INHIBITORY EFFECTS OF N-ACETYLCYSTEINE ON SUPEROXIDE ANION GENERATIONIN HUMAN POLYMORPHONUCLEAR LEUKOCYTES

It has been suggested that reactive oxygen species released by activat ed polymorphonuclear leukocytes (PMN) in man is one mechanism of tissu e injury. Therapeutic action aimed at increasing antioxidant defence m echanisms is still a clinical challenge. This study examines the activ ity of N-acetylcysteine, a known antioxidant, in the protection of PMN exposed in-vitro to the chemoattractant peptide fMet-Leu-Phe (FMLP), the protein kinase C activator phorbol myristate acetate or the lipid peroxidation promoter t-butyl hydroperoxide. FMLP (3-300 nM) and phorb ol myristate acetate (160 pm-160 nM) induced concentration-related sup eroxide anion generation. Pre-treatment with N-acetylcysteine (33-333 mu M) resulted in concentration-related inhibition of superoxide produ ction induced by FMLP (30 nM) or phorbol myristate acetate (16 nM); -l og IC50 values were 3.97 +/- 0.07 and 3.91 +/- 0.10, respectively. Cha nges in intracellular calcium ion concentration ([Ca2+](i)) induced by FMLP (30 nM) were studied in fura-2-loaded human PMN. FMLP produced a transient calcium response, i.e. a peak followed by decay to a residu al value above baseline. N-Acetylcysteine (333 mu M) did not affect ei ther basal [Ca2+](i) values or changes in [Ca2+](i) values after treat ment with FMLP. Activation by phorbol myristate acetate caused a reduc tion in glutathione levels from 5.94 +/- 0.86 (control) to 1.84 +/- 0. 51 nmol/3 x 10(6) cells (P<0.05 compared with control). Pre-treatment with N-acetylcysteine (333 mu M) fully reversed the reduction in gluta thione levels induced by phorbol myristate acetate (4.83 +/- 0.68 nmol /3 x 10(6) cells; P>0.05 compared with control). Exposure to t-butyl h ydroperoxide (0.5 mM, 30 min) markedly increased malondialdehyde level s (from 0.03 +/- 0.02 to 0.73 +/- 0.07 nmol/10(6) cells), and index of lipid peroxidation. Malondialdehyde levels were significantly reduced in PMN treated with N-acetylcysteine (333 mu M; 0.55 +/- 0.04 nmol/10 (6) cells; P<0.05 compared with untreated cells exposed to t-butyl hyd roperoxide). In conclusion, N-acetylcysteine reduces superoxide genera tion in response to FMLP and phorbol myristate acetate and partially p rotects against lipid peroxidation in PMN from man. The protection aff orded by N-acetylcysteine is not related to alteration of the intracel lular calcium signal but might be effected by replenishment of the int racellular glutathione levels.