2.5. Islets exposed to silk and alginate hydrogel fragments in vitro

Silk fibroin solutions were prepared as previously described[12]. Silk self-assembly and gelation was induced by sonication of 375 μL of 2 wt. % silk fibroin mixed with 625 μL of sterile water in a glass vial and vortexed for 7 min at 3200 rpm (VWR International, Radnor, PA). After phase separation, the white solid-like material was removed, and the remaining silk solution was diluted two-fold with media and allowed to gel for 2 h at 37°C. Subsequently, a sterile pipette tip was used to disperse the silk gel into fragments.

Alginic acid sodium salt from brown algae was obtained commercially (Sigma, Cream Ridge, NJ) and dissolved to 2% w/v in purified water. Alginate gel fragments were formed using 30 μL 2% w/v alginate dripped into 100 mM CaCl₂ to crosslink and create gel pellets. These gel pellets were incubated at 37°C for 1 h, placed on an 80 μm strainer, and washed with PBS to remove residual CaCl₂. Alginate pellets were then re-suspended in 600 μL medium and sonicated until broken up homogeneously in the medium. Islet samples (30 IEQs) were then plated with either silk or alginate fragments for 24 h (with medium alone serving as a control), followed by treatment with an inflammatory cytokine cocktail (10 ng/mL tumor necrosis factor (TNF)α, 5 ng/mL IL-1β, and 100 ng/mL IFNγ) for 4 h. Islet function was assessed by GSIS assay before and after cytokine exposure.