FIGURE 2.

In vitro and in vivo viability of hASC. (A) MTS activity of hASC cultured in 2D. Cell viability was evaluated in 2D after molding Si-HPMC or Si-HPCH hydrogels on top of the cell layer (10,000 cells/cm²). As described in the section “Materials and Methods,” an MTS assay was performed on days 0, 1, and 7. The positive control (CTRL) was obtained by growing hASC alone, while the negative control was obtained by growing hASC in the presence of actinomycin D (Actino; 5 μg/mL). The results are expressed as the percentage of day 0 for each respective condition. ^∗∗∗p < 0.001 compared to day 0 (Kruskal-Wallis). (B,C) 3D viability of hASC cultured in Si-HPMC or Si-HPCH hydrogels. Cell viability was evaluated in 3D after molding hydrogels mixed with 1 × 10⁶ hASC on days 0, 1, and 7 by the Live/Dead Cell Viability assay. The negative control was obtained by adding actinomycin-D (Actino; 5 μg/mL) to the culture medium. Pictures of representative samples of hASC into Si-HPMC cultured in the presence of actinomycin-D (Actino), into Si-HPMC (Si-HPMC) or into Si-HPMC (Si-HPCH) at day 7 are shown (B). The scale bar represents 100 μm. Live and dead cells were then counted (C). The results are expressed as the percentage of day 0 for each respective condition. ^∗p < 0.05; ^∗∗∗p < 0.001 compared to day 0 (Kruskal-Wallis). (D) In vivo hASC viability after subcutaneous implantation. 250 μL of Si-HPMC (panels a and b) or Si-HPCH (panels c and d) were implanted alone (panels a and c) or mixed with human ASC (panels b and d; 1 × 10⁶ cells/mL of hydrogel) in the subcutis of nude mice. After 6 weeks, explanted samples were histologically prepared for Alcian Blue staining. The back arrows indicate hASC while the white arrows indicate chitosan. The scale bar represents 100 μm.