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    L-cysteine influx and efflux in human erythrocytes: the role of red blood cells in redox and metabolite homeostasis in the plasma
    (Pharmaceutical Soc Japan, 2006) Yildiz, D; Ates, BH; Uslu, C; Oztas, H
    The objective of this study was to investigate L-cysteine influx and efflux in human erythrocytes. L-cysteine is an amino acid required for glutathione synthesis in erythrocytes. In addition to being incorporated into glutathione, the soluble antioxidant L-cysteine plays a role in the maintenance of a proper intracellular or extracellular redox status. Recent investigations have pointed out that L-cysteine may contribute to redox homeostasis in the plasma and in the periplasm of some bacteria. Thus L-cysteine availability in the plasma may influence the oxidized/reduced state of several other metabolites normally found in the plasma. Our L-cysteine uptake studies demonstrated that erythrocytes can respond to an increase in the L-cysteine concentration in the extracellular media and influx L-cysteine in a concentration dependent-manner. The L-cysteine efflux is also time and concentration dependent. Erythrocytes pretreated with higher concentration of L-cysteine displayed higher efflux rates. Erythrocytes pretreated with L-cysteine 1 mM displayed efflux and increased the free-SH concentrations up to 0.184 +/- 0.010 mM in the incubation media in 1 hr. While this concentration reached 0.843 +/- 0.012 mM in 10 mM-L-cysteine pretreated erythrocytes. Our results also showed that the L-cysteine efflux is partly mediated by the Alanine-Serine-Cysteine (ASC) system. The presence of alanine or serine in the incubation media decreased the rate of efflux by about 16%. Our results also showed that the L-cysteine efflux process is not a simple diffusion but a carrier-mediated process. When compared with N-acetyl-L-cysteine (NAC), which is known to diffuse through the membranes, L-cysteine displayed a higher efflux rate under the same conditions. Pretreatment of erythrocytes with L-cysteine 4 mM increased the free-SH concentration to 0.48 +/- 0.005 mM whereas the same concentration of NAC brought the free-SH concentration to 0.36 +/- 0.01 mM in the incubation media. Our results suggest that erythrocytes may contribute to redox and metabolite homeostasis of the plasma.
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    L-Cysteine influx and efflux: A possible role for red blood cells in regulation of redox status of the plasma
    (Taylor & Francis Ltd, 2006) Yildiz, D; Uslu, C; Cakir, Y; Oztas, H
    The objective of this study was to investigate if erythrocytes play a role in the maintenance of redox homeostasis of the plasma. Thus, we studied l-cysteine efflux and influx in vitro in human erythrocytes. In the present study, we exposed the erythrocytes to different concentrations of l-cysteine and then measured the intracellular free -SH concentrations. Erythrocytes treated in the same manner were later utilized for the cysteine efflux studies. The effect of temperature on the influx and the efflux processes were also evaluated. Change in the free -SH content of the buffer was evaluated as a measure for the presence of an efflux process. The effects of free -SH depletion on l-cysteine transport is also investigated. We also determined the rate of l-cysteine efflux in the presence and absence of buthionine sulfoximine (BSO) in erythrocytes that are pretreated with 1-chloro-2,4-dinitro benzene, a glutathione (GSH) depletory. Our l-cysteine influx studies demonstrated that erythrocytes can respond to increases in l-cysteine concentration in the extracellular media and influx l-cysteine in a concentration-dependent manner. Free -SH concentrations in erythrocytes treated with 1 mM l-cysteine reached to 1.64 0.06 mM in 1 h whereas this concentration reached to 4.30 0.01 mM in 10 mM l-cysteine treated erythrocytes. The l-cysteine efflux is also determined to be time-and concentration-dependent. Erythrocytes that are pretreated with higher l-cysteine concentrations displayed a higher efflux process. Outside concentration of free -SH in 1 mM l-cysteine pretreated erythrocytes reached to 0.200 0.005 mM in 1 h whereas this concentration reached to 1.014 0.002 with 10 mM l-cysteine pretreated erythrocytes. Our results also indicate that the rate of inward and outward transport of l-cysteine is affected by the oxidative status of the erythrocytes. When GSH is depleted and GSH synthesis is blocked, the l-cysteine uptake and the efflux processes are significantly decreased. Depending on our results, it could be concluded that erythrocytes play a role in the regulation of the plasma redox status and intracellular level of GSH determines the rate of the l-cysteine efflux.