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Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes

Hepatitis B virus (HBV) infection and its sequelae remain a major public health burden, but both HBV basic research and the development of antiviral therapeutics have been hindered by the lack of an efficient in vitro infection system. Recently, sodium taurocholate cotransporting polypeptide (NTCP)...

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Autores principales: Yan, Ran, Zhang, Yongmei, Cai, Dawei, Liu, Yuanjie, Cuconati, Andrea, Guo, Haitao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4466484/
https://www.ncbi.nlm.nih.gov/pubmed/26070202
http://dx.doi.org/10.1371/journal.pone.0129889
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author Yan, Ran
Zhang, Yongmei
Cai, Dawei
Liu, Yuanjie
Cuconati, Andrea
Guo, Haitao
author_facet Yan, Ran
Zhang, Yongmei
Cai, Dawei
Liu, Yuanjie
Cuconati, Andrea
Guo, Haitao
author_sort Yan, Ran
collection PubMed
description Hepatitis B virus (HBV) infection and its sequelae remain a major public health burden, but both HBV basic research and the development of antiviral therapeutics have been hindered by the lack of an efficient in vitro infection system. Recently, sodium taurocholate cotransporting polypeptide (NTCP) has been identified as the HBV receptor. We herein report that we established a NTCP-complemented HepG2 cell line (HepG2-NTCP12) that supports HBV infection, albeit at a low infectivity level following the reported infection procedures. In our attempts to optimize the infection conditions, we found that the centrifugation of HepG2-NTCP12 cells during HBV inoculation (termed “spinoculation”) significantly enhanced the virus infectivity. Moreover, the infection level gradually increased with accelerated speed of spinoculation up to 1,000g tested. However, the enhancement of HBV infection was not significantly dependent upon the duration of centrifugation. Furthermore, covalently closed circular (ccc) DNA was detected in infected cells under optimized infection condition by conventional Southern blot, suggesting a successful establishment of HBV infection after spinoculation. Finally, the parental HepG2 cells remained uninfected under HBV spinoculation, and HBV entry inhibitors targeting NTCP blocked HBV infection when cells were spinoculated, suggesting the authentic virus entry mechanism is unaltered under centrifugal inoculation. Our data suggest that spinoculation could serve as a standard protocol for enhancing the efficiency of HBV infection in vitro.
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spelling pubmed-44664842015-06-22 Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes Yan, Ran Zhang, Yongmei Cai, Dawei Liu, Yuanjie Cuconati, Andrea Guo, Haitao PLoS One Research Article Hepatitis B virus (HBV) infection and its sequelae remain a major public health burden, but both HBV basic research and the development of antiviral therapeutics have been hindered by the lack of an efficient in vitro infection system. Recently, sodium taurocholate cotransporting polypeptide (NTCP) has been identified as the HBV receptor. We herein report that we established a NTCP-complemented HepG2 cell line (HepG2-NTCP12) that supports HBV infection, albeit at a low infectivity level following the reported infection procedures. In our attempts to optimize the infection conditions, we found that the centrifugation of HepG2-NTCP12 cells during HBV inoculation (termed “spinoculation”) significantly enhanced the virus infectivity. Moreover, the infection level gradually increased with accelerated speed of spinoculation up to 1,000g tested. However, the enhancement of HBV infection was not significantly dependent upon the duration of centrifugation. Furthermore, covalently closed circular (ccc) DNA was detected in infected cells under optimized infection condition by conventional Southern blot, suggesting a successful establishment of HBV infection after spinoculation. Finally, the parental HepG2 cells remained uninfected under HBV spinoculation, and HBV entry inhibitors targeting NTCP blocked HBV infection when cells were spinoculated, suggesting the authentic virus entry mechanism is unaltered under centrifugal inoculation. Our data suggest that spinoculation could serve as a standard protocol for enhancing the efficiency of HBV infection in vitro. Public Library of Science 2015-06-12 /pmc/articles/PMC4466484/ /pubmed/26070202 http://dx.doi.org/10.1371/journal.pone.0129889 Text en © 2015 Yan et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Yan, Ran
Zhang, Yongmei
Cai, Dawei
Liu, Yuanjie
Cuconati, Andrea
Guo, Haitao
Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title_full Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title_fullStr Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title_full_unstemmed Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title_short Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes
title_sort spinoculation enhances hbv infection in ntcp-reconstituted hepatocytes
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4466484/
https://www.ncbi.nlm.nih.gov/pubmed/26070202
http://dx.doi.org/10.1371/journal.pone.0129889
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