Mitogen-induced upregulation of hepatitis C virus expression in human lymphoid cells.
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 ex vivo 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 ex vivo 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.
Pham
Tram N Q
TNQ
Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6.
MacParland
Sonya A
SA
Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6.
Coffin
Carla S
CS
Liver Unit, Division of Gastroenterology, University of Calgary, Calgary, Alberta, Canada.|Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6.
Lee
Samuel S
SS
Liver Unit, Division of Gastroenterology, University of Calgary, Calgary, Alberta, Canada.
Bursey
Ford R
FR
Gastroenterology Unit, General Hospital, Faculty of Medicine, Memorial University, St John's, Newfoundland, Canada.
Michalak
Tomasz I
TI
Discipline of Laboratory Medicine, Faculty of Medicine, Memorial University, St John's, Newfoundland, Canada.|Molecular Virology and Hepatology Research, Division of Basic Medical Science, Faculty of Medicine, Health Sciences Centre, Memorial University, St John's, Newfoundland, Canada A1B 3V6.
D000328:Adult; D005260:Female; D015870:Gene Expression; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D006526:Hepatitis C / Q000188:drug therapy / Q000821:virology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology*; D008297:Male; D008934:Mitogens / Q000494:pharmacology*; D012367:RNA, Viral / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015854:Up-Regulation; D014779:Virus Replication
D008934:Mitogens; D012367:RNA, Viral