Clotrimazole enhances lysis of human erythrocytes induced by t-BHP.

Clotrimazole (CLT) is an antifungal and antimalarial agent also effective as a Gardos channel inhibitor. In addition, CLT possesses antitumor properties. Recent data provide evidence that CLT forms a complex with heme (hemin), which produces a more potent lytic effect than heme alone. This study addressed the effect of CLT on the lysis of normal human erythrocytes induced by tert-butyl hydroperoxide (t-BHP). For the first time, it was shown that 10 microM CLT significantly enhanced the lytic effect of t-BHP on erythrocytes in both Ca(2+)-containing and Ca(2+)-free media, suggesting that the effect is not related to Gardos channels. CLT did not affect the rate of free radical generation, the kinetics of GSH degradation, methemoglobin formation and TBARS generation; therefore, we concluded that CLT does not cause additional oxidative damage to erythrocytes treated with t-BHP. It is tempted to speculate that CLT enhances t-BHP-induced changes in erythrocyte volume and lysis largely by forming a complex with hemin released during hemoglobin oxidation in erythrocytes: the CLT-hemin complex destabilizes the cell membrane more potently than hemin alone. If so, the effect of CLT on cell membrane damage during free-radical oxidation may be used to increase the efficacy of antitumor therapy.

Factors raising intracellular calcium increase red blood cell heterogeneity in density and critical osmolality.

BACKGROUND: In normal human blood, RBC volume (V), surface area (A) and hemoglobin content (H) exhibit Gaussian distributions with coefficients of variation (CV) of 10-15%. Strikingly narrower distributions are observed for their cell density (CV approximately 0.5%) and filterability (CV approximately 5-7%). This implies that V is highly correlated with H and A. We hypothesize that the RBC is able to adjust its volume to parameters H and A. It is tempting to speculate that intracellular free calcium (Cai) is a mediator in this process, acting as an activator of the Gardos channel. We tested this hypothesis by experimentally varying Cai and measuring changes in RBC density and filterability distributions. MATERIAL/METHODS: Three different approaches were used to raise Cai: (i) RBCs were loaded with Ca2+ in the presence of the calcium ionophore A23187; (ii) RBCs were incubated with Ca2+ in the presence of 1.0 mM ortho-vanadate; and (iii) the calcium pump was switched off by ATP depletion of RBCs. The density distribution of RBCs was determined by a phthalate technique. The distributions in filterability were obtained using a kinetic filtrometer. RESULTS: Whatever the approach used, the density and filterability distributions of treated RBCs broadened significantly in comparison with those of control cells. CONCLUSIONS: The results obtained suggest that (i) in vivo regulation of RBC volume is mediated by Cai and (ii) Cai probably depends on the A/V ratio, which determines the stress experienced by the RBC membrane in the circulation and, thereby, the calcium influx rate.