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. 1994 May;62(5):1534–1541. doi: 10.1128/iai.62.5.1534-1541.1994

Tumor necrosis factor alpha augments nitric oxide-dependent macrophage cytotoxicity against Entamoeba histolytica by enhanced expression of the nitric oxide synthase gene.

J Y Lin 1, R Seguin 1, K Keller 1, K Chadee 1
PMCID: PMC186349  PMID: 7513301

Abstract

Nitric oxide (NO measured as nitrite, NO2-) is the major effector molecule produced by activated macrophages for in vitro cytotoxicity against Entamoeba histolytica trophozoites. In this study, we determine whether tumor necrosis factor alpha (TNF-alpha) produced by activated bone marrow-derived macrophages (BMM) is involved in the induction of the inducible NO synthase gene (mac-NOS) for NO-dependent amebicidal activity. TNF-alpha alone did not directly induce macrophage NO2- production to kill amebae; however, in combination with increasing concentrations of TNF-alpha and gamma interferon (IFN-gamma), BMM amebicidal activity and NO2- production progressively increased and showed a significant linear correlation. Antiserum to TNF-alpha and the NO synthase inhibitor NG-monomethyl L-arginine (L-NMMA) inhibited the synergistic effects of TNF-alpha and IFN-gamma. BMM activated with increasing concentrations of lipopolysaccharide (LPS) and IFN-gamma showed a significant linear correlation between TNF-alpha release and NO2- production. Antiserum to TNF-alpha suppressed TNF-alpha release, NO2- production, and amebicidal activity by 93, 53, and 86%, respectively. L-NMMA diminished NO2- production by 74% and macrophage amebicidal activity by 83% but had no effect on TNF-alpha release. Quantification by Northern (RNA) blot analyses demonstrated that IFN-gamma in combination with TNF-alpha or LPS increased markedly the accumulation of mac-NOS and TNF-alpha mRNAs in a time-dependent manner with a concomitant increase in NO and TNF-alpha production. Peak induction of mac-NOS occurred after 24 h, whereas TNF-alpha mRNA was rapidly expressed after 4 h and remained stable for 48 h. Taken together, these data argue that TNF-alpha augments NO-dependent macrophage cytotoxicity against E. histolytica via elevated levels of mac-NOS mRNA expression which may be associated with the accumulation of TNF-alpha mRNA.

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Selected References

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