alpha(4)-Integrin mediates neutrophil-induced free radical injury to cardiac myocytes

Previous work has demonstrated that circulating neutrophils (polymorphonucl ear leukocytes [PMNs]) adhere to cardiac myocytes via beta (2)-integrins an d cause cellular injury via the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enzyme system. Since PMNs induced to leave the vasculature (emigrated PMNs) express the alpha (4)-integrin, we asked whether (a) thes e PMNs also induce myocyte injury via NADPH oxidase, (b) beta (2)-integrins (CD18) still signal oxidant production, or if this process is now coupled to the alpha (4)-integrin; and (c) dysfunction is superoxide dependent with in the myocyte or at the myocyte-PMN interface. Emigrated PMNs exposed to c ardiac myocytes quickly induced significant changes in myocyte function. My ocyte shortening was decreased by 30-50% and rates of contraction and relax ation were reduced by 30% within the first 10 min. Both alpha (4)-integrin antibody (Ab)-treated PMNs and NADPH oxidase-deficient PMNs were unable to reduce myocyte shortening. An increased level of oxidative stress was detec ted in myocytes within 5 min of PMN adhesion. Addition of an anti-alpha (4) -integrin Ab, but not an anti-CD18 Ab, prevented oxidant production, sugges ting that in emigrated PMNs the NADPH oxidase system is uncoupled from CD18 and can be activated via the alpha (4)-integrin. Addition of exogenous sup eroxide dismutase (SOD) inhibited all parameters of dysfunction measured, w hereas overexpression of intracellular SOD within the myocytes did not inhi bit the oxidative stress or the myocyte dysfunction caused by the emigrated PMNs. These findings demonstrate that profound molecular changes occur wit hin PMNs as they emigrate. such that CD18 and associated intracellular sign aling pathways leading to oxidant production are uncoupled and newly expres sed alpha (4)-integrin functions as the ligand that signals oxidant product ion. The results also provide pathological relevance as the emigrated PMNs have the capacity to injure cardiac myocytes through the alpha (4)-integrin -coupled NADPH oxidase pathway that can be inhibited by extracellular, but not intracellular SOD.