Gelatin A (300 bloom grade, from porcine skin) and methacrylic anhydride were purchased from Sigma-Aldrich (USA). GelMA was synthesized as previously described60 (link). Briefly, a 10% w/v gelatin solution was prepared in PBS at pH 7.4. methacrylic anhydride (8 mL) was added dropwise to methacrylate amine groups along the gelatin backbones. The polymeric mixture was stirred vigorously and maintained at 60 °C for two hours after dilution with PBS. The polymeric solution was then transferred to dialysis membranes made of regenerated cellulose (~12–14 kDa cutoff) and dialyzed with deionized water for one week with two daily water changes. The GelMA solution was then frozen at −80 °C and lyophilized for 72 hours.
To fabricate GelMA-ND nanocomposite hydrogels, NDs were sonicated for 30 minutes (20 kHz, 2 sec on, 1 sec off) in deionized water at the concentration of 0.4% w/v prior addition to the GelMA solution in PBS. Nanocomposite hydrogels composition was modified varying the amount of NDs from 0.05% up to 0.2% w/v keeping constant gelatin concentration at 7% w/v and the amount of photoinitiator Irgacure 2959 (Sigma-Aldrich, USA) at 0.1% w/w. Photochemical hydrogels were then obtained after UV irradiation at 350–400 nm (Omnicure S200, Lumen Dynamics, Canada) for 6 minutes at an intensity of 7 mW/cm2.
To fabricate GelMA-ND nanocomposite hydrogels, NDs were sonicated for 30 minutes (20 kHz, 2 sec on, 1 sec off) in deionized water at the concentration of 0.4% w/v prior addition to the GelMA solution in PBS. Nanocomposite hydrogels composition was modified varying the amount of NDs from 0.05% up to 0.2% w/v keeping constant gelatin concentration at 7% w/v and the amount of photoinitiator Irgacure 2959 (Sigma-Aldrich, USA) at 0.1% w/w. Photochemical hydrogels were then obtained after UV irradiation at 350–400 nm (Omnicure S200, Lumen Dynamics, Canada) for 6 minutes at an intensity of 7 mW/cm2.