Modulation of human neutrophil inflammatory responses by nitric oxide: studies in unprimed and LPS-primed cells

Abstract

Because nitric oxide (NO) can act both as a regulatory and as a toxic molecule, we have studied N-formyl-methionyl-leucyl-phenylalanine (fMLF) -stimulated responses of human neutrophils (PMNs) during various conditions of NO modulation in unprimed and bacterial lipopolysaccharide (LPS) -primed cells. Effects of various NO modulators were assessed on stimulated superoxide (O2-) generation, granule exocytosis, homotypic aggregation, and rises in intracellular free Ca2+ ([Ca2+]i). Significant differences in the effects of various NO modulators on inflammatory responses of PMNs kept in stirred suspension versus those kept under static and/or adherent conditions, were observed. L-arginine, the physiological substrate for NO synthase (NOS), and NG-nitro-L-arginine methyl ester, an inhibitor of NOS, both caused a 40-50% inhibition of LPS-induced priming of O2- generation in PMNs in stirred suspension, but not in LPS-primed PMNs under static or adherent conditions. The NO donors, sodium nitroprusside and S-nitroso-N-acetylpenicillamine, completely abrogated the LPS-induced priming of 02- generation in PMNs in suspension, while causing only a 40-50% inhibition in PMNs under static or adherent conditions. The Ca2+ ionophore, A23187, prevented the LPS-induced priming of 02-generation without affecting 02- generation in unprimed PMNs. LPS priming of PMNs induced about a twofold increase in fMLF-stimulated homotypic aggregation, exocytosis of secondary granules, and rises in [Ca2+]i. In related studies, we also provide definitive evidence for enzymatic formation of NO in human PMNs and demonstrate a significant decrease in NO levels in LPS-primed PMNs. Taken together, these findings indicate that NO modulates PMN inflammatory responses and plays a protective role in priming and activation processes of inflammatory PMNs.

Interactive role of nitric oxide and superoxide anion in neutrophil-mediated endothelial cell injury

“…Nitrite levels were measured as an index of NO generation in the culture supernatants, by a sensitive fluorimetric method of Misko et al [15], the details of which have been recently reported from our laboratory [16]. For optimal NO generation, PMN or EC were supplemented with L-arginine (200 µM).…”

Induction of NOS in rat blood PMN in vivo and in vitro: modulation by tyrosine kinase and involvement in bactericidal activity

“…We recently demonstrated that NO is involved in the candidacidal activity of rat inflammatory peritoneal neutrophils [15] and other authors have shown that NO contributes to the tumoricidal activity of those cells [16]. In addition, the NO-generating system in neutrophils has been suggested to function as an autocrine mechanism that may regulate PMN activation during various pathophysiological conditions in vivo [17]. Therefore, it is important to elucidate the modulatory mechanisms involved in the induction of the NOS pathway in neutrophils.…”

Activation of p38 Mitogen-activated Protein Kinase by Lipopolysaccharide in Human Neutrophils Requires Nitric Oxide-dependent cGMP Accumulation

“…In Jurkat T-cells, long-term exposure to nitric oxide (NO) leads to activation of several MAP kinases, with ERK being the most responsive (21). LPS induces NO production in neutrophils, and this process is thought to mediate many of its inflammatory activities (22)(23)(24)(25)(26)(27). Stimulation of neutrophils with fMLF can also induce NO production, which is thought to play an important role in chemotaxis and degranulation elicited by this agent (28 -32).…”