Enhancing the stiffness of collagen hydrogels for delivery of encapsulated chondrocytes to articular lesions for cartilage regeneration.

This study investigated a dual crosslinking paradigm, consisting of (a) photocrosslinking with Rose Bengal (RB) and green light followed by (b) chemical crosslinking with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), and N-hydroxysuccinimide (NHS) to enhance collagen gel stiffness. In group 1, 50 μL collagen constructs of 2% (w/v) type I collagen containing 10 μM RB were allowed to gel spontaneously at 37 °C. In group 2, the spontaneous gels were exposed to green light (532 nm). In group 3, the photochemically crosslinked gels were subsequently treated with a 1-h exposure to 33 mM EDC/6 mM NHS. Samples (n = 18) were subjected to 0.08% (w/v) collagenase digestion, and the storage modulus of samples was measured by rheometry. Viability of encapsulated chondrocytes was measured by live/dead assay. Chondrocytes were ≥ 95% viable in all constructs at 10 days in vitro. Resistance to collagenase digestion increased as; spontaneous gels (2 h) < photochemical gels (3-4 h) < dual crosslinked gels (>24 h). The storage modulus of dual-crosslinked constructs was increased 5-fold over both photocrosslinked and spontaneous gels. As the dual crosslinking paradigm did not reduce encapsulated chondrocyte viability, these crosslinked collagen hydrogels could be a useful tool for the practical delivery of encapsulated chondrocytes to articular defects.