Guangxi Collaborative Innovation Center for Biomedicine, Guangxi Medical University, Nanning 530021, China.
National Engineering Research Center for Biomaterials, Sichuan University, Chengdu 610064, China.
ACS Biomater Sci Eng. 2020 Jun 8;6(6):3491-3501. doi: 10.1021/acsbiomaterials.0c00149. Epub 2020 May 6.
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 , 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, , 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.
在天然软骨的发育过程中，间充质凝聚是软骨发生的起始事件，间充质干细胞（MSCs）经历了一个微环境的转变，从最初主要的细胞-细胞相互作用到后来细胞-细胞外基质（ECM）相互作用占主导地位。虽然微团培养已被开发用于模拟间充质凝聚，但分子机制仍不清楚，细胞-细胞到细胞-ECM 相互作用的转变也没有得到很好的再现。在本研究中，我们首先构建了 MSC 微球（MMs），并研究了它们的软骨分化，同时对 N-钙粘蛋白进行功能阻断。结果表明，与对照组相比，实验组 MSC 的早期软骨分化和软骨特异性基质沉积明显延迟。接下来，我们用聚赖氨酸（poly(l-lysine)）短暂处理以促进 N-钙粘蛋白基因的表达，结果促进了 MSC 的软骨分化。在建立细胞-细胞黏附的 MMs 中培养一天后，构建了胶原水凝胶包被的 MMs（CMMs），以模拟细胞-ECM 相互作用，胶原微环境补偿了 N-钙粘蛋白阻断的抑制作用。令人惊讶的是，在没有 N-钙粘蛋白阻断的情况下，我们发现了具有重要意义的软骨分化细胞的迁移。总之，我们的研究表明 N-钙粘蛋白在早期间充质凝聚中起着关键作用，胶原水凝胶为晚期软骨分化提供了一个支持性的微环境。因此，在工程化微环境中呈现出细胞-细胞黏附与细胞-ECM 相互作用的顺序似乎是促进 MSC 软骨分化的一种有前途的策略。
