Adrenomedullin significantly and dose-dependently increases the adhesion of RA-FLSs to vitronectin, fibronectin, and type I collagen. RA-FLSs are resuspended in HAM-F12 without fetal calf serum and incubated for 1 hour with adrenomedullin (AM) at the indicated concentrations. Then 105 cells with or without adrenomedullin are seeded on 96-well plates previously coated overnight with the extracellular matrix proteins, bovine serum albumin (BSA), or polylysin. After 2-hour adhesion, nonadherent cells are removed, and the remaining RA-FLSs are stained with violet crystal for 15 minutes. The plates are washed, and the cells are lysed. Adherent cells are quantified by measuring absorbance at 550 nm with a spectrophotometer. (a) Effect of AM on RA-FLS adhesion to different surfaces. (b) The effect of AM is dose dependent. (c) Phase-contrast microscopy photographs demonstrating adhesion of RA-FLSs to BSA (negative control) and vitronectin with or without 10-7 M AM. (d) Time course of RA-FLSs adhesion to vitronectin under basal conditions (left panel) and with AM (right panel). Bars indicate mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.0001 versus control; ##P < 0.01 and ###P < 0.0001 versus 10-10 M AM; and, §§P < 0.01 and §§§P < 0.0001 versus 10-9 M AM. Results are given as the ratio of optical density (OD) in each condition over control OD without AM and reported as mean ± SEM of three experiments (three different donors) performed in quadruplicate.