A hydrogel system based on oxidized alginate covalently crosslinked with gelatin (ADA-GEL) has been utilized for different biofabrication approaches to design constructs, in which cell growth, proliferation and migration have been observed. However, cell–bioink interactions are not completely understood and the potential effects of free aldehyde groups on the living cells have not been investigated. In this study, alginate, ADA and ADA-GEL were characterized via FTIR and NMR, and their effect on cell viability was investigated. In the tested cell lines, there was a concentration-dependent effect of oxidation degree on cell viability, with the strongest cytotoxicity observed after 72 h of culture. Subsequently, primary human cells, namely fibroblasts and endothelial cells (ECs) were grown in ADA and ADA-GEL hydrogels to investigate the molecular effects of oxidized material. In ADA, an extremely strong ROS generation resulting in a rapid depletion of cellular thiols was observed in ECs, leading to rapid necrotic cell death. In contrast, less pronounced cytotoxic effects of ADA were noted on human fibroblasts. Human fibroblasts had higher cellular thiol content than primary ECs and entered apoptosis under strong oxidative stress. The presence of gelatin in the hydrogel improved the primary cell survival, likely by reducing the oxidative stress via binding to the CHO groups. Consequently, ADA-GEL was better tolerated than ADA alone. Fibroblasts were able to survive the oxidative stress in ADA-GEL and re-entered the proliferative phase. To the best of our knowledge, this is the first report that shows in detail the relationship between oxidative stress-induced intracellular processes and alginate di-aldehyde-based bioinks.

A novel approach, in the context of bioprinting, is the targeted printing of a defined number of cells at desired positions in predefined locations, which thereby opens up new perspectives for life science engineering. One major challenge in this application is to realize the targeted printing of cells onto a gel substrate with high cell survival rates in advanced bioinks. For this purpose, different alginate-dialdehyde—polyethylene glycol (ADA-PEG) inks with different PEG modifications and chain lengths (1–8 kDa) were characterized to evaluate their application as bioinks for drop on demand (DoD) printing. The biochemical properties of the inks, printing process, NIH/3T3 fibroblast cell distribution within a droplet and shear forces during printing were analyzed. Finally, different hydrogels were evaluated as a printing substrate. By analysing different PEG chain lengths with covalently crosslinked and non-crosslinked ADA-PEG inks, it was shown that the influence of Schiff’s bases on the viscosity of the corresponding materials is very low. Furthermore, it was shown that longer polymer chains resulted in less stable hydrogels, leading to fast degradation rates. Several bioinks highly exhibit biocompatibility, while the calculated nozzle shear stress increased from approx. 1.3 and 2.3 kPa. Moreover, we determined the number of cells for printed droplets depending on the initial cell concentration, which is crucially needed for targeted cell printing approaches.