TAURINE CHLORAMINE INHIBITS PRODUCTION OF NITRIC-OXIDE AND TNF-ALPHA IN ACTIVATED RAW-264.7 CELLS BY MECHANISMS THAT INVOLVE TRANSCRIPTIONAL AND TRANSLATIONAL EVENTS

We previously reported that taurine chloramine (Tau-Cl) inhibits the p roduction of both nitric oxide and TNF-alpha by activated RAW 264.7 ce lls. The current studies were conducted to gain insight into the mecha nisms through which Tau-Cl exerts these effects. RAW 264.7 cells were activated by LPS (10 mu g/ml) and rIFN-gamma (50 U/ml) in the absence or presence of either 0.8 mM Tau-Cl or taurine. Production of NO and T NF-alpha by RAW 264.7 cells was monitored: NO was measured spectrophot ometrically as nitrite and TNF-alpha was measured by ELISA. Cell lysat es were analyzed for the inducible form of nitric oxide synthase (iNOS ) by Western blot analyses, and TNF-alpha and iNOS mRNAs were assessed by northern blot analyses. Tau-Cl inhibited transcription of the iNOS gene, or some earlier event in the signal transduction pathway, becau se iNOS protein and iNOS mRNA were undetected in lysates of cells acti vated in the continuous presence of Tau-Cl. In contrast, steady-state levels of TNF-alpha mRNA increased in the presence of Tau-Cl to at lea st the same extent as that in untreated activated cells and persisted for a longer period of time. Metabolic labeling experiments demonstrat ed that Tau-Cl inhibited translation of TNF-alpha mRNA because the pre sence of the presecretory 26-kDa form and the secreted 17-kDa form of TNF-alpha were greatly reduced in lysates and culture media, respectiv ely, of cells activated in the presence of Tau-Cl. Inhibition of TNF-a lpha synthesis by Tau-Cl is not the result of a generalized effect on protein synthesis because the amount of radiolabeled protein precipita ted from metabolically labeled cells by TCA was unaffected by Tau-Cl, and cell viability was unaffected. The results of these studies demons trate that Tau-Cl decreases production of tissue-damaging inflammatory mediators and thus may act as a physiologic modulator of macrophage f unction.