The traditional two-dimensional (2D) cell culture methods have a long history of mimicking in vivo cell growth. However, these methods cannot fully represent physiological conditions, which lack two major indexes of the in vivo environment; one is a three-dimensional 3D cell environment, and the other is mechanical stimulation; therefore, they are incapable of replicating the essential cellular communications between cell to cell, cell to the extracellular matrix, and cellular responses to dynamic mechanical stimulation in a physiological condition of body movement and blood flow. To solve these problems and challenges, 3D cell carriers have been gradually developed to provide a 3D matrix-like structure for cell attachment, proliferation, differentiation, and communication in static and dynamic culture conditions. 3D cell carriers in dynamic culture systems could primarily provide different mechanical stimulations which further mimic the real in vivo microenvironment. In this review, the current advances in 3D dynamic cell culture approaches have been introduced, with their advantages and disadvantages being discussed in comparison to traditional 2D cell culture in static conditions.

Accumulating evidence indicates that the interaction between immune and skeletal systems is vital in bone homeostasis. However, the detailed mechanisms between macrophage polarization and osteogenic differentiation of mesenchymal stromal cells (BMSCs) remain largely unknown. We observed enhanced macrophage infiltration along with bone formation in vivo, which showed a transition from early stage M1 phenotype to later stage M2 phenotype, cells at the transitional stage expressed both M1 and M2 markers actively participated in osteogenesis, which was mimicked by stimulating macrophages with lower inflammatory stimulus (as compared with typical M1). Using conditioned medium (CM) from M0, typical M1, low-inflammatory M1 (M1semi), and M2 macrophages, it was found that BMSCs treated with M1semi CM showed significantly induced migration, osteogenic differentiation, and mineralization, compared to others. Along with the induced osteogenesis, the autophagy level was the highest in M1semi CM treated BMSCs, which was responsible for BMSC migration and osteogenic differentiation, as autophagy-interruption significantly abolished this effect. This study indicated that low-inflammatory macrophage could activate autophagy in BMSCs to improve osteogenesis.