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

## TL;DR

This study shows how RGD peptides can be added to a special type of gel to improve cell adhesion and change material properties.

## Contribution

The novel contribution is the functionalization of poly(2-oxazoline) cryogels with RGD peptides and the demonstration of enhanced cell adhesion.

## Key findings

- RGD-functionalized cryogels showed faster and stronger cell adhesion compared to non-functionalized gels.
- Functionalization altered thermal, mechanical, and swelling properties of the cryogels.
- Cell spreading was observed only on RGD-functionalized and amino side chain gels, not on ethyl side chain gels.

## Abstract

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).

Poly(2‐oxazoline) based cryogels with amino side chains and piperazine derived end groups (CG(B‐Am‐PipA)) which were functionalized with an RGD peptide (CG(B‐Am‐PipA/RGD)) demonstrated changes in their thermal, mechanical, and swelling properties upon functionalization. Cultivation of L929 cells on the cryogels showed that both CG(B‐Am‐PipA) and CG(B‐Am‐PipA/RGD) facilitate cellular adhesion and spreading, a phenomenon which was not observed for the pure poly(2‐ethyl‐2‐oxazoline) based gel (CG(B‐Et‐PipA)).