Inhibition of ␣4␤1/vascular cell adhesion molecule-1 (VCAM-1) interactions have therapeutic potential in treating allergic airway disease because of the importance of these adhesion molecules in the trafficking of eosinophils, lymphocytes, and monocytes. We examined several small molecule inhibitors of ␣4␤1/VCAM-1 interactions with in vitro potencies (IC 50 values) ranging from 0.52 nM (CP-664511; 3-The same compounds were evaluated in vivo using a murine model of ovalbumin-induced pulmonary eosinophilia. In this model, systemic administration of antibodies against ␣4 reduced bronchoalveolar lavage (BAL) eosinophilia ϳ60%. Small molecule ␣4␤1 antagonists were administered by intratracheal instillation and demonstrated dose-dependent inhibition of BAL eosinophil numbers and achieved a maximum inhibition of ϳ60%. In general, the rank order of potency for these compounds in vitro was consistent with that observed in vivo, which confirms that their efficacy is likely via blockade of ␣4␤1/VCAM-1 interactions. The most potent compound, CP-664511, also inhibited BAL eosinophilia following s.c. administration (1-10 mg/kg, s.c.). These data support the utility of small molecule ␣4␤1 antagonists in the treatment of relevant diseases, such as asthma.

p59fynT is a protein tyrosine kinase in the src family that has been associated with and believed to function in the signaling of many receptors, including the T-cell receptor. A role for the kinase in antigen-driven pulmonary inflammation was examined using mice whose p59fynT gene had been genetically ablated. FynKO mice that were sensitized to ovalbumin exhibited a marked increase in bronchoalveolar lavage eosinophils and cytokines, including interleukin (IL)-4 and IL-5, relative to wild-type mice in response to antigen aerosol exposure. Ovalbumin-stimulated IL-5 production was also increased in cultured splenocytes derived from fynKO mice relative to wild-type mice, whereas interferon-gamma levels were unchanged. Diminished concanavalin A--stimulated IL-4 levels from fynKO splenocytes were consistent with reduced serum immunoglobulin (Ig)E levels observed in sensitized/saline aerosol-challenged animals and may reflect defective natural killer 1.1(+) T cell development. Normalization of IgE levels in sensitized fynKO mice relative to wild-type mice occurred after repeat antigen challenge, which suggests a secondary source of IL-4. Overall, these data demonstrate fyn is a negative regulator of allergic airway inflammation in mice because its absence promotes a shift to a T helper-2 phenotype that may reflect the kinase's role in T-cell receptor signaling.

1 The ability of various C-C chemokines to elicit tissue eosinophil in®ltration following intradermal injection or peripheral blood eosinophilia following intravenous injection were compared in the Brown-Norway rat. 2 Eotaxin (0.1 ± 3 mg site 71 ) of human and murine origin produced equivalent, dose-dependent increases in eosinophil peroxidase activity in rat dermis 4 h post-injection. 3 Human eotaxin-2 was equipotent with human eotaxin in terms of dermal eosinophil recruitment. Other human CCR3 agonists, such as MCP-3, RANTES and MCP-4 failed to increase dermal eosinophil peroxidase activity at doses up to 1 mg site 71 whereas the latter did produce a small e ect at 3 mg site 71 . 4 Consistent with observations in vivo, human eotaxin displaced [ 125 I]-eotaxin from rat spleen membranes more potently (IC 50 =2 nM) than did MCP-4 (IC 50 =500 nM). RANTES did not compete with the radiolabelled chemokine at concentrations up to 1 mM. 5 Human eotaxin (5 mg) administered intravenously increased circulating eosinophils *3 fold whereas MCP-4 (5 mg, i.v.) increased circulating monocytes *3 fold without a ecting eosinophil numbers. 6 Dexamethasone pretreatment inhibited eotaxin-induced dermal eosinophil in¯ux only at a steroid dose (0.1 mg kg 71 , s.c.) which signi®cantly reduced circulating eosinophil numbers. The steroid also reduced eosinophilia in peripheral blood resulting from systemic eotaxin administration (5 mg, i.v.). 7 These data suggest di erences in rat CCR3 relative to other species as surmised from a distinctive rank order of chemokine potency. In addition to its chemotactic e ects eotaxin, but not MCP-4, promotes eosinophil recruitment into the circulation. One of the mechanisms by which glucocorticoids, such as dexamethasone, acutely inhibits eotaxin-induced dermal eosinophil in¯ux is to diminish the circulating numbers of these cells available for tissue recruitment.

No abstract

Synergistic interactions between cytokines, chemokines and adhesion molecules may facilitate the selective recruitment of eosinophils into sites of allergic inflammation. Ovalbumin-sensitized IL5TG mice responded to antigen challenge with robust airway eosinophilia 24 and 72 hr post-exposure. Adhesion molecule expression and functional responsiveness of immune cells derived from IL5TG mice to various inflammatory mediators were evaluated. IL5TG-derived eosinophils, but not neutrophils, expressed higher levels of CD49d and CD11b relative to WT. Functional responsiveness to eotaxin was increased in IL5TG eosinophils as demonstrated by a 10x increase in its potency in producing actin polymerization and 3x increase in CD11b upregulation relative to WT. These data are consistent with increased CCR3 expression on IL5TG eosinophils. Responsiveness of eosinophils to LTB4 or MIP-1alpha was similar between WT and IL-5TG mice. These data provide evidence of synergy between eosinophil-specific cytokines and chemokines that may promote accumulation of this cell type under conditions of allergic inflammation in vivo.