Detection of hepatitis C virus RNA sequences in hepatocellular carcinoma and its precursors by microdissection polymerase chain reaction.

Abstract

Chronic hepatitis C virus (HCV) infection is frequently associated with the development of hepatocellular carcinoma (HCC), but the mechanism of malignant transformation is unknown. To analyze the association of HCV with HCC, we developed a microdissection technique for the detection by polymerase chain reaction of positive (genomic)- and negative (replicative)-strand HCV RNA in histologically confirmed HCC and the surrounding cirrhotic and macroregenerative nodules. Five HCCs and one macroregenerative nodule and the surrounding cirrhotic liver tissues of all cases were selected for this study. The method entails extraction of RNA from selected areas of formalin-fixed, hematoxylin-stained histologic sections, followed by strand-specific reverse-transcription double polymerase chain reaction and Southern blotting. Positive- and negative-strand HCV RNA sequences were detected in five of six tumors and the surrounding cirrhotic livers. These results verify the method of polymerase chain reaction detection of HCV RNA from histologically defined, selected lesions. In addition, the findings suggest that HCV RNA persists and replicates in hepatocytes during malignant transformation.