Fonseca et. al. Enzymatic, physicochemical and biological properties of MMP-sensitive alginate hydro

Fonseca et. al. Enzymatic, physicochemical and biological properties of MMP-sensitive alginate hydro

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Enzymatic, physicochemical and biological properties of MMP-sensitive alginate hydrogels Keila B. Fonseca, ab F. Raquel Maia, ab Filipe A. Cruz, ab Douglas Andrade, c Maria A. Juliano, c Pedro L. Granja abd and Cristina C. Barrias * a Protease-sensitive hydrogels that recapitulate the mechanisms of cell-driven enzymatic remodelling of the natural extracellular matrix (ECM) have been gaining popularity as arti f cial 3D cell-microenvironments. Here, the matrix metalloproteinase (MMP)-sensitive peptide Pro-Val-Gly-Leu-Iso-Gly (PVGLIG) was double-end grafted to alginate forming water-soluble PVGLIG alginate conjugates. The PVGLIG peptide was synthesized as a Fluorescence Resonance Energy Transfer (FRET) sensor and showed to be a good substrate for MMP-2, MMP-9, MMP-13 and MMP-14. After demonstrating that human MSC (hMSC) expressed both MMP-2 and MMP-14 under basal and osteogenic in vitro conditions, the e F ect of 3D- culture within MMP-sensitive alginate hydrogels on hMSC behaviour was addressed. In situ -forming alginate hydrogels containing only cell-adhesive RGD peptides (RGD alginate, MMP-insensitive) or both peptides (PVGLIG/RGD alginate, MMP-sensitive) were used. Cell matrix and cell cell interactions were enhanced in hMSC-laden MMP-sensitive alginate hydrogels, as compared to MMP-insensitive hydrogels with identical viscoelastic and microstructural properties. hMSC underwent osteogenic di F erentiation in both types of matrices. However, the presence of PVGLIG stimulated the secretion of proteases (most likely MMP-2) by hMSC, in both undi F erentiated and di F erentiated cultures. By using the FRET sensor, it was possible to demonstrate that the cocktail of hMSC-secreted MMPs was e F ectively active in cleaving the PVGLIG motif. Protease-sensitive alginates can be used to create cell-responsive 3D microenvironments and o F er promise as injectable carriers for therapeutic hMSC-delivery. 1 Introduction In situ gel-forming hydrogels, such as alginate, o f er promise not only as 3D models for in vitro mechanistic studies but also as injectable vehicles for the therapeutic delivery of cells and/or biomolecules. 1 4 Alginate hydrogels can be formed under cyto- compatible conditions, giving rise to highly hydrated and permeable 3D networks with tuneable viscoelastic properties. 5 8 Alginates are intrinsically non-fouling, lack cell-adhesion signals and cannot be cleaved by mammalian enzymes, but possess a number of reactive functional groups that can be chemically derivatized to enhance their biological properties. 6,9 This, in turn, makes alginates excellent blank state polymers for engineering biofunctional derivatives, namely through the incorporation of cell-signalling moieties. The incorporation of ECM-like peptides on the polymer backbone is among the strategies most used to promote cell interactions with hydrogels.
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Fonseca et. al. Enzymatic, physicochemical and biological properties of MMP-sensitive alginate hydro

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