特异性T细胞在抗流感病毒感染交叉免疫保护中的作用研究

The outbreaks of infectious disease caused by influenza A virus has been reported more and more frequently and severely in recent years. Thus, developing cross-protective influenza vaccines against the spread of a new influenza virus is an important strategy against pandemic emergence. Recent studies suggest an important role for CD8+ T cells in driving protection against IAVs, especially the newly-emerged viruses with pandemic potential, thus identification of new vaccine targets manifesting T cell mediated response may serve as an ideal approach for influenza vaccine. In other to research CD8 T Cell-mediated cross-protection against influenza virus, the present study involves the application of an immunoinformatics-based consensus approach for HLA-A2-restricted and HLA-A2-restricted epitope prediction of several subtypes of influenza virus epidemic frequently and severely in recent years. And followed in vitro refolding of MHC/peptides and the immunogenicity and cross-protection activity were further determined in TAP and HLA-A2/A11 transgenic mice. Identify and characterize novel immunogenic CD8+ T cell epitopes, and also analysis their longevity and functionality in order to rationally design prime-and-boost strategies. These studies may contribute to the further understanding of influenza virus specific cellular immunity and the development of relevant vaccines.

多种流感病毒亚型开始不断突破种间屏障感染人类，具有造成大流行的潜能。因此亟待开发一种新型流感疫苗以应对流感跨种传播及多亚型并存的现状。基于此，我们对特异性T细胞在抗流感病毒感染交叉免疫保护中的作用进行研究，首先通过生物信息学的方法对我国主要的HLA-A类型HLA-A2和HLA-A11限制性的流感病毒全蛋白质CTL表位进行预测，并通过体外HLA分子折叠试验、细胞结合试验全面筛选CTL表位；在此基础上，查明以病毒感染和gp96为佐剂的流感裂解苗免疫人TAP和HLA-A2或A-11转基因小鼠后，活化了哪些特异性T细胞，特别是保守表位特异性T细胞，解析表位特异性T细胞在流感病毒感染交叉保护中的作用。本研究研究不仅全面揭示流感病毒特异性T细胞应答在交叉保护中的作用机制，还可为设计新型流感病毒T细胞佐剂疫苗提供新的思路，对研究可预防多种亚型流感病毒感染、提供机体长久免疫保护的通用流感疫苗提供理论基础。

疫苗免疫依旧是预防流感的主要方法。但是，目前使用的疫苗只对特定的病毒株具有保护效力，一旦出现流感病毒新亚型或新毒株，现有疫苗就会失去其保护效力。因此，需要针对多种流感亚型提供广泛保护的新疫苗。在本研究中，我们证明gp96佐剂的季节性流感单价H1N1裂解疫苗在BALB/c小鼠模型中可诱导对不同亚型的甲型流感病毒的交叉保护。在gp96的参与下使疫苗的免疫应答倾向Th1型，与单独的裂解疫苗相比，具有明显更强的抗原特异性T细胞应答。用gp96佐剂疫苗免疫引起明显的针对不同流感病毒株的保守表位的交叉反应CD8+T细胞应答。值得注意的是，来自病毒结构蛋白的6个高度保守的CD8+T表位（包括KD限制的HA437-445、M131-39、NP39-47、NP147-155、NP218-226和PB181-89）在gp96介导的交叉保护中起主要作用。这些交叉反应的CD8+T细胞可能从接种疫苗后建立的记忆细胞池中召回，并从肺外部位招募，以促进病毒清除。比较分析表明，大多数由异源病毒感染诱导的保守表位特异性CTL也被gp96佐剂疫苗特异性激活，从而产生更广泛的保护性CD8+T细胞应答。我们的结果证明了在现有季节性流感疫苗中加入gp96蛋白的优势，为疫苗提供更好交叉保护的能力。