Effects of Three-Dimensional Culture of Mouse Calvaria-Derived Osteoblastic Cells in a Collagen Gel with a Multichannel Structure on the Morphogenesis Behaviors of Engineered Bone Tissues

Bone has a complex hierarchical structure that contributes to its superior mechanical properties. Therefore, reproducing the complex hierarchical structure of bone tissue is a promising strategy to construct functional engineered bone tissues. In this study, we aimed to reproduce this complex hierarchical structure by developing a method for the three-dimensional culture of MC3T3-E1 osteoblastic cells in a collagen gel with a multichannel structure (MCCG), which mimics the parallel arrangement of Haversian canals in bone tissue. MCCG was homogeneously calcified via the biomineralization properties of MC3T3-E1s. Confocal laser scanning microscopy revealed that MCCG could support the growth and proliferation of MC3T3-E1 cells in the deeper parts of the engineered bone tissue and that the cells formed a toroidal structure on the channel surface and a network-like structure in the gel matrix region. Furthermore, quasi-quantitative measurement of osteocalcin and dentin matrix protein 1 expression indicated the coexistence of two types of cells with different morphologies and differentiation phenotypes. Thus, three-dimensional culture of MC3T3-E1 cells in MCCG yielded engineered tissues mimicking the hierarchical structures of bone tissues. Engineered bone tissues with a biomimetic hierarchical structure could be used as a model system for investigating bone metabolism and evaluating the efficacy of novel drugs