Surface-Functionalized Poly(2-Oxazoline)-Based Grids Produced by Two-Photon Polymerization as 3D-Structured Hydrogels Promoting Cell Adhesion.

We present the development of resins for two-photon polymerization (2PP) printing based on sidechain-functionalized poly(2-oxazoline) (POx) cross-linkers. These POx cross-linkers carry piperazine acrylamide end groups (B-Et-PipA) and amino side-chain-functionalized POx (B-Am-PipA). They were converted by 2PP into 3D hydrogel grids, designated as P(B-Et-PipA) and P(B-Am-PipA), respectively. The morphology of the obtained structures was analyzed using scanning electron microscopy (SEM), confocal laser-scanning microscopy (CLSM), and atomic force microscopy. P(B-Am-PipA) based hydrogel grids were covalently functionalized with the FITC-labeled GCWRGDSP (RGD) peptide containing the RGD unit, resulting in P(B-Am-PipA/FITC-RGD). To investigate the hydrogels' suitability to be applied as potential cell culture matrices, grids of the samples P(B-Et-PipA), P(B-Am-PipA), and P(B-Am-PipA/RGD) were prepared possessing different mesh sizes (10 and 25 µm). The initial cellular responses of L929 cells cultured on the different hydrogels were monitored using fluorescence microscopy and SEM. On P(B-Et-PipA) hydrogels, the cells exhibited no apparent interaction with the hydrogel scaffold. In contrast, P(B-Am-PipA) and P(B-Am-PipA/RGD) surfaces promoted cellular attachment and progressive adhesion, resulting in flattened cell morphologies. Among these, P(B-Am-PipA/RGD) grids with a mesh size of 25 µm demonstrated the most rapid and pronounced cell adhesion, indicating that RGD functionalization is particularly effective in enhancing initial cell adhesion.