Osteogenic Response of MC3T3-E1 and Raw264.7 in the 3D-Encapsulated Co-Culture Environment



Three-dimensional (3D) in vitro cultures recapitulate the physiological microenvironment and exhibit high concordance with in vivo conditions. Improving co-culture models with different kind of cell types cultured on a 3D scaffold can closely mimic the in vivo environment. In this study, we examined the osteogenic response of pre-osteoblast MC3T3-E1 cells and Raw264.7 mouse monocytes in a 3D-encapsulated co-culture environment composed of the Cellrix® 3D culture system, which provides a physiologically relevant environment.


The Cellrix® 3D Bio-Gel scaffolds were used to individually culture or co-culture two type cells in 3D microenvironment. Under 3D culture conditions, osteoblastic behavior was evaluated with an ALP assay and staining. ACP assay and TRAP staining were used as osteoclastic behavior indicator.


Treatment with osteoblastic induction factors (+3F) and RANKL had on positively effect on alkaline phosphatase activity but significantly inhibited to acid phosphatase activity during osteoclastic differentiation in 3D co-culture. Interestingly, alkaline phosphatase activity or acid phosphatase activity in 3D co-culture was stimulated with opposite differentiation factors at an early stage of differentiation. We guess that these effects may be related to RANK–RANKL signaling, which is important in osteoblast regulation of osteoclasts.


In this study, the osteogenic response of 3D encapsulated pre-osteoblast MC3T3-E1 cells and mouse monocyte Raw264.7 cells was successfully demonstrated. Our 3D culture conditions will be able to provide a foundation for developing a high-throughput in vitro bone model to study the effects of various drugs and other agents on molecular pathways.

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This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education, Science and Technology (NRF-2018R1D1A1B07040482) and by Grant No. 03-2016-0190 from the SNUH Research Fund.

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Correspondence to Jae Hyup Lee or Misun Cha.

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Kim, J., Lyu, HZ., Jung, C. et al. Osteogenic Response of MC3T3-E1 and Raw264.7 in the 3D-Encapsulated Co-Culture Environment. Tissue Eng Regen Med (2021). https://doi.org/10.1007/s13770-020-00321-0

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  • 3D culture system
  • Co-culture
  • Encapsulation
  • Osteoblast
  • Osteoclast