Monitoring bone remodeling in a humanized 3D osteoblast-osteoclast co-culture

Introduction: Changes in resorption and formation cannot be both monitored, localized and quantified in existing in vitro 3D models of bone remodelling, but addition of this through micro-computed tomography (μCT) would make such models more useful for studying bone remodelling. The goal of this study was to develop a 3D osteoblast-osteoclast co-culture in which 1) osteoblasts deposit mineralized matrix, 2) monocytes differentiate into resorbing osteoclasts, and 3) the formation and resorption of mineralized matrix could be localized and quantified over time using μCT.
Methods and results: Bone marrow derived mesenchymal stromal cells (MSCs) were seeded on silk fibroin scaffolds, succeeded to differentiate towards osteoblasts and created mineralized constructs (Fig. 1a). Thereafter, monocytes were added and differentiated towards osteoclasts during 3 weeks. Osteoclasts and resorption pits were visualized using Scanning Electron Microscopy (Fig 1b). The presence of osteoblasts, osteoclasts and osteocytes on the constructs was confirmed using immunohistochemistry (Fig. 1c). Constructs were scanned using μCT and subsequent scans were registered to visualize remodelling over time (Fig. 1d). μCT scans were used to quantify mineralized volume of the constructs over time (Fig. 1e), revealing that the mineralized volume of the constucts still increases after initiating a co-culture, but decreases after 12 d of co-culture. Registrations were used to determine that resorption exceeded formation between d12 and 22 of co-culture.
Discussion: Both osteoblastic formation and osteoclastic resorption were happening in parallel in the co-culture throughout the construct. As expected, mineralized volume continued to increase after the co-culture was initiated, as monocytes are not capable of resorption. After d 12 however, monocytes have differentiated into active osteoclasts, and resorption even exceeds formation.
Conclusion: the current model was able to visualize, localize and quantify mineralized matrix deposition by osteoblasts and resorption by osteoclasts. Such a model could be used to facilitate fundamental research on bone remodelling, facilitate drug testing and may have clinical implications in personalized medicine by directly testing drug effectiveness on patient cells.