Background and aim: It has been a conflicting issue whether TT virus (TTV), a newly isolated DNA virus from a patient with liver injury of unknown cause, is a causative agent of acute and/or chronic hepatitis. TT Virus DNA titers were shown to be 10-100-fold greater in liver tissue than in serum, whereas the majority of TTV-positive cases had no biochemical or histological evidence of significant liver damage. We therefore attempted in situ hybridization to investigate whether TTV is hepatotropic.
Methods: Because of the marked divergence in TTV genome types, a template for TTV-DNA (coding region for N22 clone) was amplified and labeled with digoxigenin-dUTP by using hemi-nested PCR from the serum, then DNA probes were applied to the liver sections of the same case. After hybridization, the probes were visualized immunohistochemically. Besides TTV-DNA-negative cases, competitive inhibition experiments with unlabeled probes were performed to confirm the specificity.
Results: There were no positive signals in the negative controls, and the intensity of positive signals was markedly diminished in the competitive inhibition experiments. No cross-hybridization with different genotype probes also confirms the specificity. Under the optimal conditions, the positive signals were located in the cytoplasm of the hepatocytes in eight of nine TTV-DNA-positive cases. The signals were not seen in non-parenchymal cells of the liver.
Conclusion: TT Virus is proved to be hepatotropic by in situ hybridization.