Figure 3

Tol-OP-ST surface density influences cell morphology and response to BMP2. Gold coverslips were incubated with different concentrations of Tol-OP-ST and based on SPR analysis surface densities calculated (molecules/mm²). (a, b, c) Cells were plated onto the surfaces, as indicated, under serum free conditions and vinculin immunolocalized 24 hours later. Size bars = 50 μm. (d) DAPI was used to visualize nuclear DNA and images quantified to show number of adhered cells. Numbers of cells on 1.6 and 3.3 × 10¹⁰ molecules/mm² were significantly different to all other surfaces. (e) Assessment of cell area revealed that cells on 1.6 and 3.3 × 10¹⁰ molecules/mm² were significantly different to all other surfaces. Furthermore (f) levels of vinculin positivity were significantly higher in cells on 1.6 and 3.3 × 10¹⁰ molecules/mm² in comparison to lower densities. (g) Cells transfected with a SMAD-reporter construct were plated on the surfaces, treated with 100 ng/ml recombinant BMP-2 and levels of luciferase assessed and normalised to cell number. Data representative of three independent experiments; results are mean ± SEM; * P < 0.05, ** P < 0.01. Significance was determined by ANOVA with Fisher post test.