Glutathione peroxidase, thioredoxin, and membrane protein changes in erythrocytes predict ribavirin-induced anemia

Abstract

Low erythrocyte membrane protein sulfhydril concentrations are a risk factor for ribavirin-induced anemia. We further studied the role of oxidative stress and erythrocyte membrane alterations in ribavirin-induced anemia. The levels of thioredoxin, glutathione peroxidase, protein sulfhydrils, and protein-mixed disulfides, as well as the electrophoretic membrane protein pattern, were determined in freshly isolated erythrocytes from healthy control subjects, patients without severe anemia during previous ribavirin treatment (still hepatitis C virus [HCV]-positive), and patients who had had severe anemia with ribavirin (still HCV-positive or HCV-negative), 6 months after full recovery. Erythrocytes were also incubated with buffer, ribavirin, phenylhydrazine, or dehydroepiandrosterone, and concentrations of protein sulfhydrils, protein-mixed disulfides, thiobarbituric acid-reactive substances, and total and oxidized glutathione, as well as osmotic resistance, were determined. Patients with previous severe ribavirin-induced anemia had lower levels of protein sulfhydrils (30.9 nmol/mg protein versus 43.2 nmol/mg protein, P<.001) and thioredoxin (0.6 nmol/g hemoglobin versus 1.2 nmol/g hemoglobin, P<.001), higher levels of protein-mixed disulfides (1.5 nmol/g hemoglobin versus 0.5 nmol/g hemoglobin, P<.001) and glutathione peroxidase (618 mU/mg protein versus 393 mU/mg protein, P<.001), and a membrane protein pattern consistent with band 4 dimer disaggregation. These differences were independent of HCV seropositivity. There were negative correlations between levels of glutathione peroxidase and thioredoxin (r=-0.87) and between levels of protein sulfhydrils and protein-mixed disulfides (r=-0.93). In vitro studies showed that erythrocytes of patients who had had hemolysis during treatment of HCV are more susceptible to oxidative stress. Pronounced differences in markers of oxidative stress and membrane proteins exist between patients with and without a history of ribavirin-induced anemia. Our findings suggest that there are erythrocyte-related risk factors for ribavirin-induced severe anemia.