Molecularly designed alginate hydrogels susceptible to local proteolysis as three-dimensional cellular microenvironments

This work reports the development of new alginate-based hydrogels mimicking key features of the natural extracellular matrix, namely cell adhesion and cell-mediated matrix degradation. This work is part of the PhD works of Keila Fonseca.

Keila B. Fonseca, Sí­lvia J. Bidarra, Maria J. Oliveira, Pedro L. Granja and Cristina C. Barrias. Acta Biomaterialia, 2011.

The development of sophisticated three-dimensional (3-D) cell culture microenvironments that recreate some of the complexity of the natural extracellular matrix (ECM) remains a challenging task. Here, the modification of alginate through partial crosslinking with a matrix metalloproteinase (MMP) cleavable peptide (proline-valine-glycine-leucine-isoleucine-glycine, PVGLIG) is described, and its use in the preparation of injectable, in situ crosslinkable hydrogel-like matrices is proposed. PVGLIG-grafted algi- nates were synthesized by carbodiimide chemistry and characterized. Their biological performance was evaluated by comparing the response of 3-D cultured mesenchymal stem cells (MSCs) to alginate hydrogels containing only cell-adhesion peptides (RGD-alginate) or both peptides (PVGLIG/RGD-algi- nate). After 1 week, cells remained essentially round within RGD-alginate, while they exhibited an elon- gated morphology within PVGLIG/RGD-alginate hydrogels, forming cellular networks. This suggests that cells were able to structurally reorganize the matrix, through enzymatic hydrolysis of PVGLIG residues, overcoming biophysical hydrogel resistance. As shown by gelatine-zymography, MSC presented higher activity of MMP-2 when cultured within alginate functionalized with MMP-sensitive peptide, suggesting that the cell’s proteolytic phenotype was modulated by the matrix composition. Additionally, PVGLIG/ RGD-alginate hydrogels were clearly degraded in cell culture. Taken together, the results demonstrate that the co-incorporation of MMP-labile peptides in cell-adhesive RGD-alginate hydrogels improved their performance as ECM analogues, providing a more dynamic and physiological 3-D cellular microenvironment.

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