Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Okayama, 700-8558, Japan.
Division of Biological Information Technology, Joint Research Center for Human Retrovirus Infection, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima, 890-8544, Japan.
Arch Virol. 2020 Feb;165(2):331-343. doi: 10.1007/s00705-019-04461-0. Epub 2019 Dec 12.
The most characteristic feature of the hepatitis C virus (HCV) genome in patients with chronic hepatitis C is its remarkable variability and diversity. To better understand this feature, we performed genetic analysis of HCV replicons recovered from two human hepatoma HuH-7-derived cell lines after 1, 3, 5, 7, and 9 years in culture: The cell lines 50-1 and sO harbored HCV 1B-1 and O strain-derived HCV replicons established in 2002 and 2003, respectively. The results revealed that genetic variations in both replicons accumulated in a time-dependent manner at a constant rate despite the maintenance of moderate diversity (less than 1.8% difference) between the clones and that the mutation rate in the 50-1 and sO replicons was 2.5 and 2.9 × 10 base substitutions/site/year, respectively. We found that the genetic distance of both replicons increased from 7.9% to 10.5% after 9 years in culture. In addition, we observed that the guanine + cytosine (GC) content of both replicon RNAs increased in a time-dependent manner, as observed in our previous studies. Finally, we demonstrated that the high sensitivity of both replicons to direct-acting antivirals was maintained even after 9 years in culture. Our results suggest that long-term cultured HCV replicon-harboring cells are a useful model for understanding the variability and diversity of the HCV genome and the drug sensitivity of HCV in patients with chronic hepatitis C.
丙型肝炎病毒（HCV）基因组在慢性丙型肝炎患者中的最显著特征是其显著的变异性和多样性。为了更好地理解这一特征，我们对从培养了 1、3、5、7 和 9 年的两个人肝癌 HuH-7 衍生细胞系中回收的 HCV 复制子进行了遗传分析：细胞系 50-1 和 sO 分别携带 2002 年和 2003 年建立的 HCV 1B-1 和 O 株衍生的 HCV 复制子。结果表明，尽管克隆之间保持适度的多样性（差异小于 1.8%），但两个复制子的遗传变异都以时间依赖性方式以恒定速率累积，并且 50-1 和 sO 复制子的突变率分别为 2.5 和 2.9×10 个碱基取代/位点/年。我们发现，两个复制子的遗传距离在培养 9 年后从 7.9%增加到 10.5%。此外，我们观察到，与我们之前的研究一样，两种复制子的鸟嘌呤+胞嘧啶（GC）含量随时间呈依赖性增加。最后，我们证明了两种复制子对直接作用抗病毒药物的高敏感性即使在培养 9 年后也能保持。我们的研究结果表明，长期培养的 HCV 复制子携带细胞是理解 HCV 基因组变异性和多样性以及慢性丙型肝炎患者 HCV 药物敏感性的有用模型。
