Joris van Dongen

318 Chapter 13 Extracellular matrix derived hydrogels are non-cytotoxic The serial sampling at 24h intervals of possible cytotoxic elutes from hydrogels showed no difference in MTT conversion compared to medium controls. Therefore, no IC 50 could be calculated for all time points of each hydrogel indicating that the hydrogels did not release any cytotoxic compounds over time (Fig. 4). Results showed that the positive control (puromycin) induced a strong cytotoxic response (LogIC 50 3.1 – 3.6 mM). Figure 4. Optical density plotted against the serially diluted CMe from human adipose tissue-derived hydrogels after four different time points (log scale). PK84 fibroblasts were treated with hydrogel CMe or controls for 48h. Puromycin was used as positive control. Normal culture medium was used as negative control. Results are presented as mean with standard error of the mean of triplicates of three independent donors. CMe = conditioned medium. No dead cells present in extracellular matrix derived hydrogels Live/dead staining showed no dead cells (PI-stained nuclei) in hydrogels nor control (Fig. 5) ASCs appeared to adhere to the hydrogels and acquire the typical spindle-shape of mesenchymal cells; similar to the controls seeded on flat tissue culture plastic (Fig. 5B, D, F). It appeared that ASCs were more stretched when cultured in hydrogels than on tissue culture plastic (Fig. 5). Cell density did not appear to differ between ASC seeded on hydrogels as compared to tissue culture plastic controls, although quantification was not possible. The visualization even of fluorescently labelled ASCs (Fig. 5A, C, E) on and in hydrogels is challenging due to the limited depth of field with the microscope.