Effect of culture duration on chondrogenic preconditioning of equine bone marrow mesenchymal stem cells in self-assembling peptide hydrogel.

Ex vivo induction of chondrogenesis is a promising approach to improve upon the use of bone marrow mesenchymal stem cells (MSCs) for cartilage tissue engineering. This study evaluated the potential to induce chondrogenesis with days of culture in chondrogenic medium for MSCs encapsulated in self-assembling peptide hydrogel. To simulate the transition from preconditioning culture to implantation, MSCs were isolated from self-assembling peptide hydrogel into an individual cell suspension. Commitment to chondrogenesis was evaluated by seeding preconditioned MSCs into agarose and culturing in the absence of the chondrogenic cytokine transforming growth factor beta (TGFβ). Positive controls consisted of undifferentiated MSCs seeded into agarose and cultured in medium containing TGFβ. Three days of preconditioning was sufficient to produce chondrogenic MSCs that accumulated ∼75% more cartilaginous extracellular matrix than positive controls by day 17. However, gene expression of type X collagen was ∼65-fold higher than positive controls, which was attributed to the absence of TGFβ. Potential induction of immunogenicity with preconditioning culture was indicated by expression of major histocompatibility complex class II (MHCII), which was nearly absence in undifferentiated MSCs, and ∼7% positive for preconditioned cells. These data demonstrate the potential to generate chondrogenic MSCs with days of self-assembling peptide hydrogel, and the ability to readily recover an individual cell suspension that is suited for injectable therapies. However, continued exposure to TGFβ may be necessary to prevent hypertrophy indicated by type X collagen expression, while immunogenicity may be a concern for allogeneic applications. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1368-1375, 2019.