The closest environment of stem cells, their extracellular matrix (ECM), is a complex micro-environment comprised of different proteins and carbohydrates such as glycosaminoglycans (GAGs). It dictates key signaling processes related with stem cells proliferation, differentiation and survival. Thus, mimicking the ECM in terms of composition, microstructure and mechanical properties is of great importance in order to understand these processes but also to develop regenerative strategies involving stem cells [1].
Herein, we report on the use of two GAGs, namely hyaluronan (HA) and chondroitin sulfate (CS) to create functional supports for stem cells culture. We have chosen HA and CS as they are part of the native ECM of the stem cells where they have structural and signaling role [2]. Both GAGs are negatively charged but differ by the sulfation degree: CS is sulfated and has bigger negative charge as compared to the non-sulfated HA. This structural difference determines different bioactivity of HA and CS, further explored under this study. The GAGs were modified by acrylation and then assembled in hydrogels by UV-photocrosslinkig. The use of GAGs-based hydrogels has many advantages such as high content of water, structural similarity to ECM, good biocompatibility and low toxicity. Soft lithography was used to introduce different micro-patterns, namely pillars and grooves, on the surface of hydrogels containing different compositions of HA:CS (Figure 1).
Adipose derived stem cells (ASCs) were cultured on these hydrogels and the effect of the chemical composition and the pattern on the ASCs behaviour (cell viability, morphology, adhesion and gene expression) was studied at different time points (7, 14 and 21 days).
We found that ASCs adhesion depends on the gel composition: the number of attached cells is proportional of the CS content of the gel. Microstructuring of the gel also induces different ASCs response: cells align on the grooved samples independently of surface composition, but not on the pillars micro-structured hydrogels(Figure 2). Noteworthy, our preliminary results demonstrate that high CS content together with the micro-pillar features induce differentiation towards the osteogenic lineage as demonstrated by the overexpression of Runx2, collagen type I and ALP genes (Figure 3).