Adenosine metabolism in human skin fibroblasts

Abstract

When normal fibroblasts were incubated in media containing various initial concentrations of [8-14C]adenosine, ranging from 0.25 to 400 microM, under conditions where product formation was linear, greater than 90% of the intracellular label was found in adenine nucleotides, largely in the form of ATP, less than 1% of the intracellular label appeared in the nucleic acids, the remaining intracellular label was found in adenosine, inosine, and hypoxanthine, and the media contained two labeled products, inosine and hypoxanthine. Production of labeled inosine and hypoxanthine from adenosine was considerably lower in adenosine deaminase (ADA)-deficient cells than in normal cells and virtually eliminated in normal cells by the presence of 1 microM deoxycoformycin (a potent ADA inhibitor), suggesting that labeled inosine and hypoxanthine production requires ADA activity. Initial rates of deamination (inosine and hypoxanthine formation) and phosphorylation (adenine nucleotide formation) were estimated by examining the metabolic fate of [8-14C]adenosine in hypoxanthine phosphoribosyltransferase-deficient cells, which cannot recycle hypoxanthine. The estimate of the initial rate of phosphorylation exceeded that of deamination only at the lowest adenosine concentration examined (0.25 microM). The ratio of deamination to phosphorylation rose from approximately 1 at 0.41 microM to approximately 15 at 400 microM extracellular adenosine.