幽门螺杆菌黏附素HLA-DRB1限制性CD4+T细胞表位的预测与鉴定  

Identification of HLA-DRB1 restricted CD4^+ T-cell epitopes from HpaA of Helicobacter pylori

作　　者：胡健[1] 郭红[1] 李滨[2] 陈立[2] 何亚非[1] 孙合强[2] 李海波[2] 赵卓[2] 吴超[2] 

机构地区：[1]第三军医大学新桥医院消化内科,重庆400037 [2]第三军医大学医学检验系临床微生物及免疫学教研室,重庆400038

出　　处：《免疫学杂志》2014年第2期124-128,132,共6页Immunological Journal

基　　金：国家自然科学基金(81072493);重庆市自然科学基金(2011BB5043);"十一五"国家重大新药创制专项(2009ZX09102-220)

摘　　要：目的针对中国汉族高频HLA-DRB1基因型人群,预测与鉴定幽门螺杆菌(Helicobacter pylori,H.pylori)黏附素蛋白A亚单位(neuraminyllactose-binding hemagglutinin,HpaA)CD4+T细胞表位。方法通过Allele Frequency Net数据库分析中国汉族人群HLA-DRB1高频基因型,利用NetMHCIIpan对HpaA蛋白中能被这些高频基因型递呈的CD4+T细胞表位进行预测;通过PCR-SBT法筛选携带有这些高频基因型的H.pylori感染者,利用重组HpaA抗原刺激感染者外周血单个核细胞体外扩增出HpaA抗原特异性T淋巴细胞后利用合成短肽对所预测表位进行鉴定,通过抗体阻断实验对表位的HLA限制性进行分析,利用DC细胞负载重组HpaA蛋白对表位的自然递呈特征进行分析。结果中国汉族人群中HLA-DRB1高频基因型为DRB1*0901(14.4%)、DRB1*1202(13.3%)和DRB1*1501(10.8%),针对上述3种高频基因型预测出24个HpaA蛋白CD4+T细胞表位。其中,4个表位:HpaA39-53(HLA-DRB1*0901)、HpaA42-56(HLA-DRB1*0901)、HpaA89-103(HLA-DRB1*1202)和HpaA87-101(HLA-DRB1*1501)可有效刺激体外扩增的HpaA特异性T细胞产生高水平IFN-γ,且均能被DC细胞自然递呈。结论鉴定得到的4个HLA-DRB1限制性CD4+T细胞表位有可能成为H.pylori表位疫苗设计的候选抗原。This study designed to identify the immunodominant CD4＋ T cell epitopes derived from HpaA of Helicobacter pylori （H. pylori） which restricted to the most popular HLA molecules of Chinese Han population. Firstly, The frequencies of HLA-DRB1 alleles of Chinese Han population was analyzed by using Allele Frequency Net data base, and CD4＋ T cell epitopes derived from HpaA protein were predicted by using NetMHCIIpan. Then, H. pylori infected subjects expressing the most popular HLA molecules were identified by PCR-SBT, and HpaA- specific T cell lines expanded in vitro by using the recombinant HpaA protein, and peripheral blood mononuclear ceils of H. pylori infected subjects were used to identify the immunodominant epitopes. Furthermore, HLA blocking assay was used to analyze the HLA restrictions, and DCs pulsed recombinant HpaA protein were used to confirm be natural presenting characteristics of the epitopes. Result showed that the most popular three HLA alleles in Chinese Han population were DRBl＊0901（14.4%）, DRBl＊1202（13.3%） and DRBl＊1501（10.8%）, and 24 epitopes restricted to these three HLA-DRB1 molecules were predicted, in which HLA-DRBl＊0901/HpaA39_53, HLA-DRB 1 ＊ 0901/HpaA42-56, HLA-DRB 1 ＊ 1202/HpaAsg-103 and HLA-DRB 1 ＊ 15 01/HpaA87-101 aroused the strongest HpaA-specific T cells responses of IFN-＋/ production in vitro. And all of the four epitopes could be naturally presented by DCs. In conclusion, these four CD4＋ T cell epitopes identified here might be used as candidate antigens for H. pylori epitope vaccine design.

关 键 词：幽门螺杆菌 CD4+T细胞表位 幽门螺杆菌鞭毛黏附素A亚单位 人白细胞抗原-DRB1 

分 类 号：R378[医药卫生—病原生物学]