Detection of hepatitis C viral sequences in formalin-fixed, paraffin-embedded liver tissue: effect of interferon alpha therapy.
To determine the effect of interferon-alpha (IFN) therapy on hepatitis C virus (HCV) in liver, reverse transcription "nested" polymerase chain reaction (RT-PCR) was applied to detect HCV RNA in formalin-fixed, paraffin-embedded liver biopsy specimens obtained before and at the end of IFN therapy in 42 patients with chronic HCV infection. Results were correlated with the clinical and biochemical outcome in 36 cases. Fifteen patients were nonresponders to IFN; 13 patients had a complete response to IFN but relapsed shortly after IFN was stopped (responders who relapsed); and 8 patients showed a complete and sustained response to IFN therapy (sustained responders). Total RNA was extracted using proteinase K digestion and phenol/chloroform/isoamyl alcohol extraction, and HCV RNA was detected by standard RT-PCR using primers from the highly conserved 5' untranslated region. HCV RNA was detected in 41 of the 42 pretreatment specimens. Of the 36 patients with paired posttreatment samples, HCV RNA was detected in all 15 patients who did not respond to IFN and 9 of 13 who responded to IFN but relapsed shortly after IFN was stopped. In contrast, only one of the eight patients who had a sustained response to IFN therapy had HCV RNA detected by RT-PCR (P < 0.04). These data confirm 1) the feasibility of detecting HCV RNA in formalin-fixed, paraffin-embedded tissue from patients with chronic HCV infection, 2) show that sustained response to IFN is associated with loss of liver HCV RNA at the end of IFN therapy, and 3) offer an explanation for recurrence in patients who relapse.
Diamond
D A
DA
Department of Medicine, University of Florida, Gainesville.
Davis
G L
GL
Qian
K P
KP
Lau
J Y
JY
D001483:Base Sequence; D005557:Formaldehyde; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000382:microbiology* / Q000628:therapy; D006521:Hepatitis, Chronic / Q000382:microbiology* / Q000628:therapy; D006801:Humans; D007155:Immunologic Factors / Q000627:therapeutic use*; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000627:therapeutic use*; D008099:Liver / Q000382:microbiology*; D008969:Molecular Sequence Data; D010232:Paraffin; D012367:RNA, Viral / Q000032:analysis*; D011994:Recombinant Proteins; D012680:Sensitivity and Specificity; D016610:Tissue Embedding; D016707:Tissue Fixation
D007155:Immunologic Factors; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D012367:RNA, Viral; D011994:Recombinant Proteins; D005557:Formaldehyde; D010232:Paraffin