Effects of photochemical riboflavin-mediated crosslinks on the physical properties of collagen constructs and fibrils

The use of collagen scaffold in tissue engineering is on the rise, as modifications to mechanical properties are becoming more effective in strengthening constructs whilst preserving the natural biocompatibility. The combined technique of plastic compression and cross-linking is known to increase the mechanical strength of the collagen construct. Here, a modified protocol for engineering these collagen constructs is used to bring together a plastic compression method, combined with controlled photochemical crosslinking using riboflavin as a photoinitiator. In order to ascertain the effects of the photochemical crosslinking approach and the impact of the crosslinks created upon the properties of the engineered collagen constructs, the constructs were characterized both at the macroscale and at the fibrillar level. The resulting constructs were found to have a 2.5 fold increase in their Young’s modulus, reaching a value of 650 ± 73 kPa when compared to non-crosslinked control collagen constructs. This value is not yet comparable to that of native tendon, but it proves that combining a crosslinking methodology to collagen tissue engineering may offer a new approach to create stronger, biomimetic constructs. A notable outcome of crosslinking collagen with riboflavin is the collagen’s greater affinity for water; it was demonstrated that riboflavin crosslinked collagen retains water for a longer period of time compared to non-cross-linked control samples. The affinity of the cross-linked collagen to water also resulted in an increase of individual collagen fibrils’ cross-sectional area as function of the crosslinking. These changes in water affinity and fibril morphology induced by the process of crosslinking could indicate that the crosslinked chains created during the photochemical crosslinking process may act as intermolecular hydrophilic nanosprings. These intermolecular nanosprings would be responsible for a change in the fibril morphology to accommodate variable volume of water within the fibril.