ENTAMOEBA-HISTOLYTICA MODULATES THE NITRIC-OXIDE SYNTHASE GENE AND NITRIC-OXIDE PRODUCTION BY MACROPHAGES FOR CYTOTOXICITY AGAINST AMEBAS AND TUMOR-CELLS

Nitric oxide (NO) is the major cytotoxic molecule produced by activated macrophages for cytotoxicity against Entamoeba histolytica trophozoites. In the present study, we determined whether E. histolytica infection and soluble amoebic proteins affected macrophage cytotoxicity against amoebae and tumour cells by modulating the inducible NO synthase gene (iNOS) and NO (measured as nitrite, NO2-) and tumour necrosis factor-alpha (TNF-alpha) production. Amoebic liver abscess-derived macrophages [days 10, 20, 30 post-infection (p.i.)] stimulated with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS) showed increased cytotoxicity against L929 cells (TNF-alpha-sensitive), but were refractory for killing amoebae and P815 cells (both NO-sensitive), concomitant with low NO2- production (< 4 mu M/10(6) cells). In contrast, peritoneal and spleen macrophages at 10 and 20 days p.i. activated with IFN-gamma and LPS demonstrated increased killing of amoebae, and L929 and P815 cells concomitant with high NO2- production (> 12 mu M/10(6) cells). Pretreatment of mouse bone marrow-derived macrophages with amoebic proteins suppressed IFN-gamma and LPS-induced amoebicidal (33%) and tumoricidal (44-49%) activities, with a corresponding decrease in TNF-alpha (56%) and NO (41%) production as well as TNF-alpha (41%) and iNOS (27%) mRNA by Northern blot analyses as compared to untreated activated controls. Inhibition of prostaglandin E(2) (PGE(2)) biosynthesis in abscess and naive macrophages pretreated with amoebic proteins augmented IFN-gamma- and LPS-induced killing of L929 cells and TNF-alpha production, but failed to increase killing of P815 cells and amoebae as well as iNOS mRNA levels or NO production. These results suggest that E. histolytica selectively induces dysfunction of macrophage cytotoxicity by modulating iNOS mRNA expression and NO production independent from TNF-alpha and PGE(2) allowing the parasites to survive within the host by impairing host immune responses.