PHAGOCYTOSIS OF MALARIAL PIGMENT HEMOZOIN INHIBITS NADPH-OXIDASE ACTIVITY IN HUMAN MONOCYTE-DERIVED MACROPHAGES

Upon stimulation, inactive subunits of monocyte NADPH oxidase (NOX) ar e assembled in the membrane to generate the active enzyme responsible for oxidative burst. Phosphorylation of the 47 kDa NOX cytoplasmic sub unit (47 kDa band) by protein kinase C (PKC) is important for NOX asse mbly and activation. Alternatively, NOX is activated in vitro by sodiu m dodecyl sulfate (SDS) or amphiphiles via a phosphorylation-independe nt mechanism. Previous data indicate that phagocytosis of malarial pig ment hemozoin inhibits oxidative burst and PKC activity (Schwarzer, E. , Turrini, F., Giribaldi, G., Cappadoro, M. and Arese, P. (1993)Biochi m. Biophys. Acta, 1181, 51-54). We show here that SDS-stimulated NOX a ctivity and phorbol 12-myristate 13-acetate (PMA)-induced oxidative bu rst dropped by 54% and 36% of control values 2 h after hemozoin phagoc ytosis, respectively. SDS-stimulated NOX activity remained roughly con stant until 12 h: whereas oxidative burst dropped further by approx. 6 0% and 75% of control values 6 h and 12 h after hemozoin phagocytosis. Reconstitution experiments indicate that damage was localized to cyto solic NOX subunit(s). Membrane assembly of active NOX was defective in PMA- (PKC-dependent stimulation) and FMLP- (PKC-dependent and indepen dent stimulation) stimulated hemozoin-fed monocytes. Labeling experime nts with [P-32]orthophosphate or [gamma-P-32]ATP showed that endogenou s PKC-dependent phosphorylation of the 47 kDa band was unaffected 12 h and impaired only 24 h after hemozoin phagocytosis. Thus, only long-t erm inhibition of NOX may additionally depend on superimposed PKC inhi bition.