Transforming growth factor-beta 1 primes macrophages for enhanced expression of the nitric oxide synthase gene for nitric oxide-dependent cytotoxicity against Entamoeba histolytica.

Nitric oxide (NO) produced by activated macrophages is the major cytotoxic molecule for in vitro cytotoxicity against Entamoeba histolytica trophozoites. Transforming growth factor-beta 1 (TGF-beta 1) is a potent negative regulator of several macrophage functions, including NO production. In this study, we investigated the effect of TGF-beta 1 on macrophage nitric oxide synthase (mac-NOS) mRNA expression and NO production for macrophage cytotoxicity against E. histolytica trophozoites. TGF-beta 1 by itself was incapable of inducing mouse bone marrow-derived macrophage (BMM) amoebicidal activity and NO production (as measured by nitrite). In contrast, TGF-beta 1 pretreatment (4 hr) primed BMM for an enhanced amoebicidal activity of 15% and 23% in response to (interferon-gamma) IFN-gamma+tumour necrosis factor-alpha (TNF-alpha) or IFN-gamma+lipopolysaccharide LPS, concomitant with increased NO production of 85% and 27%, respectively. TGF-beta 1 pretreatment increased NO production in response to IFN-gamma+TNF-alpha/LPS stimulation in a time- and dose-dependent manner. By Northern blot analysis, the increased NO production of TGF-beta 1-pretreated BMM was preceded by markedly enhanced expression of mac-NOS mRNA. The priming effect of TGF-beta 1 on NO production was critically dependent on both a TNF-alpha (> or = 100 U) and a LPS (> or = 100 ng) triggering dose in the presence of IFN-gamma. TGF-beta 1 pretreatment enhanced TNF-alpha mRNA expression, but had no effect on TNF-alpha production in culture supernatants after 4 hr of stimulation with IFN-gamma+TNF-alpha/LPS; however, at a later time-point (16-48 hr), even though the levels of TNF-alpha mRNA expression were unaffected, TNF-alpha production was reduced. These data demonstrate that TGF-beta 1 priming for increased mac-NOS mRNA expression for NO-dependent cytotoxicity against E. histolytica in response to IFN-gamma+TNF-alpha/LPS stimulation may be involved in the modulation of a TNF-alpha triggering signal by TGF-beta 1.