α5β1 integrin mediates the biologic activity of peptide 12. (A) Peptide 12 or BSA, as a nonspecific adhesion control, was coated onto microtiter plates, and HUVECs were added in the presence of chelating agents (EDTA or EGTA). Alternatively, HUVECs were preincubated with an antibody against α5β1 integrin, against α5 or β1 integrin subunits, or against α6 integrin subunit and allowed to adhere on coated wells. Data are mean plus or minus SEM; a representative experiment performed in triplicate is shown. (B) Fibronectin-adherent HUVECs were incubated with magnetic beads coated with RGE peptide (i), RGD peptide (ii,iv), or peptide 12 (iii,v). HUVECs were then immunostained for α5β1 integrin. Integrin clusters around the beads are indicated (arrowheads). Bars represent 10 μm.