肝细胞自噬对乙型肝炎病毒感染抗原交叉提呈免疫应答的影响及机制研究

Current literature showed that chronic HBV infection is often accompanied by low immune response or even immune tolerance to HBV antigens. Our previous research showed that the cross-presentation of tumor antigen is closely related with autophagy. The efficiency of cross-presentation is greatly enhanced by the induction of autophagy. However, it is unknown how the cross-presentation of HBV antigen is affected by autophagy of hepatocytes infected by HBV. This project aims to establish cell models that mimic HBV infection on the basis of the defect of autophagy ex vivo; to determine whether hepatocyte specific deletion of autophagy gene could affect HBV infection and CTL immune response to HBV antigens; to analyze the immune responses of antigen-specific T cells cross-primed by the antigen derived from hepatocytes infected with HBV. Results from this project will have important implication for the development of novel strategies for the generation of strong anti-HBV immunity and effective treatment of HBV chronic infection.

HBV慢性感染者处于特异性免疫应答低下或免疫耐受状态，缺乏有效的抗原提呈。我们在肿瘤的研究中发现，肿瘤抗原的交叉提呈与自噬密切相关，采用自噬诱导剂处理肿瘤细胞可以提高交叉提呈的效率。HBV感染肝细胞自噬是否具有调节HBV抗原的交叉提呈，增强HBV特异性CTL应答尚不清楚。本项目建立肝细胞自噬不同条件下（自噬缺陷和正常）的乙肝感染细胞模型，将其作为抗原供给细胞，联合DC进行体外抗原交叉提呈实验。旨在研究HBV感染相关的肝细胞自噬对HBV抗原交叉提呈的作用，阐明干预肝细胞自噬对抑制HBV复制和增强特异性CTL应答的机理，为打破机体对慢乙肝的免疫耐受、活化特异性CTL应答，诱导有效清除乙肝病毒感染寻找新方法。

HBV慢性感染者处于特异性免疫应答低下或免疫耐受状态，缺乏有效的抗原提呈。我们在肿瘤的研究中发现，肿瘤抗原的交叉提呈与自噬密切相关，采用自噬诱导剂处理肿瘤细胞可以提高交叉提呈的效率。HBV感染肝细胞自噬是否具有调节HBV抗原的交叉提呈，增强HBV特异性CTL应答尚不清楚。本项目构建了来源于HepG2.215细胞的自噬小体疫苗（HBV+ DRibbles），检测了其在急慢性HBV感染小鼠模型中的抗HBV作用，发现HBV+ DRibbles能够交叉激活体内HBV抗原特异性T细胞反应，可作为预防性疫苗诱导机体的HBV保护性免疫，作为治疗性疫苗治疗急慢性HBV感染。随后，我们发现包含CMV抗原DRibbles能够激活人外周血中记忆性CMV抗原特异性T细胞，在这一过程中，CD141+ DC是PBMC中能够最有效交叉提呈UbiLT3 pp65 DRibbles，激活CMV pp65抗原特异性CD8+ T细胞的抗原提呈细胞。最后，我们比较了IL-4 DC及IFN-α DC负载自噬小体疫苗后激活人抗原特异性T细胞的活性，结果提示与IL-4 DC相比，IFN-α DC负载HEK293T CEF DRibbles后能更有效激活人CEF抗原特异性CD8+及CD4+ T细胞。我们的研究提示DRibbles可运用于病毒感染的治疗，为打破机体对慢乙肝的免疫耐受、活化特异性CTL应答，诱导有效清除乙肝病毒感染提供了一种新方法。