HBV pgRNA剪接变异体的复制周期及其在慢性乙型肝炎疾病进展中的作用

申请人近年专注HBV RNA相关基础研究并在病毒学权威期刊发表论文。血清HBV RNA以pgRNA为主，可作为预测抗病毒治疗应答的重要指标。我们已发现剪接变异体是血清pgRNA的重要组成成分，并存在于病毒样颗粒中，但它在血清中大量存在的原因和机制不明。我们前期系统研究了pgRNA剪接变异体的胞内复制和分泌过程，揭示了逆转录合成DNA效率的不足可能是它大量存在于血清中的原因，并发现其血清水平是肝硬化的危险因素。本研究拟通过稳定表达细胞系、HepG2-NTCP和人原代肝细胞以及人源化肝脏小鼠模型，证明pgRNA剪接变异体复制形成的病毒颗粒能感染肝细胞并形成cccDNA，并在长期随访队列中探究它的血清水平与慢乙肝疾病进展间的关系。本研究将阐明pgRNA剪接变异体的复制周期及它的临床应用价值，是对现有HBV复制理论的补充和完善，有助于确立新的临床参考指标，扩大血清HBV RNA的临床应用。

Serum HBV RNA plays an important role in predicting responses of antiviral treatment. It has been demonstrated that pgRNA is the major type of serum HBV RNA. The applicant has published papers related to basic research of HBV RNA in a top journal of virology. We have found that splicing variants exist in virion-like particles, which is the main component of serum pgRNA, yet the reason and mechanism of their abundant presence in serum is still largely unknown. We have systematically studied the intracellular replication and secretion process of pgRNA splicing variants, it reveals that the insufficient efficiency of DNA synthesis through reverse transcription may account for their abundant presence in serum. We also found that the serum level of pgRNA splicing variants were closely related to the progression of liver cirrhosis. In the study, the stably expressed cell line, HepG2-NTCP, human primary hepatocytes and humanized liver mouse model will be used to prove viral particles formed by the replication of pgRNA splicing variants can infect liver cells and then generate cccDNA. Moreover, we will explore the correlation between serum pgRNA splicing variants level and the progression of chronic hepatitis B in a long-term follow-up cohort. This study will clarify the replication cycle of pgRNA splicing variants and its clinical application value. Our study could contribute to a better understanding of HBV life cycle, it also helps to establish a new clinical marker and broaden the area for clinical application of serum HBV RNA.