ZHANG Zheng-mao1, TIAN Yong-jun, YANG Yan, WANG Bao-ju, GUO Pei-xuan, YANG Dong-liang1**
(1.Division of Clinical Immunology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030,
China; 2. Department of Pathobiology and Purdue Cancer Research Center, Purdue University, West Lafayette, IN 47907, USA))
Abstract：The objective of our present study is to explore the potential use of pRNA as a bio-carrier of
siRNA to inhibit HBV gene expression and replication. After co-transfected with pHA-HBs into 293T
cells, HBVsi18-42, a pRNA-escorted siRNA, suppressed HBsAg and accumulated in the cells in a
dose-dependent fashion. HBVsi18-42 substantially inhibited HBV gene expression and replication
initiated by pHBV1.3 in HepG2 cells. In hydrodynamic injection mouse model, Balb/cJ mice were
co-injected with pHBV1.3 and HBVsi18-42. Serum concentrations of HBsAg were analyzed by ELISA
on days 1, 2, and 3 post-injection. HBV core protein in mouse liver was visualized by
immunohistochemical staining. The results showed that the HBsAg levels in mice sera were reduced by
60%~90% in consecutive days relative to the control group, and the number of HBcAg positive
hepatocytes in the mouse liver sections was decreased substantially by 79.1%. Our preliminary data
showed that pRNA could be used as bio-carrier for the delivery of siRNA to knock down HBV gene
expression and repress viral replication.
Key words：RNAi；pRNA；Hepatitis B virus
摘要：为了研究由pRNA 携带的siRNA（HBVsi18-42）所介导的RNAi 过程能有效地抑制HBV 的基因表达和病
毒复制，我们利用细胞模型和高压注射小鼠模型评价HBVsi18-42 对HBV 复制和基因表达的抑制作用。通过
Western 印迹检测细胞内的HBsAg 含量，用ELISA 检测细胞培养上清和小鼠血清中的HBsAg 水平，采用Southern
印迹检测HBV 的复制中间体，通过免疫组织化学检测肝组织切片中HBcAg 的表达情况。试验结果显示，
HBVsi18-42 能以剂量依赖的方式在293T 细胞中抑制HBsAg 的表达以及在HepG2 细胞中下调病毒HBsAg 和
HBeAg 的表达和病毒复制中间体的水平。在小鼠模型中，注射后的3d 内HBVsi18-42 使小鼠血清中HBsAg 的水
平分别下降了98.98％、77.07％和60.73％，免疫组织化学检测显示，在注射后的第3 天小鼠肝组织内HBcAg 阳
性细胞数减少了79.1％。初步结果显示HBVsi18-42 无论是在细胞或是在小鼠模型中都能下调HBV 的复制和基因
的表达。本研究为我们下一步实现由pRNA 介导的靶向RNAi 及基因治疗提供了理论和技术支持。
关键词：RNA 干扰；pRNA；乙型肝炎病毒
