Cell Ratio-Dependent Osteoblast-Endothelial Cell Crosstalk Promoting Osteogenesis-Angiogenesis Coupling via Regulation of Microfluidic Perfusion and Paracrine Signaling.

Osteogenesis-angiogenesis coupling, a dynamic and coordinated interaction between skeletal and vascular cells, is essential for fracture healing. However, the effects of these cell ratios and their interactions under microfluidic perfusion and paracrine signaling on osteogenesis-angiogenesis coupling have rarely been reported. In this study, dynamic and static models of osteogenesis-angiogenesis coupling were developed and the osteogenic and angiogenic effects of the two models were compared. Static co-cultures of MC3T3-E1 and bEnd.3 cells in Transwell inserts showed a cell ratio-dependent reciprocal relation: a ratio of 1:1 (MC3T3-E1:bEnd.3) favored osteogenesis, whereas a ratio of 2:1 (MC3T3-E1:bEnd.3) promoted angiogenesis. On that basis, we developed an osteogenesis-angiogenesis coupling chip based on microfluidic technology. The microfluidic perfusion within the chip further enhanced the mineralizing effect of osteoblasts and the angiogenic effect of endothelial cells, respectively, and increased the secretion of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) compared to the static Transwell insert model. The results suggest that the microfluidic chip enhanced the potential of osteogenesis-angiogenesis coupling mediated by paracrine signaling. Overall, the chip is not only a powerful model for understanding bone-vascular interaction but also a scalable platform for high-throughput drug screening and personalized therapy development for fractures.