Airway remodelling in chronic severe asthma is associated with proliferation of airway wall smooth muscle and reticular basement membrane thickening due to increased extracellular matrix (ECM) protein deposition. We have characterised the effects of specific ECM proteins that are altered in the airways of asthmatics on airway smooth muscle (ASM) cell attachment (by Methylene Blue staining; Oliver et al. 1989) and on proliferation (MTT reduction assay; Hirst et al. 1992) induced by PDGF-BB (5 ng ml^-1). Additionally, the cell surface integrin repertoire was examined by immunocytochemistry, and its functional relevance determined using monoclonal anti-human blocking antibodies.

This study was approved by the Guy’s and St Thomas’ Hospitals’ Research Ethics Committee and written, informed consent was obtained. Growth-arrested human ASM cells (2 Ω 10⁴ per well; Hirst et al. 1992) were seeded into 24-well plates pre-coated with (1-30 µg ml^-1) collagen (Col) types I (fibrillar or monomeric), III, IV and V, biglycan (BN), decorin (DN), fibronectin (FN), laminin (LN) or tenascin-C (TN). Significant attachment occurred to the ECM substrates (P < 0.001, n = 4) with more than 70 % of cells on 10 µg ml^-1 Col I, III, IV, FN or LN being attached by 2 h; maximum attachment occurred at 8 h. Attachment to BN or DN was reduced (~50 %) compared with plastic (P > 0.001, n = 4, ANOVA) and other ECM substrates. PDGF-BB-stimulated proliferation was significantly (P < 0.001, n = 6) enhanced in cells growing on FN (EC₅₀ 10 µg ml^-1) or monomeric Col I (EC₅₀ 3 µg ml^-1), compared with growth on plastic. However, proliferation induced by PDGF-BB on Col V, LN, BN or DN was inhibited. Col I (fibrillar), III and IV, and TN, had little effect on PDGF-BB-stimulated proliferation. ASM cells expressed α2, α3, α5, αVβ3 and β1 integrins. Cellular attachment to ECM (Col I, III, IV and V, and TN) substrates was blocked (~50 %) by an anti-β1 integrin blocking antibody (10 µg ml^-1), while no inhibitory effect was seen with blocking antibodies (10 µg ml^-1) to individual α integrins (α1-6 or αV). When α integrin blocking antibodies were combined, attachment to Col I was inhibited (~40 %). This was reduced further by 10 µg ml^-1 of β1 but not β2 integrin blocking antibody (~85 %).

The data suggest that the ECM is important in regulating mitogen-stimulated proliferation of human ASM cells, and attachment to several ECM proteins is mediated partially by β1 integrins.This work was supported by The Wellcome Trust (051435) and the Special Trustee’s of Guy’s Hospital.

Hirst, S.J., Barnes, P.J. & Twort, C.H.C. (1992). Am. J. Resp. Cell Mol. Biol. 7, 574-581.

Oliver, H.M., Harrison, N.K., Bishop, J.E., Cole, P.J. & Laurent, G.J. (1989). J. Cell Sci. 92, 513-518.