RGD Modification of Poly(2-oxazoline) Cryogels: Investigation of Material Properties and Cellular Adhesion

In this study, we demonstrate the preparation of a poly(2-oxazoline) based RGD-functionalized cryogels and cell culture studies on these cryogels. The present work also involves the investigation of cryogels with ethyl side chains (CG(B-Et-PipA)) and primary amino groups in their side chain (CG(B-Am-PipA)) which have been synthesized previously. CG(B-Am-PipA) was subsequently functionalized with a peptide containing an RGD motif (GCWGRGDSP), resulting in the formation of CG(B-Am-PipA/RGD). The coupling was verified by confocal laser-scanning microscopy (CLSM), fourier-transform infrared spectroscopy (FT-IR), high-resolution magic-angle-spinning (HR-MAS) NMR spectroscopy, and gravimetrical measurements. The degree of functionalization was found to be around 45%. The functionalized gels exhibited a change in their thermal properties, which was examined using thermogravimetric analysis (TGA). Rheological analysis was employed to study the mechanical properties and showed the formation of a stiffer material after peptide coupling. Swelling tests revealed a reduced swelling behavior for CG(B-Am-PipA/RGD). Cell biological investigations were conducted with L929 cells which were incubated with the samples CG(B-Et-PipA)RhoB, CG(B-Am-PipA)RhoB and CG(B-Am-PipA/RGD)RhoB. CLSM measurements after 0.5, 1, 2 and 6 h revealed an initial adhesion for both CG(B-Am-PipA)RhoB and CG(B-Am-PipA/RGD)RhoB, with the RGD-functionalized cryogel exhibiting the fastest cell adhesion and the most pronounced adherent phenotype, characterized by distinctly spread cell bodies as confirmed by cell area measurements. Cultivation of cells over a period of 7 days, analyzed by scanning electron microscopy (SEM), further showed pronounced cellular adhesion exclusively on CG(B-Am-PipA) and CG(B-Am-PipA/RGD).