THE ROLE OF INTEGRINS ALPHA-2-BETA-1 AND ALPHA-3-BETA-1 IN CELL CELL AND CELL SUBSTRATE ADHESION OF HUMAN EPIDERMAL-CELLS

We have examined cultures of neonatal human foreskin keratinocytes (HFKs) to determine the ligands and functions of integrins α2β1, and α3β1 in normal epidermal stratification and adhesion to the basement membrane zone (BMZ) in skin. We used three assay systems, HFK adhesion to purified extracellular matrix (ECM) ligands and endogenous secreted ECM, localization of integrins in focal adhesions (FAs), and inhibition of HFK adhesion with mAbs to conclude: (a) A new anti-α3β1 mAb, P1F2, localized α3β1 in FAs on purified laminin > fibronectin/collagen, indicating that laminin was the best exogenous ligand for α3β1. However, in long term culture, α3β1 preferentially codistributed in and around FAs with secreted laminin-containing ECM, in preference to exogenous laminin. Anti-α3β1, mAb P1B5, detached prolonged cultures of HFKs from culture plates or from partially purified HFK ECM indicating that interaction of α3β1 with the secreted laminin-containing ECM was primarily responsible for HFK adhesion in long term culture. (b) In FA assays, α2β1 localized in FAs coincident with initial HFK adhesion to exogenous collagen, but not laminin or fibronectin. However, in inhibition assays, anti-α2β1 inhibited initial HFK adhesion to both laminin and collagen. Thus, α2β1 contributes to initial HFK adhesion to laminin but α3β1 is primarily responsible for long-term HFK adhesion to secreted laminin-containing ECM. (c) Serum or Ca2+-induced aggregation of HFKs resulted in relocation of α2β1 and α3β1 from FAs to cell-cell contacts. Further, cell-cell adhesion was inhibited by anti-α3β1 (P1B5) and a new anti-β1 mAb (P4C10). Thus, interaction of α3β1 with either ECM or membrane coreceptors at cell-cell contacts may facilitate Ca2+-induced HFK aggregation. (d) It is suggested that interaction of α3β1 with a secreted, laminin-containing ECM in cultured HFKs, duplicates the role of α3β1 in basal cell adhesion to the BMZ in skin. Further, relocation of α2β1 and α3β1 to cell-cell contacts may result in detachment of cells from the BMZ and increased cell-cell adhesion in the suprabasal cells contributing to stratification of the skin.