| Abstract |
Although the germicide role of H2O2 released during inflammation is well established, a hypothetical regulatory function, either promoting or inhibiting inflammation, is still controversial. In particular, after 15 years of highly contradictory results it remains uncertain whether H2O2 by itself activates NF-kappa B or if it stimulates or inhibits the activation of NF-kappa B by proinflammatory mediators. We investigated the role of H2O2 in NF-kappa B activation using, for the first time, a calibrated and controlled method of H2O2 delivery-the steady-state titration-in which cells are exposed to constant, low, and known concentrations of H2O2. This technique contrasts with previously applied techniques, which disrupt cellular redox homeostasis and/or introduce uncertainties in the actual H2O2 concentration to which cells are exposed. In both MCF-7 and HeLa cells, H2O2 at extracellular concentrations up to 25 mu M did not induce significantly per se NF-kappa B translocation to the nucleus, but it stimulated the translocation induced by TNF-alpha. For higher H2O2 doses this stimulatory role shifts to an inhibition, which may explain published contradictory results. The stimulatory role was confirmed by the observation that 12.5 mu M H2O2, a concentration found during inflammation, increased the expression of several proinflammatory NF-kappa B-dependent genes induced by TNF-alpha (e.g., IL-8, MCP-1, TLR2, and TNF-a). The same low H2O2 concentration also induced the anti-inflammatory gene coding for heme oxygenase-1 (HO-1) and IL-6. We propose that H2O2 has a fine-tuning regulatory role, comprising both a proinflammatory control loop that increases pathogen removal and an anti-inflammatory control loop, which avoids an exacerbated harmful inflammatory response. |
