Collagen nanofiber-covered porous biodegradable carboxymethyl chitosan microcarriers for tissue engineering cartilage

Nanofibrous collagen-coated porous carboxymethyl chitosan microcarriers (CMC-MCs) were successfully fabricated for use as injectable cell microcarriers. A modified phase separation method combined with temperature controlled freeze-extraction was used for formulating the CMC-MCs. Collagen nanofibers were immobilized onto the surfaces of the CMC-MCs via covalently anchoring some collagen molecules first and more molecules self-assembling into nano-scale fibrous networks afterward. Scanning electron microscopy and hydroxyproline colorimetry analysis revealed that more collagen was immobilized on the CMC-MCs with collagen molecules anchored initially. In vitro cell culture revealed that chondrocytes could adhere, proliferate, and remain differentiated on the nanofiber-coated CMC-MCs. Optical microscopy and confocal laser scanning microscopy showed that chondrocytes grew to confluence on the CMC-MCs within 3 days post-seeding. Subsequently, several confluent CMC-MCs attached to each other, forming tissue-like aggregates after 7 days culture. The mRNA expression of type II collagen was much stronger in chondrocytes cultured on the nanofiber-coated CMC-MCs for 7 days than those cultured in 24-well plates or on CMC-MCs without initial treatment. These porous CMC-MCs could be utilized for cultivating cells and for application in cartilage tissue engineering as injectable scaffolds for cell delivery.