Mesenchymal stromal cell differentiation into osteoblasts on modular artificial proteins with bioactive ligands

Abstract

Modular self-assembling de novo triblock proteins (designated CRC) that contain a central bioactive domain with associating ends allowing preferential formation bundles have been developed. These proteins subsequently self assemble into hydrogel networks. The modularity of these triblock systems allows for different combinations of end and centre domains to modulate cell differentiation and phenotypic changes. In this study, we examined the effect of CRC peptides with bioactive ligands from extracellular matrix proteins involved in differentiation and repair activities, fibronectin (RGDS) and laminin (LQVQ), on mesenchymal stromal cell (MSC) differentiation into osteoblasts. Briefly, CRCs were covalently surface grafted onto collagen scaffolds via 1-ethyl-3(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) chemistry. Four peptide combinations tested were: P-CRC, P-CRC-RGDS/CRC, P-CRC-LQVQ/CRC and CRC-RGDS/CRC-LQVQ. They were examined for effects on proliferation and differentiation of MSCs in culture. No significant differences in proliferation rates were observed amongst samples. MSCs grown on CRC alone were unhealthy. All cells cultured on the peptide substrates, except for a combination of CRC-RGDS/CRC-LQVQ, had calcium deposits that were indicators of osteogenic differentiation. Cells grown on CRC/CRC-RGDS substrates showed most differentiation. The combination of RGDS/LQVQ peptides appeared to promote retention of an undifferentiated progenitor cell state. We have therefore shown that modular, self-assembling peptides have the potential to be useful in modulating differentiation or retention of the stem cell phenotype. With further development, they could possibly be used to develop or enhance biomaterials scaffolds for use in modulating stem cell behaviour in potential therapeutic applications.