼠，分析其所引起的细胞和体液免疫反应；攻虫后计算减虫率，评估疫苗的免疫效果。本课题将为旋毛虫病表位疫苗的设计提供实验依据和奠定基础。

旋毛虫病是一种常见的人畜共患寄生虫病，因进食感染的猪肉或其它肉类而传播。在中国，通过给家畜注射疫苗阻断该病的传播是一项切实有效的措施。本课题组前期研究发现旋毛虫副肌球蛋白（Paramyosin of Trichinella spiralis, TsPmy）是一个具有应用前景的抗旋毛虫病疫苗候选抗原分子，其重组蛋白对免疫小鼠具有一定保护性。宿主的体液免疫对旋毛虫的清除十分重要，因此鉴定TsPmy 的辅助性T细胞表位并研究其特异性的免疫应答特征具有重要的意义。本课题以BALB/c小鼠为模型，首先利用SYFPEITHI数据库预测了TsPmy的H-2d限制性Th表位，选取得分最高的12个Th表位肽进行合成，然后结合CD4+T淋巴细胞功能实验和细胞因子检测等方法确定4个Th2表位；然后将筛选出的Th2表位和本课题组前期鉴定的TsPmy中一个具有保护性的B细胞表位串联构建Th2-B表位肽嵌合疫苗并在大肠杆菌中进行表达，将此纯化后的多表位蛋白（recombinant multi-epitope protein, rMEP）疫苗免疫小鼠，攻虫后获得了55.4%的肌幼虫减虫率。脾淋巴细胞增殖实验、抗体亚型分析及细胞因子检测显示rMEP诱导小鼠产生了以Th2型免疫反应为主的混合性Th1/Th2反应。本课题为抗旋毛虫病表位疫苗的设计提供了实验依据。