ABSTRACT
We compared the utility of restriction endonuclease cleavage to type herpes simplex virus DNA polymerase gene amplicons from two well established PCR primer sets. Amplicons typed using Ava II had a 96% correlation to type determined by monoclonal antibody staining, while amplicons typed using Drd I had a 72% correlation to type determined by monoclonal antibody staining.
Subject(s)
DNA-Directed DNA Polymerase/genetics , Herpesvirus 1, Human/classification , Herpesvirus 2, Human/classification , DNA Primers , DNA, Viral/analysis , Herpesvirus 1, Human/genetics , Herpesvirus 1, Human/isolation & purification , Herpesvirus 2, Human/genetics , Herpesvirus 2, Human/isolation & purification , Humans , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Sequence Analysis, DNAABSTRACT
BACKGROUND: Thymosin-alpha 1 is a biological response modifier that has been used clinically, alone and in combination with interferon-alpha for the treatment of chronic hepatitis B viral infection. Both immunomodulatory and immediate intracellular mechanisms have been postulated to explain the effect of these two agents on HBV-infected hepatocytes. METHODS: In this study, hepatitis B transfected HepG2 hepatoblastoma cells (HepG2-Nu2), derived from 2.2.15 cells, were used as an in vitro model to determine the efficacy of thymosin-alpha 1 and interferon-alpha, individually and combined, as proliferation inhibitors of HBV-infected cells. For comparison, parental HepG2 cells and an SV40-transfected HepG2 cell line (HepG2P9T2) were also evaluated. RESULTS: In a clonogenic soft agar assay, thymosin-alpha 1 inhibited the anchorage-independent growth of the HepG2-Nu2 cells by 40% compared with untreated controls, but did not inhibit parental HepG2 or HepG2P9T2 clonal growth. The response was dose dependent over concentrations spanning three log units. In comparison, 10000 units/ml of interferon-alpha inhibited parental HepG2, HepG2-N4Z and HepG2P9T2 by 33%, 41% and 87%, respectively. The combination of thymosin-alpha 1 and interferon-alpha consistently inhibited HepG2-Nu2 clonal growth more effectively than either treatment alone, reaching maximum inhibition levels of 51%. CONCLUSIONS: Thymosin-alpha 1 specifically inhibits the tumorigenic growth of HBV-transfected HepG2 cells in contrast to the general inhibition displayed by interferon-alpha. This panel of cell lines may be an important resource for dissecting the mechanism by which thymosin, alone or in combination with other drugs, influences HBV-infected hepatocytes and/or HBV-associated carcinoma.
