Designing novel hybrid hydrogel scaffolds for driving the fate of human cardiac MSC

The design of appropriate scaffolds remains one of the most important challenges for regenerative medicine. Current idea is that the interaction between stem cells and scaffold could drive cell fate by regulating gene expression and protein organization. In this study, the combination of biochemical and mechanical properties was optimized in order to produce novel 3D protein hydrogels able to favors cell-structure interactions, cell alignment and to promote differentiation of human cardiac mesenchymal stem cells (cMSCs). Firstly, novel injectable hybrid hydrogels (PSFHy) were produced by combining the bio-functionality of silk fibroin protein with the structural versatility of poly(ethylene glycol) diacrylate. Functionalization of the 3D-PSFHy with protein microspheres was required to increase the porosity and cell-adhesive properties of the material. MSembedding PSFHy were able to sustain a good viability of cMSCs – far beyond what reported for unfunctionalized hydrogels – and cMSCs exhibited the expression of proteins that are characteristic of the initial phases of cardiac muscle differentiation process. A cell-preconditioning of the scaffold was also performed, suggesting a potential application of these sponge-like hydrogels for analysing the effects of several extracellular microenvironments, produced by different kinds of cells, on the stem cells fate. In the context of 3D-bioprinting developments, novel 3D Scaffold-in-scaffold were designed revealing able to provide a multitextured 3D environment to the cells and to guide cMSCs alignment and promote their differentiation inducing the development of a cardiac phenotype. Finally, a novel formulation of diallyl disulfide (DADS) and αlinolenic acid (ALA) as protein nanoemulsions (BAD-NEs) was produced. BAD-NEs ability to release the bioactive molecule H2S, their antioxidant and radical scavenging property and their effects in stimulating cMSCs growth were demonstrated. Studies aimed at limiting hydrogel photopolymerization damage on embedded cells – among others in 3Dbioprinting technologies – were carried out and to this purpose cMSCs pretreatment with BAD-NEs was proven effective.