PDF(Pubmed)
Abstract：
Individuals chronically infected with hepatitis B virus (HBV) and hepatitis Delta virus (HDV) present an increased risk of developing cirrhosis and hepatocellular carcinoma in comparison to HBV mono-infected individuals. Although HDV only replicates in individuals coinfected or superinfected with HBV, there is currently no in vitro model that can stably express both viruses simultaneously, mimicking the chronic infections seen in HBV/HDV patients. Here, we present the HepG2BD cell line as a novel in vitro culture system for long-term replication of HBV and HDV. HepG2BD cells derive from HepG2.2.15 cells in which a 2 kb HDV cDNA sequence was inserted into the adeno-associated virus safe harbor integration site 1 (AAVS1) using CRISPR-Cas9. A Tet-Off promoter was placed 5\' of the genomic HDV sequence for reliable initiation/repression of viral replication and secretion. HBV and HDV replication were then thoroughly characterized. Of note, non-dividing cells adopt a hepatocyte-like morphology associated with an increased production of both HDV and HBV virions. Finally, HDV seems to negatively interfere with HBV in this model system. Altogether, HepG2BD cells will be instrumental to evaluate, in vitro, the fundamental HBV-HDV interplay during simultaneous chronic replication as well as for antivirals screening targeting both viruses.
摘要：
与HBV单感染的个体相比，慢性感染乙型肝炎病毒（HBV）和丁型肝炎病毒（HDV）的个体出现肝硬化和肝细胞癌的风险增加。虽然HDV仅在共感染或重叠感染HBV的个体中复制，目前还没有能够同时稳定表达两种病毒的体外模型，模仿在HBV/HDV患者中看到的慢性感染。这里,我们提出了HepG2BD细胞系作为一种新型的体外培养系统，用于HBV和HDV的长期复制。HepG2BD细胞来源于HepG2.2.15细胞,其中使用CRISPR-Cas9将2kb的HDVcDNA序列插入腺相关病毒安全港整合位点1(AAVS1)。将Tet-Off启动子放置在基因组HDV序列的5'5'上,以可靠地启动/抑制病毒复制和分泌。然后彻底表征HBV和HDV复制。值得注意的是，非分裂细胞采用与HDV和HBV病毒体产量增加相关的肝细胞样形态。最后,在这个模型系统中，HDV似乎对HBV产生了负面影响。总之,HepG2BD细胞将有助于评估，在体外，在同时慢性复制以及针对两种病毒的抗病毒药物筛查期间，HBV-HDV的基本相互作用。
