血吸虫感染中CTL效应与CD8α+DCs分化成熟关系的研究

CD8+ cytotoxic T lymphocytes (CTL) contribute to immunity against Schistosoma japonicum infection through mechanisms yet to be elucidated. It awaits further investigation whether vaccine designs should include strategies to induce CTL responses. Dendritic cells (DCs) are crucial components in the cascade of immune responses including CTL activation. DCs in mouse spleen can be divided into CD8α+ and CD8α- subpopulations, CD8α+DCs have specialized functions to produce high amounts of IL-12 and IFN-γ, and to active MHC classⅠ-restricted CTL responses to exogenous antigens by cross-priming. It is not clear if schistosomal antigens can also be cross-presented to and activate CD8 + T cells by CD8α+ DCs. However, the functions of CD8α+DCs could be modulated by their activation state, the nature of the antigens, the concentration of the antigen and special cytokine milieu. Our previous studies suggested that during schistosome infection CD8α+DCs and CD8α-DCs ratio changed with different antigen components in the environment. In our proposed study, we plan to further investigate the key role of CD8α+DCs during schistosome infections, its function of cross-priming CD8+T cells to participate in the immunity and the abilities of different schistosome antigens to influence DCs differentiation and polarize immature DC maturation towards a specific subtype. This study may provide theoretical guidance and experimental evidence to the rational design of vaccines against schistosomes, and deepen our understanding of the important roles of DCs and CD8+T cells during helminth infection.

CD8+T细胞在抗血吸虫感染免疫中发挥功能的具体机制尚不清楚，但可发挥作用已较明确。研究证实成熟CD8α+DCs在交叉递呈活化CD8+T细胞方面具有独特作用。血吸虫抗原作为外源性抗原是否可通过CD8α+DCs交叉递呈激活CD8+T细胞仍需实验证据。研究表明DCs亚群对抗原识别递呈方式及诱导应答类型与其分化成熟状态有直接关系。本申请前期研究提示CD8α+/-DCs变化与血吸虫感染后抗原环境变化相关。血吸虫是否通过抗原环境改变影响CD8α+/-DCs分化成熟及功能从而调控整个免疫应答方向尚待验证。本研究拟使用CD8α+DCs缺失小鼠日本血吸虫感染模型探索感染后CTL效应产生的分子机制，体内外实验观察血吸虫不同抗原对CD8α+/-DCs亚群分化及成熟阶段递呈、交叉递呈功能的影响，阐明CD8α+/-DCs分化成熟及功能和特定抗原环境相关，为抗血吸虫疫苗设计包括CTL作用提供新思路及相应实验基础。

CD8+T 细胞在抗血吸虫感染免疫中发挥功能的具体机制尚不清楚，但可发挥作用已较明确。研究证实成熟CD8α+DCs 在交叉递呈活化CD8+T 细胞方面具有独特作用。本研究首先建立了体外Flt3L刺激快速制备大量的具有免疫活性的未成熟CD11c+CD8α+BMDCs的方法，为后续的实验室研究，动物实验以及临床研究奠定了基础；第二，本研究发现SEA、SWAP刺激未成熟CD8α+DCs可检测到显著表达的CD86，SEA刺激组同时检测到高表达的CD40、CD80、MHCI、MHCII，SWAP刺激组同时检测到高表达的CD40、MHCI、MHCII，表明血吸虫抗原刺激活化了CD8α+DC，不同性质的血吸虫抗原可能通过不同的方式影响DCs成熟活化并可能进一步影响其递呈及交叉递呈功能；第三，本研究发现SEA刺激组CD8α+DCs细胞表面TRL1,TLR2, TLR5, TLR6, TLR11表达显著高于CD8α-DCs细胞，CD8α-DCs细胞表面TLR4表达显著高于CD8α+DCs细胞；SWAP刺激组CD8α+DCs细胞表面TLR11表达显著高于CD8α-DCs细胞，CD8α-DCs细胞表面TLR6表达显著高于CD8α+DCs细胞。提示血吸虫不同性质抗原可能进一步通过调节不同CD8α+/-DCs亚群不同Toll样受体表达影响DCs功能从而影响感染的结局；第四，本研究观察了CD8α+DCs-/-亚群缺失小鼠在日本血吸虫感染过程中免疫应答情况。结果显示CD8α+DCs-/-组单个虫卵肉芽肿面积显著低于野生型C57BL/6J组小鼠，提示在血吸虫自然感染过程中，CD8α+DCs亚群的缺失会减轻小鼠肝脏病理损伤。CD8α+DCs-/-组Tc1在3w,6w,9w均显著高于C57BL/6J组，提示CD8α+DCs-/-缺失的情况下，血吸虫产卵后宿主体内仍维持了较强的1型免疫反应以及活化的CD3+ CD8+IFN-γ+细胞，为抗血吸虫疫苗设计包括CTL作用提供新思路及相应实验基础,深化了对CD8+T应答在蠕虫感染中作用的理解。