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Journal of Virology, September 2009, p. 8396-8408, Vol. 83, No. 17
0022-538X/09/$08.00+0 doi:10.1128/JVI.00700-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Detection of Clonally Expanded Hepatocytes in Chimpanzees with Chronic Hepatitis B Virus Infection 
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William S. Mason,1*,
Huey-Chi Low,2,3
Chunxiao Xu,1
Carol E. Aldrich,1
Catherine A. Scougall,2,3
Arend Grosse,2,3
Andrew Clouston,4
Deborah Chavez,5
Samuel Litwin,1
Suraj Peri,1
Allison R. Jilbert,2,3, and
Robert E. Lanford5,
Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, Pennsylvania 19111,1 School of Molecular and Biomedical Science, University of Adelaide, Adelaide SA 5005, Australia,2 Infectious Diseases Laboratories, Institute of Medical and Veterinary Science, Adelaide SA 5000, Australia,3 University of Queensland Clinical Research Centre, Brisbane QLD 4029, Australia,4 Department of Virology and Immunology, Southwest Foundation for Biomedical Research and Southwest National Primate Research Center, 7620 NW Loop 410, San Antonio, Texas 782275
Received 3 April 2009/ Accepted 8 June 2009
During a hepadnavirus infection, viral DNA integrates at a low rate into random sites in the host DNA, producing unique virus-cell junctions detectable by inverse nested PCR (invPCR). These junctions serve as genetic markers of individual hepatocytes, providing a means to detect their subsequent proliferation into clones of two or more hepatocytes. A previous study suggested that the livers of 2.4-year-old woodchucks (Marmota monax) chronically infected with woodchuck hepatitis virus contained at least 100,000 clones of >1,000 hepatocytes (W. S. Mason, A. R. Jilbert, and J. Summers, Proc. Natl. Acad. Sci. USA 102:1139-1144, 2005). However, possible correlations between sites of viral-DNA integration and clonal expansion could not be explored because the woodchuck genome has not yet been sequenced. In order to further investigate this issue, we looked for similar clonal expansion of hepatocytes in the livers of chimpanzees chronically infected with hepatitis B virus (HBV). Liver samples for invPCR were collected from eight chimpanzees chronically infected with HBV for at least 20 years. Fifty clones ranging in size from 
35 to 10,000 hepatocytes were detected using invPCR in 32 liver biopsy fragments (1 mg) containing, in total, 3 x 107 liver cells. Based on searching the analogous human genome, integration sites were found on all chromosomes except Y, 30% in known or predicted genes. However, no obvious association between the extent of clonal expansion and the integration site was apparent. This suggests that the integration site per se is not responsible for the outgrowth of large clones of hepatocytes.
* Corresponding author. Mailing address: Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, PA 19111. Phone: (215) 728-2462. Fax. (215) 728-2412. E-mail: ws_mason{at}fccc.edu
Published ahead of print on 17 June 2009.
Supplemental material for this article may be found at http://jvi.asm.org/.
W.S.M., A.R.J., and R.E.L. contributed equally to this work.
Journal of Virology, September 2009, p. 8396-8408, Vol. 83, No. 17
0022-538X/09/$08.00+0 doi:10.1128/JVI.00700-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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