N-Acetyl Cysteine Inhibits Induction of No Production By Endotoxin or Cytokine Stimulated Rat Peritoneal Macrophages, C 6 Glial Cells and Astrocytes

Abstract

The present study underscores the importance of N-acetyl cysteine (NAC), a potent antioxidant, in inhibiting the induction of NO production by lipopolysaccharides (LPS) and cytokines in peritoneal macrophages, C 6 glial cells and primary astrocytes. LPS, interleukin-1 beta (IL-1β), interferon-gama (IFN-γ) and tumor necrosis factor-alpha (TNF-α) alone or in combinations induced the production of NO to different degrees. NAC when added 2 h earlier to the addition of these stimuli potentially blocked the increase in NO production in macrophages, astrocytes and C 6 glial cells. The decrease in NO production by NAC was accompanied by a decrease in inducible nitric oxide synthase (iNOS) activity, in iNOS protein detected by immunoblot analysis with antibodies against iNOS, and in iNOS mRNA determined by reverse-transcriptase coupled polymerase chain reaction (RT-PCR). Time course studies show that inhibition was maximum when NAC was added 2 h prior to the addition of LPS and the degree of inhibition decreased progressively with the increase in time interval when NAC was added after the addition of LPS. In addition to NAC, another antioxidant pyrrolidine dithiocarbamate (PDTC) was also found to inhibit the induction of NO production effectively. Since activation of NF-κB is necessary for the induction of iNOS, we examined the effect of NAC on the activation of NF-κB. Inhibition of LPS-induced activation of NF-κB by NAC in rat peritoneal macrophages suggests that the inhibitory effect of NAC on the induction of iNOS is due to the inhibition of NF-κB. Besides NO, NAC also blocked the production of TNF-α in rat peritoneal macrophages activated with endotoxin. These results suggest that expression of iNOS and TNF-α in macrophages do involve oxygen radicals. The importance of these results in relation to controlling various harmful effects of cytokines released by activated macrophages and glial cells is discussed.