EOSINOPHIL VLA-4 BINDING TO FIBRONECTIN AUGMENTS BRONCHIAL NARROWING THROUGH 5-LIPOXYGENASE ACTIVATION

We examined the effect of ligation of human eosinophils activated by p latelet-activating factor (PAF) to soluble human fibronectin (FN) on t he augmented contractile response of human bronchial explants. Styrene microplate wells were FN-coated, and eosinophils were allowed to adhe re in the presence of 1) buffer control, 2) 20 mu g/ml monoclonal anti body (HP2/1) to the alpha(4) beta(1) ligand (VLA-4) on the eosinophils , 3) 20 mu g/ml anti-CD18 R15.7, 4) 20 mu g/ml anti-CD16 3G8, or 5) 10 (-6) M A63162, a 5-lipoxygenase inhibitor. Sixty minutes later, treate d cells were activated with either buffer or 10(-6) M PAF. Airway lumi nal diameter was assessed by computerized videomicrometry as a functio n of pixel number, and activation of eosinophils was confirmed by meas urement of leukotriene C-4 (LTC(4)) secretion. Ligation with FN caused an increase in PAF-stimulated LTC(4) secretion from 276 +/- 75.6 pg/1 0(6) cell at baseline to 606 +/- 90.2 pg/10(6) cell (P < 0.01). This c orresponded to augmented luminal narrowing of human bronchial explants from 25.3 +/- 9.39% (PAF activation alone) to 42.9 +/- 8.0% (PAF-acti vated eosinophils + FN) (P < 0.012. Both augmented airway luminal narr owing and in creased LTC(4) secretion caused by PAF-activated cells af ter FN ligation were blocked completely by anti-VLA-4 MAb (P < 0.05 vs . control). Pretreatment with 10(-6) MA63162 inhibited completely the PAF-stimulated LTC(4) secretion to baseline level (P < 0.001). Inhibit ion of 5-lipoxygenase similarly blocked luminal narrowing caused by eo sinophils stimulated by PAF by > 95% (P < 0.001). We demonstrate that the binding of human eosinophils to the matrix protein FN causes augme nted secretion of LTC(4) which, in turn, causes augmented luminal narr owing of explanted human bronchi in vitro. We also demonstrate that th e augmented activity is blocked selectively by pretreatment with speci fic monoclonal antibody against VLA-4 and blockade of eosinophil 5-lip oxygenase inhibits both LTC(4) secretion and airway narrowing after PA F stimulation.