Cytotoxicity of redox radical initiators for encapsulation of mesenchymal stem cells

We have developed a novel in situ crosslinkable hydrogel system based on oligo(poly(ethylene glycol) fumarate) that is being investigated as an injectable carrier for mesenchymal stem cells (MSCs) for orthopaedic tissue engineering applications. This hydrogel is crosslinked using the redox radical initiators ammonium persulfate and ascorbic acid. However, it is believed that, when oxidized, ascorbic acid produces a radical anion that decreases the pH of the solution during crosslinking. Therefore, in this study, combinations of two different persulfate oxidizing agents with three reducing agents derived from ascorbic acid were examined to determine the relationship between pH and cytotoxicity for rat MSCs. pH was recorded over two hours for solutions of the various reagents in cell culture media. After two hours of exposure, viability of MSCs was determined on a plate reader using the LIVE/DEAD fluorescent assay. pH and cell viability data for samples from 1-1000 mM showed that there was a smaller change in pH and a corresponding higher viability at lower concentrations, regardless of the reagent used. This indicates that low pH, even without the generation of radicals, has a negative impact on cell viability. Further results from combinations of oxidizing and reducing agents support the hypothesis that radical anion formation contributes to the pH drop in these solutions.