Cell Culture of Sporadic Hepatitis E Virus in China

Huang, Rutong; Li, Derong; Wei, Shao-jing; Li, Qinghong; Yuan, Xitong; Li, Geng; Li, Xiaoyü; Liu, Minxia

doi:10.1128/cdli.6.5.729-733.1999

Cited by 63 publications

(33 citation statements)

References 16 publications

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“…In 2007, efficient HEV cell-culture propagation systems were established by using PLC/PRF/5, a hepatocarcinoma cell line, and A549, an adenocarcinomic human alveolar basal epithelial cell line (23). The replication efficiencies of these new systems are several orders of magnitude higher than those of previous culturing systems (23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33). The increased availability of HEV reagents provided by the new culturing systems has permitted many groups to construct HEV infectious clones, thereby facilitating multiple recent reverse genetic studies (34)(35)(36)(37)(38).…”

mentioning

confidence: 99%

Establishment of hepatitis E virus infection‐permissive and ‐non‐permissive human hepatoma PLC/PRF/5 subclones

Shiota

Yoshizaki

et al. 2015

Microbiology and Immunology

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PLC/PRF/5 cells show limited permissiveness, meaning that almost all subclones are permissive; however, some subclones do not exhibit permissiveness for hepatitis E virus (HEV) infection. In this study, the single-cell cloning of PLC/PRF/5 was performed and heterogeneous subclones characterized. Notably, the efficiency of intracellular virus replication did not correlate with the permissiveness for HEV infection. However, as well as binding permissive subclones, virus-like particles bound non-permissive subclones on various levels, suggesting that these subclones have some deficiencies in the attachment and entry steps of infection. Our data would be useful for investigating the HEV life cycle.

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mentioning

confidence: 99%

Establishment of hepatitis E virus infection‐permissive and ‐non‐permissive human hepatoma PLC/PRF/5 subclones

Shiota

Yoshizaki

et al. 2015

Microbiology and Immunology

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“…A549 (human lung carcinoma, ATCC, VA, USA) cell line was used in this study and maintained as described previously (Huang et al, 1999). A549 cells were trypsinized and transferred to 500 mL antibiotic-free growth medium in 24-well plates at 0.5-2.0 Â 10 5 /well.…”

Section: Sirna and Plasmid Transfectionmentioning

confidence: 99%

“…HEV has been successfully cultured in A549 cells as previously described (Huang et al, 1999). For HEV infection challenge, A549 cells were transfected with each of siRNAs (si-ORF2-1, si-ORF2-2, si-ORF2-3, si-ORF2-4, and scrambled siRNA) as described previously 24 h before the virus challenge.…”

Section: Live Virus Sirna Antiviral Assaysmentioning

confidence: 99%

“…For HEV infection challenge, A549 cells were transfected with each of siRNAs (si-ORF2-1, si-ORF2-2, si-ORF2-3, si-ORF2-4, and scrambled siRNA) as described previously 24 h before the virus challenge. The viral challenge was performed as the previously described (Huang et al, 1999;Tanaka et al, 2007). The virus at a viral count of 1-2 Â 10 5 /mL as calculated by viral genomic titer determined by Real-Time quantitative PCR (Li et al, 2006;Kasorndorkbua et al, 2004) was inoculated into A549 cells, and CPEs were observed daily.…”

Section: Live Virus Sirna Antiviral Assaysmentioning

confidence: 99%

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RNA interference inhibits hepatitis E virus mRNA accumulation and protein synthesis in vitro

Huang

Zhou

et al. 2010

Veterinary Microbiology

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Hepatitis E virus (HEV) is a zoonotic pathogen to which several species, including human beings, pigs and rodents, are reported to be susceptible. To date, vaccines developed against HEV still need to be improved and a structural gene (ORF2), which encodes a capsid protein with high sequence conservation found across HEV genotypes, is a potential candidate. To exploit the possibility of using RNA interference (RNAi) as a strategy against HEV infection, four small interference RNA (siRNA) duplex targeting ORF2 gene were constructed. A challenge against HEV infection by RNAi was performed in A549 cells. Real-Time quantitative polymerase chain reaction (Real-Time qPCR) and Western blot assay demonstrated that four HEV specific siRNAs (si-ORF2-1, si-ORF2-2, si-ORF2-3 and si-ORF2-4) were capable of protecting cells against HEV infection with very high specificity and efficiency. The results suggest that RNAi is a potent anti-HEV infection prophylaxis strategy.

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“…HEV has been successfully cultured in A549 and PLC/PRF/5 line cells, as previously described (Huang et al, 1999;Lorenzo et al, 2008;Tanaka et al, 2007) and only A549 cells show a typical cytopathic effect (CPE) induced by the virus. HEV positive feces suspension was centrifuged at 12,000 × g at 4 • C for 10 min, and filtered through 0.22 m microfilters, followed by treatment with penicillin and streptomycin for 1 h. The virus was 10-fold serially diluted, and each dilution was added to A549 cells in triplicate.…”

Section: Virus Challenge In A549 Cellsmentioning

confidence: 99%

Effective inhibition of hepatitis E virus replication in A549 cells and piglets by RNA interference (RNAi) targeting RNA-dependent RNA polymerase

et al. 2009

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RNA interference (RNAi) is a natural mechanism for suppressing or silencing expression of aberrant or foreign genes. It is a powerful antiviral strategy that has been widely employed to protect hosts from viral infection. Hepatitis E (HE) is an acute fulminant hepatitis in adults that has particularly high mortality in pregnant women. At this point in time, there is no vaccine or antiviral treatment that is effective against the infectious agent, HEV. The nonstructural polyprotein region possesses an RNA-dependent RNA polymerase (RdRp) that is responsible for the replication of the viral RNA genome. RdRp is therefore regarded as one of the most attractive candidates for RNA interference (RNAi). In the present study, the high efficiency and specificity of siRNA were evaluated by Real-Time quantitative PCR and Western blot assays. Protective effects against HEV infection were achieved in A549 cells and in piglets. In piglets treated with a shRNA-RdRp-1 expression plasmid prior to HEV inoculation, HEV antigens were significantly reduced in the liver, spleen, and kidneys, and the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBIL) were clearly decreased. These results suggested that RNAi is a potentially effective antiviral strategy against HEV replication and infection.

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Cell Culture of Sporadic Hepatitis E Virus in China

Cited by 63 publications

References 16 publications

Establishment of hepatitis E virus infection‐permissive and ‐non‐permissive human hepatoma PLC/PRF/5 subclones

Establishment of hepatitis E virus infection‐permissive and ‐non‐permissive human hepatoma PLC/PRF/5 subclones

RNA interference inhibits hepatitis E virus mRNA accumulation and protein synthesis in vitro

Effective inhibition of hepatitis E virus replication in A549 cells and piglets by RNA interference (RNAi) targeting RNA-dependent RNA polymerase

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