Tuning the resorption-formation balance in an in vitro 3D osteoblast-osteoclast co-culture model of bone

Stefan J A Remmers; Freek C van der Heijden; Bregje W M de Wildt; Keita Ito; Sandra Hofmann

doi:10.1016/j.bonr.2022.101646

Tuning the resorption-formation balance in an in vitro 3D osteoblast-osteoclast co-culture model of bone

Bone Rep. 2022 Dec 12:18:101646. doi: 10.1016/j.bonr.2022.101646. eCollection 2023 Jun.

Authors

Stefan J A Remmers¹, Freek C van der Heijden¹, Bregje W M de Wildt¹, Keita Ito¹, Sandra Hofmann¹

Affiliation

¹ Orthopaedic Biomechanics, Department of Biomedical Engineering and Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, the Netherlands.

Abstract

The aim of the present study was to further improve an in vitro 3D osteoblast (OB) - osteoclast (OC) co-culture model of bone by tuning it towards states of formation, resorption, and equilibrium for their future applications in fundamental research, drug development and personalized medicine. This was achieved by varying culture medium composition and monocyte seeding density, the two external parameters that affect cell behavior the most. Monocytes were seeded at two seeding densities onto 3D silk-fibroin constructs pre-mineralized by MSC-derived OBs and were co-cultured in one of three different media (OC stimulating, Neutral and OB stimulating medium) for three weeks. Histology showed mineralized matrix after co-culture and OC markers in the OC medium group. Scanning Electron Microscopy showed large OC-like cells in the OC medium group. Micro-computed tomography showed increased formation in the OB medium group, equilibrium in the Neutral medium group and resorption in the OC medium group. Culture supernatant samples showed high early tartrate resistant acid phosphatase (TRAP) release in the OC medium group, a later and lower release in the Neutral medium group, and almost no release in the OB medium group. Increased monocyte seeding density showed a less-than-proportional increase in TRAP release and resorption in OC medium, while it proportionally increased TRAP release in Neutral medium without affecting net resorption. The 3D OB-OC co-culture model was effectively used to show an excess of mineral deposition using OB medium, resorption using OC medium, or an equilibrium using Neutral medium. All three media applied to the model may have their own distinct applications in fundamental research, drug development, and personalized medicine.

Keywords: Cell culture; Co-culture; In vitro; Iβ3, integrin β3; M-CSF, macrophage colony-stimulating factor; MSC, mesenchymal stromal cell; OB, osteoblast; OC, osteoclast; Osteoblast; Osteoclast; RANKL, receptor activator of nuclear factor kappa-B ligand; Remodeling; Resorption; TRAP, tartrate resistant acid phosphatase; pNPP, (4-nitrophenyl) dihydrogen phosphate.