Cystine transport and glutathione metabolism in human erythrocytes

Cystine was transported into human erythrocytes in the presence of tertiary-butyl hydroperoxide (t-BH) or 1-chloro-2, 4-dinitrobenzene (CDNB). The transport rate of cystine was dependent on the extracellular concentration of t-BH or CDNB, and on the incubation time. By Dowex-1 column chromatography, the transported cystine was incorporated into fractions of glutathione disulfide (GSSG) and glutathione-S (GSH-S) conjugate. Cystine was also transported into reconstituted erythrocyte ghost with GSSG. The transport of cystine was Na+ dependent and decreased in the presence of N-ethylmaleimide, and it was competitively inhibited by DL-homocystine and L-alanine. The inhibition rates by DL-homocystine and L-alanine were 75% and 68%, with similar Ki values of 0.7 mM and 0.6 mM, respectively. The Km value for cystine transport was 0.15 mM. The activity of GSH-cystine transhydrogenase was detected in the hemolysate and this enzyme is thought to catalyze the action of incorporation of cystine into GSH. This enzyme was partially purified from normal human erythrocytes. In the presence of CDNB, similar rates of cystine transport were observed among the diabetic patients (n = 11), hypoxemic patients (n = 10) and the control subjects (n = 20). It is suggested that cystine transport is induced for glutathione synthesis when human erythrocytes are exposed to oxidative stresses.