Toxicity and photosensitizing assessment of gelatin methacryloyl-based hydrogels photoinitiated with lithium phenyl-2,4,6-trimethylbenzoylphosphinate in human primary renal proximal tubule epithelial cells

Gelatin methacryloyl (GelMA) and lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) photoinitiator are commonly used in combination to produce a photosensitive polymer but there are concerns that must be addressed: the presence of unreacted monomer is well known to be cytotoxic, and lithium salts are known to cause acute kidney injury. In this study, acellular 10% GelMA hydrogels cross-linked with different LAP concentrations and cross-linking illumination times were evaluated for their cytotoxicity, photosensitizing potential, and elastic moduli. Alamar Blue and CyQuant Direct Cell viability assays were performed on human primary renal proximal tubule epithelial cells (hRPTECs) exposed to extracts of each formulation. UV exposure during cross-linking was not found to affect extract cytotoxicity in either assay. LAP concentration did not affect extract cytotoxicity as determined by the Alamar Blue assay but reduced hRPTEC viability in the CyQuant Direct cell assay. Photocatalytic activity of formulation extracts toward NADH oxidation was used as a screening method for photosensitizing potential; longer UV exposure durations yielded extracts with less photocatalytic activity. Finally, elastic moduli determined using nanoindentation was found to plateau to approximately 20-25 kPa after exposure to 342 mJ/cm² at 2.87 mW of UV-A exposure regardless of LAP concentration. LAP at concentrations commonly used in bioprinting (<0.5% w/w) was not found to be cytotoxic although the differences in cytotoxicity evaluation determined from the two viability assays imply cell membrane damage and should be investigated further. Complete cross-linking of all formulations decreased photocatalytic activity while maintaining predictable final elastic moduli.