Alterations in Normal and G-6-PD Deficient Human Erythrocytes of Various Ages after Exposure to Metabolites of Hemolytic Drugs

Abstract

Since in vivo studies indicate that older erythro cytes hemolyze preferentially in drug-induced hemolysis, differential centrifugation was used to separate young and old normal and G-6-PD deficient erythrocytes. Older cells were identified by their lower level of G-6-PD activity; in corporation of ⁵⁹Fe into erythrocytes confirmed this division of cells by age. Mechanical fragility (MF) of old cells was greater than that of young cells. Effects of 8-amino-5,6-quino linediol (AQD) (a possible primaquine metabolite) and of 2-hydroxyphenetidine (HP) (a phenacetin metabolite) on hemolysis, MF and reduced glutathione (GSH) and methemoglobin content were examined in vitro. AQD (2.5 × 10^–4 m) did not produce hemolysis but selectively increased MF of old G-6-PD deficient cells. HP (5 × 10^–4 m) produced considerable hemolysis of both normal and G-6-PD deficient cells with a selective effect in old G-6-PD deficient cells. HP also selectively increased the MF of old G-6-PD deficient cells. Incubation with AQDor HP depleted GSH content more in old than in young G-6-PD deficient cells. This depletion of GSH was followed by increased MF of old G-6-PD deficient cells. Hemolysis of normal cells occurred without GSH depletion but was accompanied by GSH depletion in G-6-PD deficient cells. Smaller differences in methemoglobin formation occurred but were not well correlated with hemolysis or increased MF. Although not applicable as a routine diagnostic test in the clinical laboratory, increased MF in vitro of old G-6-PD deficient erythrocytes is suggested as a useful research tool in assessing the potential of drugs or drug metabolites to cause hemolysis in vivo, particularly in combination with parallel studies of glutathione depletion.