Na+ and K+ transport damage induced by oxygen free radicals in human red cell membranes.

The treatment of human erythrocytes with phenazine methosulfate (PMS) results in a sustained intracellular production of oxygen free radicals as shown by the reduction of nitroblue tetrazolium and methemoglobin production.Inhibition of superoxide dismutase by diethyldithiocarbamate further enhances nitroblue tetrazolium reduction indicating an increase in the PMSdependent oxygen free radicals.NADH is one of the hydrogen donors able to react with PMS and O2 in order to generate oxygen free radicals in cyclic reactions which last far more than 4 h.The attack of red cell membranes by oxygen free radicals greatly alters their chemical structure.We observed lipid peroxidation, as shown by the increase in malondialdehyde above endogenous levels and methemoglobin binding to the cell membrane.PMS treatment markedly modifies the ionic equilibrium of the erythrocytes.At low PMS concentrations they lose intracellular K' .This is a prehemolytic effect, since hemolysis occur at high PMS concentrations.Both toxic effects are enhanced by superoxide dismutase inhibition with diethyldithiocarbamate, strongly suggesting that they result from the attack of cell membranes by oxygen free radicals.A study of the effect of PMS on the different transport pathways for K' across the human red cell membrane showed that at low PMS concentrations there is a specific increase in passive K+ permeability with no major effect on the specific K' carriers such as the (Na+,K+)-pump or the (Na+,K+)cotransport system.The increase in passive K+ permeability has the following properties: (i) it is not affected by ethylene glycol bis(P-aminoethyl ether)-N,Nflfltetraacetic acid, quinine, or the replacement of C1by NO3-; (ii) it is enhanced by inhibition of the anion carrier with 4,4'-diisothiocyano-2,!2'-disulfonic acid stilbene; (iii) it is partially inhibited by external superoxide dismutase; and (iv) it is not affected by external catalase or by inhibition of endogenous catalase with 3-amino-1,2,4-triazole.Our results strongly suggest that the PMS-dependent erythrocyte K' loss results from membrane lipid peroxydation by oxygen free radicals, particularly 0 $ The generation of free radicals following irradiation (1, 2), disturbance of tissue oxygen supply (3-7), inflammation (8lo), and other physiopathological conditions (11, 12) may be extremely noxious to the structure and function of cell mem-