Cell volume-dependent regulation of L-selectin shedding in neutrophils - Arole for p38 mitogen-activated protein kinase

Neutrophil-mediated organ damage is a common feature of many disease states . We previously demonstrated that resuscitation with hypertonic salt soluti ons prevented the endotoxin-induced leukosequestration and consequent lung injury, and this effect was partially attributed to an altered surface expr ession of adhesion molecules, CD11b and L-selectin. In this study we invest igated the mechanisms whereby osmotic stress evokes L-selectin shedding, Th e metalloprotease inhibitor RO 31-9790 prevented the osmotic down-regulatio n of L-selectin, indicating that this process was catalyzed by the same "sh eddase" responsible for L-selectin cleavage induced by diverse inflammatory stimuli. The trigger for hypertonic shedding was cell shrinkage and not in creased osmolarity, ionic strength, or intracellular pH. Volume reduction c aused robust tyrosine phosphorylation and its inhibition by genistein and e rbstatin abrogated shedding. Shrinkage stimulated tyrosine kinases Hck, Syk , and Pyk2, but prevention of their activation by the Src-family inhibitor PP1 failed to affect the L-selectin response, Hypertonicity elicited the Sr c family-independent activation of p38, and the inhibition of this kinase b y 8B203580 strongly reduced shedding, p38 was also essential for the N-form yl-methionyl-leucyl-phenylalanine- and lipopolysaccharide-induced shedding but not the phorbol ester-induced shedding. Thus, cell volume regulates L s electin surface expression in a p38-mediated, metalloprotease dependent man ner. Moreover, p38 has a central role in shedding induced by many inflammat ory mediators.