Novel Recombinant Hepatitis B Virus Vectors Efficiently Deliver Protein and RNA Encoding Genes into Primary Hepatocytes
Novel Recombinant Hepatitis B Virus Vectors Efficiently Deliver Protein and RNA Encoding Genes into Primary Hepatocytes
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新型重组乙型肝炎病毒载体可有效地将蛋白质和RNA编码基因递送至原代肝细胞中
DOI：
10.1128/jvi.03328-12
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发表时间：
2013-06-01
影响因子：
5.4
通讯作者：
Xie, Youhua
中科院分区：
文献类型：
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作者：
Hong, Ran;Bai, Weiya;Xie, Youhua
B型肝炎病毒（HBV）具有极其有限的宿主嗜性和肝细胞嗜性。基于HBV的载体可以形成慢性B型肝炎和其他肝脏疾病的新疗法的基础，并且对于HBV感染的研究也是非常宝贵的。先前开发基于HBV的载体的尝试遇到重组病毒的低产量和/或在原代肝细胞中缺乏足够的感染性/货物基因表达，这阻碍了后续应用。在这项工作中，我们构建了一个新的载体的基础上，自然发生的，高度复制的HBV突变体与207 bp的缺失在preS 1/聚合酶间隔区。通过应用保持聚合酶开放阅读框（ORF）连续性的新插入策略，获得了携带蛋白或小干扰RNA（siRNA）基因的重组HBV（rHBV），其在培养的肝细胞中有效地复制和包装。我们证明了表达荧光报告基因（DsRed）的rHBV在原代树鼩肝细胞中具有高度感染性，并且表达HBV靶向siRNA的rHBV成功抑制了共感染野生型HBV的抗原表达。这种新型的HBV载体将为肝细胞靶向基因递送以及HBV感染的研究提供有力的工具。
Hepatitis B virus (HBV) has extremely restricted host and hepatocyte tropism. HBV-based vectors could form the basis of novel therapies for chronic hepatitis B and other liver diseases and would also be invaluable for the study of HBV infection. Previous attempts at developing HBV-based vectors encountered low yields of recombinant viruses and/or lack of sufficient infectivity/cargo gene expression in primary hepatocytes, which hampered follow-up applications. In this work, we constructed a novel vector based on a naturally occurring, highly replicative HBV mutant with a 207-bp deletion in the preS1/polymerase spacer region. By applying a novel insertion strategy that preserves the continuity of the polymerase open reading frame (ORF), recombinant HBV (rHBV) carrying protein or small interfering RNA (siRNA) genes were obtained that replicated and were packaged efficiently in cultured hepatocytes. We demonstrated that rHBV expressing a fluorescent reporter (DsRed) is highly infective in primary tree shrew hepatocytes, and rHBV expressing HBV-targeting siRNA successfully inhibited antigen expression from coinfected wild-type HBV. This novel HBV vector will be a powerful tool for hepatocyte-targeting gene delivery, as well as the study of HBV infection.