EVIDENCE FOR A CYCLIC GUANOSINE MONOPHOSPHATE-DEPENDENT, CARBON MONOXIDE-MEDIATED, SIGNALING SYSTEM IN THE REGULATION OF TNF-ALPHA PRODUCTION BY HUMAN PULMONARY MACROPHAGES

Background and Objectives: The second messenger cyclic guanosine 3',5' -monophosphate (cGMP) seems to be implicated in the release of tumor n ecrosis factor alpha (TNF-alpha) by activated macrophages. There is co ntroversy regarding the potential of human macrophages to produce nitr ic oxide (NO). Since guanylate cyclase can be activated also by carbon monoxide (GO) and this gas may be formed endogenously, we examined th e ability of human pulmonary macrophages to produce CO in the presence of lipopolysaccharide (LPS) or LPS + interferon gamma (IFN-gamma). In addition, the source and the relative contribution of this molecule t o the LPS-induced increase in cell cGMP content and TNF-alpha release were explored. Design: Interstitial macrophages were obtained from mul tiple organ donor lungs by enzymatic digestion. After 24-hour precultu re, purified macrophages were cultured for 24 hours in the presence or absence of LPS, LPSS+IFN-gamma, CO (250 and 500 mu mol/L), sodium nit roprusside, 8-Br-cGMP, hemoglobin, methylene blue, zinc-protoporphyrin IX, hemin, S-adenosylmethionine, deferoxamine mesylate, or combinatio ns. The cGMP content of the cells and TNF-alpha, CO, and NO release to the medium were determined. Results: In the presence of LPS, TNF-alph a production was not accompanied by any detectable increase in the NO release to the medium. However, an increase in medium CO concentration (mean +/- SEM) (5.81 +/- 0.20 vs 3.74 +/- 0.08 pmol/mu g protein; n = 11; P < .01) and cell cGMP content (0.273 +/- 0.021 vs 0.138 +/- 0.01 9 pmol/mu g protein; n = 10; P < .01) was observed. These changes were more pronounced in the presence of LPS + IFN-gamma. Release of TNF-al pha also was induced by both sodium nitroprusside and 8-Br-cGMP. In co ntrast, methylene blue, a guanylate cyclase inhibitor, inhibited LPS-, LPS + IFN-gamma-, and sodium nitroprusside-induced TNF-alpha producti on and cGMP increase; hemoglobin, which traps CO, had a similar effect . Conclusion: Intracellular cGMP increase, secondary to an endogenous production of CO, participates in the release of TNF-alpha by activate d human pulmonary macrophages.