Expression of Integrated Hepatitis B Virus X Variants in Human Hepatocellular Carcinomas and Its Significance

Chen, Wei Ning; Oon, Chong Jin; Leong, Ai Lin; S, Koh; Teng, Siaw Wei

doi:10.1006/bbrc.2000.3562

Cited by 40 publications

(34 citation statements)

References 38 publications

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“…Overall, the HBV X gene appears to be well conserved during chronic hepatitis B, and conservation of the X protein seems to be essential for the virus' ability to actively replicate (2,19). On the other hand, mutations in the HBV X gene sequence have been documented in HCC (5,20). We did not identify any conserved changes in the WHV X sequence analyzed.…”

Section: Discussioncontrasting

confidence: 54%

Repeated Passage of Wild-Type Woodchuck Hepatitis Virus in Lymphoid Cells Does Not Generate Cell Type-Specific Variants or Alter Virus Infectivity

Mulrooney‐Cousins¹,

Michalak

2008

J Virol

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Woodchuck hepatitis virus (WHV), which is closely related to human hepatitis B virus, infects the liver but also invariably establishes persistent infection in the lymphatic system. Although the dose of invading virus appears to be the main factor in determining whether WHV infection is restricted to the lymphatic system or also engages the liver, the nature of WHV lymphotropism remains unclear and a role for a specific lymphotropic variant was not excluded. The availability of woodchuck lymphocyte and hepatocyte cultures susceptible to WHV infection allows investigation of this issue in vitro. We hypothesized that repeated passage of wild-type WHV in lymphoid cells should lead to enrichment of a lymphotropic virus variant, if in fact such a variant exists. For this purpose, wild-type WHV with a homogeneous sequence was used as the inoculum, while lymphoid cells from a single healthy woodchuck donor and a normal woodchuck WCM-260 hepatocyte line served as infection targets. The serial passage of the wild-type virus repeated up to 13 times for both cell types did not lead to the emergence of cell type-specific WHV variants, as revealed by sequence analysis of the virus envelope and the core and X gene sequences. Moreover, the virus passaged in both cell types remained infectious for naive woodchucks, produced infection profiles that depended upon virus dose but not on virus cellular origin, and retained its initial DNA sequence. These results imply that WHV lymphotropism is a natural propensity of the wild-type virus and is not a consequence of infection with a viral variant.Accumulated evidence indicates that hepadnaviruses are capable of replication not only in hepatocytes but also in cells of the immune system. In regard to symptomatic, chronic hepatitis B virus (HBV) infection, which is characterized by the continuous presence of HBV surface antigen (HBsAg) in serum and protracted liver necroinflammation, long-term persistence of virus has been shown in both the liver and the lymphatic system (6,21,26,35,42,43,45). However, liver and lymphoid cells have also been found to be the sites of HBV genome carriage in serum HBsAg-negative patients in whom acute hepatitis (AH) had apparently resolved completely when sensitive PCR-based assays were applied to detect the virus genome (3,4,30,37,40,46). This occult form of HBV infection might be a source of infectious virus available for transmission to healthy individuals through blood or organ donations (1,12,21,24,27), as well as a potential cause of diseases of seemingly unknown etiology engaging the liver and the lymphatic system. In this regard, retrospective studies of hepatocellular carcinoma (HCC) of previously undetermined etiology showed low levels of HBV genomes in 63.5% of liver tissue samples tested (38). In some of those cases, integration of viral DNA into the host's genome was detected. This implies that trace amounts of HBV, identifiable by molecular assays of a sensitivity greater than those currently used in clinical laboratories, may retain pathogeni...

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Section: Discussioncontrasting

confidence: 54%

Repeated Passage of Wild-Type Woodchuck Hepatitis Virus in Lymphoid Cells Does Not Generate Cell Type-Specific Variants or Alter Virus Infectivity

Mulrooney‐Cousins¹,

Michalak

2008

J Virol

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“…26,27 Although HBxAg has been implicated in the process of HCC, the viral DNA-integrated genome is commonly found partial or complete deletion of HBx gene. 28,29 Moreover, the development of HCC is preceded by chronic liver injury and inflammation. Studies on HBsAg transgenic mice revealed that a severe, prolonged hepatocellular injury, regenerative hyperplasia, and a secondary inflammation preceded HCC.…”

Section: Discussionmentioning

confidence: 99%

siRNA pool targeting different sites of human hepatitis B surface antigen efficiently inhibits HBV infection

Yong

Mahato

2008

Journal of Drug Targeting

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The main objective was to determine whether a pool of siRNAs targeting different regions of hepatitis B virus surface antigen (HBsAg) efficiently inhibits HBV infection. siRNAs targeting different regions of HBsAg were transfected into HBV-producing HepG2.2.15 cells and at 72h post transfection, the culture medium was collected for ELISA to determine HBsAg, while total RNA was isolated from the cells for real-time PCR. Three siRNA sequences which efficiently inhibited HBV infection were converted into small hairpin RNAs (shRNAs) and then cloned into a single plasmid psiSTRIKE™ driven by a single U6 promoter. These shRNA expressing plasmids were tested for HBsAg gene silencing in HepG2.2.15 cells. A pool of siRNAs targeting HBsAg efficiently inhibited HBV replication and antigen expression when transfected into HepG2.2.15 cells, compared to the use of single siRNA. Similarly, the plasmid encoding three different shRNAs driven by a single U6 promoter was more effective in silencing HBsAg at DNA, mRNA and protein levels compared to the plasmid encoding single shRNA. No apoptotic change was observed in the cells when the plasmid was transfected at a dose of 0.5-2μg/1×10 6 cells after complex formation with Lipofectamine LTX™. Furthermore, transfection with siRNA or shRNA did not increase IFN-γ release, suggesting no induction of interferon response. In conclusion, a pool of chemically synthesized siRNAs as well as the shRNA expression plasmid encoding multiple shRNAs targeting different regions of HBsAg showed high gene silencing in HepG2.2.15 cells.

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“…This implies that HBV may have direct hepato-carcinogenic properties. Beside the chromosomal instability arising from HBV genome integration in the host chromosome (Beasley et al, 1981), the smallest HBX protein could also influence the onset of HCC by promoting the survival and growth of transformed hepatocytes (Andrisani and Barnabas, 1999;Birrer et al, 2003;Chen et al, 2000a).…”

Section: Introductionmentioning

confidence: 99%

A proteomics analysis of cellular proteins associated with HBV genotype-specific HBX: potential in identification of early diagnostic markers for HCC

Tan

Chen

2005

Journal of Clinical Virology

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Expression of Integrated Hepatitis B Virus X Variants in Human Hepatocellular Carcinomas and Its Significance

Cited by 40 publications

References 38 publications

Repeated Passage of Wild-Type Woodchuck Hepatitis Virus in Lymphoid Cells Does Not Generate Cell Type-Specific Variants or Alter Virus Infectivity

Repeated Passage of Wild-Type Woodchuck Hepatitis Virus in Lymphoid Cells Does Not Generate Cell Type-Specific Variants or Alter Virus Infectivity

siRNA pool targeting different sites of human hepatitis B surface antigen efficiently inhibits HBV infection

A proteomics analysis of cellular proteins associated with HBV genotype-specific HBX: potential in identification of early diagnostic markers for HCC

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