NO is a potent anti-inflammatory agent. Exogenous NO donors decrease leukocyte adherence to the vessel wall; conversely, NOS inhibitors increase leukocyte rolling along the endothelium [1
]. NO derived from each of the NOS isoforms inhibits leukocyte interactions with the vessel wall. Leukocyte adherence to the vessel wall is elevated 10-fold in eNOS knockout mice and 4-fold in nNOS knockout mice compared to wild-type mice [2
]. LPS induces a 3-fold higher number of leukocytes rolling on venules in iNOS knockout mice than in wild-type mice [3
]. NO also inhibits inflammation in diverse murine models of vascular disease, including myocardial infarction, glomerulonephritis, lung injury, and stroke [4
]. Thus NO derived from each of the NOS isoforms inhibits vascular inflammation. How?
Clues to the anti-inflammatory mechanism of NO came from studies of leukocyte interactions with the vessel wall. Leukocyte trafficking involves 5 discrete stages: (1) Rolling: loose attachment of leukocytes to endothelial cells is mediated by selectins and their glycoprotein ligands. (2) Activation: chemokines and other pro-inflammatory signals activate leukocytes and endothelial cells, inducing a variety of rapid responses including integrin surface expression and conformational changes. (3) Adherence: leukocytes stop rolling and become fixed to endothelial cells, mediated by intercellular adhesion molecules (ICAM) and their integrin receptors. (4) Diapedesis: leukocytes travel between endothelial cells into the vascular wall. (5) Migration: leukocytes follow a chemotactic gradient to the site of tissue injury.
NO inhibits the first stage of leukocyte trafficking, leukocyte rolling. Leukocyte rolling is mediated in part by P-selectin, and NO decreases expression of P-selectin on the endothelial surface [7
]. P-selectin is a transmembrane protein normally stored within resting endothelial cells in granules called Weibel-Palade bodies (WPB) [8
]. However, inflammatory signals such as histamine, hypoxia, or oxidized LDL activate endothelial exocytosis of these granules [11
]. During exocytosis, the membranes of the WPB rapidly fuse with the endothelial plasma membrane, translocating P-selectin to the exterior. On the outer surface of the endothelial cell, P-selectin interacts with its ligand P-selectin glycoprotein ligand-1 (PSGL-1) on the surface of leukocytes, causing transient leukocyte rolling along the vessel wall [12
]. Since exocytosis of WPB triggers the initial phase of leukocyte rolling, NO inhibition of exocytosis limits vascular inflammation.
NO also inhibits other stages of leukocyte trafficking. NO decreases expression of integrins and intercellular adhesion molecules necessary for the third stage of leukocyte rolling, leukocyte adherence [14
]. One possible mechanism may be through S-nitrosylation of nuclear factor kappaB, a transcription factor that drives expression of many pro-inflammatory molecules [18
]. Furthermore, NO blunts the second sage of leukocyte trafficking, leukocyte and endothelial activation. For example, NO limits cellular activation by acting as an anti-oxidant. Superoxide is an innate immune effector produced by NADPH oxidases in neutrophils and macrophages. Superoxide can injure cells in part by oxidizing proteins, lipids, and nucleic acids. However, NO can react rapidly with superoxide, and diminish superoxide toxicity [19
]. Furthermore, NO can directly inhibit the NADPH oxidase that produces superoxide [22
]. NO also limits inflammation induced by other oxygen radicals such as hydrogen peroxide [23
]. Thus NO limits leukocyte trafficking at multiple stages.