In situ analysis of intrahepatic virological events in chronic hepatitis B virus infection
Autor: | Liang Chen, Wei Lu, Yanling Feng, Ye Zheng, Zhanqing Zhang, Xiaonan Zhang, Weixia Wang, Zhenghong Yuan, Lu Bai |
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Jazyk: | angličtina |
Rok vydání: | 2016 |
Předmět: |
0301 basic medicine
Hepatitis B virus Hybridization probe General Medicine cccDNA In situ hybridization Biology Hepatitis B medicine.disease_cause medicine.disease Virology Molecular biology Virus 03 medical and health sciences 030104 developmental biology Hepatitis B Chronic Viral replication medicine Humans Southern blot Research Article |
Popis: | Persistent hepatitis B virus (HBV) infection is established by the formation of an intranuclear pool of covalently closed circular DNA (cccDNA) in the liver. Very little is known about the intrahepatic distribution of HBV cccDNA in infected patients, particularly at the single-cell level. Here, we established a highly sensitive and specific ISH assay for the detection of HBV RNA, DNA, and cccDNA. The specificity of our cccDNA probe set was confirmed by its strict intranuclear signal and by a series of Southern blot analyses. Use of our in situ assay in conjunction with IHC or immunofluorescence uncovered a surprisingly mosaic distribution of viral antigens and nucleic acids. Most strikingly, a mutually exclusive pattern was found between HBV surface antigen-positive (HBsA-positive) and HBV DNA- and cccDNA-positive cells. A longitudinal observation of patients over a 1-year period of adeforvir therapy confirmed the persistence of a nuclear reservoir of viral DNA, although cytoplasmic DNA was effectively depleted in these individuals. In conclusion, our method for detecting viral nucleic acids, including cccDNA, with single-cell resolution provides a means for monitoring intrahepatic virological events in chronic HBV infection. More important, our observations unravel the complexity of the HBV life cycle in vivo. |
Databáze: | OpenAIRE |
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