Role of N-Cadherin in a Niche-Mimicking Microenvironment for Chondrogenesis of Mesenchymal Stem Cells In Vitro.

During the development of natural cartilage, mesenchymal condensation is the starting event of chondrogenesis, and mesenchymal stem cells (MSCs) experienced a microenvironment transition from primarily cell-cell interactions to a later stage, where cell-extracellular matrix (ECM) interactions dominate. Although micromass pellet culture has been developed to mimic mesenchymal condensation in vitro, the molecular mechanism remains elusive, and the transition from cell-cell to cell-ECM interactions has been poorly recapitulated. In this study, we first constructed MSC microspheres (MMs) and investigated their chondrogenic differentiation with functional blocking of N-cadherin. The results showed that early cartilage differentiation and cartilage-specific matrix deposition of MSCs in the group with the N-cadherin antibody were significantly postponed. Next, poly(l-lysine) treatment was transiently applied to promote the expression of N-cadherin gene, CDH2, and the treatment-promoted MSC chondrogenesis. Upon one-day culture in MMs with established cell-cell adhesions, collagen hydrogel-encapsulated MMs (CMMs) were constructed to simulate the cell-ECM interactions, and the collagen microenvironment compensated the inhibitory effects from N-cadherin blocking. Surprisingly, chondrogenic-differentiated cell migration, which has important implications in cartilage repair and integration, was found in the CMMs without N-cadherin blocking. In conclusion, our study demonstrated that N-cadherin plays the critical role in early mesenchymal condensation, and the collagen hydrogel provides a supportive microenvironment for late chondrogenic differentiation. Therefore, sequential presentations of cell-cell adhesion and cell-ECM interaction in an engineered microenvironment seem to be a promising strategy to facilitate MSC chondrogenic differentiation.