Freezing adhesion molecules in a state of high-avidity binding blocks eosinophil migration.

Abstract

SummaryLeukocyte extravasation is mediated by multiple interactions of adhesive surface structures with ligands on endothelial cells and matrix components. The functional role of 31 (CD29) integrins (or very late antigen [VLA] proteins) in eosinophil migration across polycarbonate filters was examined under several in vitro conditions. Eosinophil migration induced by the chemoattractant C5a or platelet-activating factor was fully inhibited by monoclonal antibody (mAb) 8A2, a recently characterized "activating" CD29 mAb. However, inhibition by mAb 8A2 was observed only under filter conditions that best reflected the in vivo situation, i.e., when the eosinophils migrated over filters preincubated with the extracellular matrix (ECM) protein fibronectin (FN), or when the filters were covered with confluent monolayers of cultured human umbilical vein endothelial cells (HUVEC). When bare untreated filters were used, mAb 8A2 had no effect, whereas the C5a-directed movement was prevented by CD18 mAb. Studies with c~-subunit (CD49)-specific mAbs indicated that the integrins VLA-4 and -5 mediated migration across FN-preincubated falters, and VLA-2, -4, -5, and -6 were involved in eosinophil migration through filters covered with HUVEC. In contrast with the activating CD29 mAb 8A2, a combination of blocking CD49 mAbs or the nonactivating but blocking CD29 mAb AIIB2 failed to inhibit completely eosinophil migration over FN-preincubated or HUVEC-covered filters, mAb 8A2 stimulated binding to FN but not to HUVEC. Moreover, eosinophil migration over FN-preincubated or HUVEC-covered filters was significantly inhibited by anti-connecting segment 1 (CS-1) mAbs, as well as the sohble CS-1 peptide (unlike migration across bare untreated falters). Thus, inhibition of eosinophil migration by mAb 8A2 depended upon the presence of ECM proteins and not upon the presence of HUVEC per se. In conclusion, "freezing" adhesion receptors of the 31 integrin family into their high-avidity binding state by the activating CD29 mAb 8A2 results in a complete inhibition of eosinophil migration under physiological conditions. Hence, activation of 31 integrin-mediated cell adhesion may represent a new approach to prevent influx of inflammatory cells. though eosinophils constitute a minor fraction of the leukocytes in the circulation, they are a major component of the cellular infiltrate at extravascular sites of inflammation under certain pathological circumstances, such as allergic late phase responses in asthma and atopic skin reactions, parasitic infestation, and some delayed-type hypersensitivity reactions (1)(2)(3)(4)(5).At the beginning of this process of localized eosinophilic infiltration, the eosinophils leave the vascular compartment.The extracellular matrix (ECM) t of the vascular endothelial cells comprises all kinds of proteins and heavily glycosylated structures (proteoglycans) to which the infiltrating immune cells can attach during this migration along gradients of chemotactic factors. Platelet-activating factor (PAF) and the 1 Abbrevi...

IL-8 Induces a Transient Arrest of Rolling Eosinophils on Human Endothelial Cells

“…To investigate the changes in intracellular free Ca 2ϩ of adherent eosinophils upon IL-8 and eotaxin stimulation, cells were incubated with 8A2 and loaded on fibronectin-coated cover slips or nontreated cells were loaded on 7-h TNF-␣-stimulated HUVEC (see Materials and Methods (35)). After stimulation with 10 Ϫ8 M IL-8, a clear increase in [Ca 2ϩ ] i (Ͼ 200 nM) was observed in 42 Ϯ 7% and 30 Ϯ 7% of the cells adhering to fibronectin and activated HUVEC, respectively (Fig.…”

During human thymic development, β1 integrins regulate adhesion, motility, and the outcome of RHAMM/hyaluronan engagement

“…Inhibition of migratory behavior in vivo or of motility in vitro occurs on induction of a high-avidity integrin binding conformation [55,62]. For thymocytes, the in vivo regulatory events that control the transition from strong adhesion via ␣ 5 ␤ 1 , to weak adhesion via both ␣ 4 ␤ 1 and ␣ 5 ␤ 1 , and to motility via either ␣ 4 ␤ 1 or ␣ 5 ␤ 1 , are as yet unknown.…”

“…Activating mAbs to integrins increases adhesion [47,55,56,61]. Inhibition of migratory behavior in vivo or of motility in vitro occurs on induction of a high-avidity integrin binding conformation [55,62].…”

“…For mature T cells, integrins enter a high-avidity binding conformation only after activation. Adhesion-activating mAbs force integrins into a high-avidity Fn-binding state [47,[54][55][56]. To determine whether the alternate uses of ␤ 1 integrins in adhesion and motility reflected modulation of integrin conformation in vivo, thymocytes were pretreated with the activating mAb QE.2E5 [47], which induces high-avidity binding to Fn.…”

Expression and function of beta1 integrins on human eosinophils

“…Some studies have shown that resting eosinophils adhere to fibronectin in a VLA-4-dependent manner and exhibit prolonged survival via autocrine production of cytokines such as GM-CSF (Anwar et al 1994, Neeley et al 1994, Walsh et al 1995. Other studies, however, have found little or no adhesion without prior activation with platelet-activating factor, Mn ++ or a β1 integrin-activating antibody (Kuijpers et al 1993, Kita et al 1996. A possible explanation for these discrepancies may arise from the fact that eosinophils express the β2 integrin Mac-1, and engagement through this receptor to a different site on fibronectin, or to the blocking protein (typically albumin), may be occurring.…”