The membrane was blocked with blocking solution [3% BSA in Tris-buffered saline + Tween 20 (TBST)] for 1 h at room temperature. injury. A similar lack of congruence between effects on PMN migration and tissue injury has been reported in other disease models and for other adhesion molecules in models of ALI. Our results thus confirm the crucial role of JAM-A in PMN transmigration but demonstrate that transmigration is not essential for other aspects of inflammation or for lung injury in ALI. Keywords: myeloperoxidase, oxidant stress, lipopolysaccharide, acute respiratory distress syndrome, endothelial a variety of pulmonary and extrapulmonary insults can produce acute lung injury (ALI) (23). Pathophysiology of this disease is complex, but the earliest phases consistently feature severe neutrophil (PMN)-rich alveolar inflammation and resulting pulmonary injury (18). A major aspect of this injury is usually capillary leakage and pulmonary edema associated with hypoxemia and respiratory failure (17). Since there is currently no effective pharmacotherapy, the need for identification of novel therapeutic targets is urgent. Junctional adhesion molecule (JAM)-A is usually a member of the immunoglobulin superfamily expressed on endothelial and epithelial cells, leukocytes, and platelets (14, 25). On endothelial and epithelial surfaces, it colocalizes with tight junctions (16) and appears to be required for their formation (13, 15) but is not itself directly part of the tight junction structure (7). Tight junctions are the main determinants of vascular and mucosal permeability, and genetic deficiency of JAM-A has been shown to increase permeability of mouse intestinal mucosa and epithelial cell monolayers to dextran and other solutes (11). Adhesion molecules also play essential functions in migration of PMNs and monocytes from the vasculature into inflamed tissues. Woodfin and colleagues (26) showed that migration of PMNs from mouse cremasteric venules in response to IL-1 sequentially requires ICAM-1, JAM-A, and platelet endothelial cell adhesion molecule (PECAM)-1. Knockout of any one of these three molecules blocks IL-1-induced PMN migration, but at distinctly different locations within the endothelium. Interestingly, if transmigration is usually induced by TNF-, rather than IL-1, these molecules are required only if direct PMN activation by TNF- is usually blocked by deletion of their TNF- receptorsthat is usually, if TNF- can act only around the endothelial cells. These results imply that the mechanism of PMN transmigration depends on the activating stimulus. This dependence on the specific stimulus involved may account for the varying results obtained in modestly different animal models of disease. For example, evidence suggests that JAM-A is required for PMN transmigration in cytokine-induced meningitis (3), but not in meningitis induced by or lymphocytic MC-Val-Cit-PAB-dimethylDNA31 choriomeningitis computer virus (12). The mechanism by which PMNs interact with endothelial JAM-A has not been fully elucidated. Binding between JAM-A and the leukocyte-surface integrin L2, also known as lymphocyte function-associated antigen-1, has been exhibited (6, 19) and provides a plausible mechanism. However, interactions between PMN and endothelial JAM-A are also possible. JAM-A molecules homodimerize (10), and MC-Val-Cit-PAB-dimethylDNA31 dimerization of molecules on different cells has been shown to be important for maintenance of epithelial barrier function (15). The possibility that PMN JAM-A might be important for transmigration is supported by the observation that JAM-A-deficient (JAM-A?/?) PMNs show reduced transendothelial migration in inflamed peritoneum and myocardial ischemia-reperfusion injury (2). Surprisingly, however, deletion of JAM-A on endothelial cells, rather than PMNs, had no effect in this model. Although there have been several investigations PGR of the functions of other adhesion molecules in models of ALI (1, 4, 8, 21), the MC-Val-Cit-PAB-dimethylDNA31 role of JAM-A has not been addressed. Employing a.