乙型肝炎病毒核DNA的分布。

StephenW

J Virol. 2017 Oct 18. pii: JVI.01391-17. doi: 10.1128/JVI.01391-17. [Epub ahead of print]
Distribution of hepatitis B virus nuclear DNA.
Li M1,2, Sohn JA1, Seeger C3.
Author information

1
    Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.
2
    Department of Infectious Diseases, Institute of Hepatology, Central South University, Second Xiangya Hospital, Changsha, Hunan 410011, PR China.
3
    Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA [email protected].

Abstract

Chronic hepatitis B affects over 300 million people who are at risk of developing liver cancer. The basis for the persistence of hepatitis B virus (HBV) in hepatocytes, even in the presence of available antiviral therapies, lies in the accumulation of covalently closed circular (ccc) DNA in nuclei of infected cells. While methods for cccDNA quantification from liver biopsies and cell lines expressing the virus are known, information about cccDNA formation, stability and turnover are lacking. In particular, little is known about the fate of cccDNA during cell division. To fill gaps in knowledge concerning cccDNA biology, we have developed a fluorescence imaging in situ hybridization (FISH)-based assay for the detection of duck hepatitis B virus (DHBV) cccDNA and HBV nuclear DNA in established cell lines. Using FISH, we determined the distribution of cccDNA under conditions mimicking chronic infections with and without antiviral therapy, which prevents de novo viral replication. Our results showed that the copy numbers of viral nuclear DNA can vary by as much as 1.8 orders of magnitude among individual cells, and that antiviral therapy leads to a reduction in nuclear DNA in a manner consistent with symmetrical distribution of viral DNA to daughter cells.Importance A mechanistic understanding of the stability of HBV cccDNA in the presence of antiviral therapy and during cell division induced by immune mediated lysis of infected hepatocytes, will be critical for the future design of curative antiviral therapies against chronic hepatitis B. To date, knowledge about cccDNA stability was largely derived from quantitative analyses of cccDNA levels present in liver samples, and little was known about the fate of cccDNA in individual cells. The development of a FISH-based assay for cccDNA tracking provided the first insights into the fate of DHBV cccDNA and nuclear HBV DNA during conditions mimicking antiviral therapy.

Copyright © 2017 American Society for Microbiology.

PMID:
    29046450
DOI:
    10.1128/JVI.01391-17

StephenW

J Virol。 2017年10月18日。pii：JVI.01391-17。 doi：10.1128 / JVI.01391-17。 [提前印刷]
乙型肝炎病毒核DNA的分布。
李M1,2，索恩JA1，Seeger C3。
作者信息

1
    美国宾夕法尼亚州费城福克斯大通癌症中心癌症研究所19111，美国。
2
    中南大学肝脏病研究所传染病科，湖南长沙第二湘雅医院410011。
3
    美国宾夕法尼亚州费城福克斯大通癌症中心癌症研究所，19111，美国[email protected]。

抽象

慢性乙型肝炎影响超过3亿人谁有发展肝癌的风险。乙肝病毒（HBV）在肝细胞中持续存在的基础，即使存在可用的抗病毒疗法，也就是共价闭合环状（ccc）DNA在感染细胞核中的积累。虽然肝活检和表达病毒的细胞系的cccDNA定量方法是已知的，但是缺乏关于cccDNA形成，稳定性和周转率的信息。特别是，细胞分裂过程中cccDNA的命运知之甚少。为了填补cccDNA生物学知识的空白，我们开发了基于荧光成像原位杂交（FISH）的检测方法，用于检测已建立的细胞系中的鸭乙型肝炎病毒（DHBV）cccDNA和HBV核DNA。使用FISH，我们确定cccDNA在有和没有抗病毒治疗的模拟慢性感染的条件下的分布，这阻止了从头病毒复制。我们的研究结果表明，病毒核DNA的拷贝数可以在单个细胞中变化多达1.8个数量级，抗病毒治疗以与病毒DNA对子细胞的对称分布一致的方式导致核DNA的减少。重要性在抗病毒治疗和感染肝细胞的免疫介导的裂解诱导的细胞分裂过程中，对于HBV cccDNA的稳定性的机理理解对于慢性乙型肝炎的治疗性抗病毒治疗的未来设计至关重要。至今，关于cccDNA稳定性主要来自肝脏样品中存在的cccDNA水平的定量分析，并且对于各个细胞中cccDNA的命运知之甚少。基于FISH的cccDNA跟踪测定的开发为模拟抗病毒治疗的病症提供了第一次了解DHBV cccDNA和核HBV DNA的命运。

版权所有©2017美国微生物学会。

结论：
    29046450
DOI：
    10.1128 / JVI.01391-17