Riboflavin-UVA crosslinking of amniotic membranes and its influence on the culture of adipose-derived stem cells.

The human amniotic membrane (hAM) is a collagen-based extracellular matrix whose applications are restricted by its moderate mechanical properties and rapid biodegradation. In this work, we investigate the use of riboflavin, a water-soluble vitamin, to crosslink and strengthen the human amniotic membrane under UVA light. The effect of riboflavin-UVA crosslinking on hAM properties were determined via infrared spectroscopy, uniaxial tensile testing, proteolytic degradation, permeability testing, SEM, and quantification of free (un-crosslinked) amine groups. Samples crosslinked with glutaraldehyde, a common and effective yet cytotoxic crosslinking agent, were used as controls. Improved hAM mechanical properties must not come at the expense of reduced cellular proliferation and induction capabilities. In this study, we assessed the viability, proliferation, immunophenotype, and multilineage differentiation ability of human adipose-derived stem cells seeded on riboflavin-UVA crosslinked membranes. Overall, hAM crosslinked with riboflavin-UVA benefited from a stable three-fold increase in mechanical properties (comparable to the increase seen with glutaraldehyde crosslinked membranes) and improved biodegradation, all while retaining their biocompatibility and abilities to support the cultivation and differentiation of adipose-derived stem cells. Together, these results suggest that riboflavin-UVA crosslinking is an effective strategy to enhance the hAM for tissue engineering and regenerative medicine applications establishing it as an attractive and tuneable biomaterial.