A collagen suspension (5 ml) containing 3.0 ml of collagen G1 (5 mg/ml; MATRIX BioScience, Mörlenbach, Germany), 0.5 ml of 10× M199 medium (Sigma), 85 μl of 1 N NaOH (Sigma-Aldrich), and sterile water was mixed with 1.0 ml (2 × 106 cells) of sh-ITGA5 or sh-NC hPSCs. Collagen gel–cell suspension (0.6 ml per well) was plated in a 24-well culture plate and allowed to polymerize for 1 hour at 37°C. Once polymerized, 1 ml of serum-free medium was added with or without TGF-β (5 ng/ml) followed by detachment of the gels from the culture wells. To study the effect of peptidomimetic AV3, 1 ml of serum-free medium with TGF-β (5 ng/ml) and 20 μM AV3 or sAV3 peptidomimetic was added to the detached gels. Representative images were made at either 72 or 96 hours using a digital camera (Nikon, Mississauga, ON, Canada). Measurement of collagen gel diameter was performed using ImageJ imaging software (NIH, Bethesda, MD).

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