1887

Abstract

Considering growing evidence indicating that hepatitis C virus (HCV) replicates in lymphoid cells, establishment of a reliable and sensitive method for detection of HCV in these cells may provide means for monitoring the infection and the efficacy of sterilizing antiviral therapy. In this study, conditions for augmentation and detection of the HCV genome in peripheral blood mononuclear cells (PBMCs) from patients with chronic hepatitis C (CHC) or after a sustained virological response (SVR) to antiviral treatment were assessed. Following stimulation with combinations of mitogens and/or cytokines, PBMCs and, in certain cases, affinity-purified T and B cells were examined for HCV positive- and negative-strand RNA by using RT-PCR followed by nucleic acid hybridization, while the presence of viral NS3 protein was determined by flow cytometry. HCV RNA augmentation was assessed by quantification of Southern and dot-blot hybridization signals. The results showed that treatment of peripheral lymphoid cells with mitogens stimulating T- and B-cell proliferation and with cytokines supporting their growth significantly increased HCV RNA detection in patients with both CHC and SVR. This enhancement was up to 100-fold for the HCV genome and fivefold for the NS3 protein compared with untreated cells. In conclusion, HCV RNA can be readily detected in circulating lymphoid cells in progressing hepatitis C and following SVR after cell stimulation. As such, this method offers a new investigative tool to study HCV lymphotropism and to monitor virus presence during the course of HCV infection.

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2005-03-01
2019-11-20
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References

  1. Bartenschlager, R. & Lohmann, V. ( 2000; ). Replication of hepatitis C virus. J Gen Virol 81, 1631–1648.
    [Google Scholar]
  2. Brams, P., Royston, I. & Boerner, P. ( 1993; ). In vitro priming of human lymphocytes. I. IL2 and IL4 requirements. Hum Antibodies Hybridomas 4, 47–56.
    [Google Scholar]
  3. Braun, R. W. & Reiser, H. C. ( 1986; ). Replication of human cytomegalovirus in human peripheral blood T cells. J Virol 60, 29–36.
    [Google Scholar]
  4. Brechot, C. ( 1996; ). Hepatitis C virus: molecular biology and genetic variability. Dig Dis Sci 41, 6S–21S.[CrossRef]
    [Google Scholar]
  5. Castillo, I., Pardo, M., Bartolome, J. & 9 other authors ( 2004; ). Occult hepatitis C virus infection in patients in whom the etiology of persistently abnormal results of liver-function tests is unknown. J Infect Dis 189, 7–14.[CrossRef]
    [Google Scholar]
  6. Coffin, C. S. & Michalak, T. I. ( 1999; ). Persistence of infectious hepadnavirus in offspring of woodchuck mothers recovered from viral hepatitis. J Clin Invest 104, 203–212.[CrossRef]
    [Google Scholar]
  7. Cohen, J. ( 1999; ). The scientific challenge of hepatitis C virus. Science 285, 26–30.[CrossRef]
    [Google Scholar]
  8. El-Serag, H. B., Hampel, H., Yeh, C. & Rabeneck, L. ( 2002; ). Extrahepatic manifestation of hepatitis C among United States male veterans. Hepatology 36, 1439–1445.[CrossRef]
    [Google Scholar]
  9. Frenkel, N., Schirmer, E. C., Katsafanas, G. & June, C. H. ( 1990; ). T cell activation is required for efficient replication of human herpesvirus 6. J Virol 64, 4598–4602.
    [Google Scholar]
  10. Gowda, S. D., Stein, B. S., Mohagheghpour, N., Benike, C. J. & Engleman, E. G. ( 1989; ). Evidence that T cell activation is required for HIV-1 entry in CD4+ lymphocytes. J Immunol 142, 773–780.
    [Google Scholar]
  11. Grosjean, I., Caux, C., Bella, I., Berger, I., Wild, F., Banchereau, J. & Kaiserlian, D. ( 1997; ). Measles virus infects human dendritic cells and blocks their allostimulatory property for CD4+ T cells. J Exp Med 186, 801–812.[CrossRef]
    [Google Scholar]
  12. Houghton, M., Selby, M., Weiner, A. & Choo, Q. L. ( 1994; ). Hepatitis C virus: structure, protein products and processing of the polyprotein precursor. Curr Stud Hematol Blood Transfus 61, 1–11.
    [Google Scholar]
  13. Hyypia, T., Korkiamaki, P. & Vainiopaa, R. ( 1985; ). Replication of measles virus in human lymphocytes. J Exp Med 161, 1261–1271.[CrossRef]
    [Google Scholar]
  14. Joshi, A., Vahlenkamp, T. W., Garg, H., Tompkins, W. A. & Tompkins, M. B. ( 2004; ). Preferential replication of FIV in activated CD4+CD25+ T cells independent of cellular proliferation. Virology 321, 307–322.[CrossRef]
    [Google Scholar]
  15. Kato, N., Nakazawa, T., Mizutani, T. & Shimotohno, K. ( 1995; ). Susceptibility of human T lymphotropic virus type 1 infected cell line MT-2 to hepatitis C virus infection. Biochem Biophys Res Commun 206, 863–869.[CrossRef]
    [Google Scholar]
  16. Laskus, T., Radkowski, M., Wang, L. F., Vargas, H. & Rakela, J. ( 1998; ). The presence of active hepatitis C virus replication in lymphoid tissue in patients coinfected with human immunodeficiency virus type 1. J Infect Dis 178, 1189–1192.[CrossRef]
    [Google Scholar]
  17. Laskus, T., Radkowski, M., Piasek, A., Nowicki, M., Horban, A., Cianciara, J. & Rakela, J. ( 2000; ). Hepatitis C virus in lymphoid cells of patients coinfected with human immunodeficiency virus type 1: evidence of active replication in monocytes/macrophages and lymphocytes. J Infect Dis 181, 442–448.[CrossRef]
    [Google Scholar]
  18. Lew, Y.-Y. & Michalak, T. I. ( 2001; ). In vitro and in vivo infectivity and pathogenicity of the lymphoid cell derived woodchuck hepatitis virus. J Virol 74, 1770–1782.
    [Google Scholar]
  19. Lucas, C. J., Ubels-Postma, J. C., Rezee, A. & Galama, J. M. ( 1978; ). Activation of measles virus from silently infected human lymphocytes. J Exp Med 148, 940–952.[CrossRef]
    [Google Scholar]
  20. Marrack, P., Mitchell, T., Bender, J., Hildeman, D., Kedl, R., Trague, K. & Kappler, J. ( 1998; ). T cell survival. Immunol Rev 165, 279–285.[CrossRef]
    [Google Scholar]
  21. Melisko, M. E., Fox, R. & Venook, A. ( 2004; ). Reactivation of hepatitis C virus after chemotherapy for colon cancer. Clin Oncol 16, 204–205.[CrossRef]
    [Google Scholar]
  22. Michalak, T. I. ( 2000; ). Occult persistence and lymphotropism of hepadnaviral infection: insights from the woodchuck viral hepatitis model. Immunol Rev 174, 98–111.[CrossRef]
    [Google Scholar]
  23. Michalak, T. I., Pasquinelli, C., Guilhot, S. & Chisari, F. V. ( 1994; ). Hepatitis B virus persistence after recovery from acute viral hepatitis. J Clin Invest 93, 230–239.[CrossRef]
    [Google Scholar]
  24. Michalak, T. I., Pardoe, I. U., Coffin, C. S., Churchill, N. D., Freake, D. S., Smith, P. & Trelegan, C. L. ( 1999; ). Occult lifelong persistence of infectious hepadnavirus and residual liver inflammation in woodchucks convalescent from acute viral hepatitis. Hepatology 29, 928–938.[CrossRef]
    [Google Scholar]
  25. Michalak, T. I., Mulrooney, P. M. & Coffin, C. S. ( 2004; ). Low doses of hepadnavirus induce infection of the lymphatic system that does not engage the liver. J Virol 78, 1730–1738.[CrossRef]
    [Google Scholar]
  26. Morsica, G., Tambussi, G., Sitia, G., Novati, R., Lazzarin, A., Lopalco, L. & Mukenge, S. ( 1999; ). Replication of hepatitis C virus in B lymphocytes (CD19+). Blood 94, 1138–1149.
    [Google Scholar]
  27. Musto, P. ( 2002; ). Hepatitis C virus infection and B cell non-Hodgkin's lymphomas: more than a simple association. Clin Lymphoma 3, 150–160.[CrossRef]
    [Google Scholar]
  28. Oldstone, M. B. ( 1996; ). Virus–lymphoid cell interactions. Proc Natl Acad Sci U S A 93, 12756–12758.[CrossRef]
    [Google Scholar]
  29. Pham, T. N. Q., MacParland, S. A., Mulrooney, P. M., Cooksley, H., Naoumov, N. V. & Michalak, T. I. ( 2004; ). Hepatitis C virus persistence after spontaneous or treatment-induced resolution of hepatitis C. J Virol 78, 5867–5874.[CrossRef]
    [Google Scholar]
  30. Radkowski, M., Wang, L. F., Vargas, H. E., Rakela, J. & Laskus, T. ( 1998; ). Detection of hepatitis C virus replication in peripheral blood mononuclear cells after orthotopic liver transplantation. Transplantation 66, 664–666.[CrossRef]
    [Google Scholar]
  31. Radkowski, M., Kubicka, J., Kisiel, E., Cianciara, J., Nowicki, M., Rakela, J. & Laskus, T. ( 2000; ). Detection of active hepatitis C virus and hepatitis G virus/GB virus C replication in bone marrow in human subjects. Blood 95, 3986–3990.
    [Google Scholar]
  32. Radkowski, M., Bednarska, A., Horban, A., Stanczak, J., Wilkinson, J., Adair, P. M., Nowicki, M., Rakela, J. & Laskus, T. ( 2004; ). Infection of primary human macrophages with hepatitis C virus in vitro: induction of tumor necrosis factor-α and interleukin 8. J Gen Virol 85, 47–59.[CrossRef]
    [Google Scholar]
  33. Shimizu, Y. K., Iwamoto, A., Hijikata, M., Purcell, R. H. & Yoshikura, H. ( 1992; ). Evidence for in vitro replication of hepatitis C virus genome in a human T cell line. Proc Natl Acad Sci U S A 89, 5477–5481.[CrossRef]
    [Google Scholar]
  34. Shimizu, Y. K., Igarashi, H., Kiyohara, T., Shapiro, M., Wong, D. C., Purcell, R. H. & Yoshikura, H. ( 1998; ). Infection of a chimpanzee with hepatitis C virus grown in cell culture. J Gen Virol 79, 1383–1386.
    [Google Scholar]
  35. Sung, V. M. H., Shimodaira, S., Doughty, A. L., Picchio, G. R., Can, H., Yen, T. S., Lindsay, K. L., Levine, A. M. & Lai, M. M. ( 2003; ). Establishment of B cell lymphoma cell lines persistently infected with hepatitis C virus in vivo and in vitro: the apoptotic effects of virus infection. J Virol 77, 2134–2146.[CrossRef]
    [Google Scholar]
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