Morphological and physiological changes of osteocytes during bone formation in Bone-on-Chip

Abstract: Osteocytes are long-term living cells responsible for bone mineralization and characterized by a body bearing numerous processes. Osteocytes orchestrate bone homeostasis in detecting microdamage and coordinate the interactions between the osteoclasts and mesenchymal stem cells (MSCs) that will differentiate into osteoblasts to form bone during remodeling. Local micro-displacements are assumed to have positive cues to prime bone cells to repair bone and to have an influence on the cell differentiation and the quality of the formed bone. However fundamental knowledge of human bone stromal cells in situ mechanobiology is challenging to quantify in particular as the mechanical properties of their environment changes as mineralization takes place.

A bones-on-chip based on human bone and human MSCs offer the opportunity to develop in situ 3D imaging and numerical modelling of concurrent measurements of mechanobiological phenomena in a controlled microenvironment for long-term culture up to 26 months [1]. The cell mechanical microenvironment can be quantified in a numerical model based on the real geometry while the cell chemical response can be observed in vitro under confocal microscopy. The method can be applied to either native osteocytes and stem cell derived osteocytes in either native or neo-formed ECM. The cell morphology, expressed genes, and their secretome can be monitored using PCR, immunohistochemistry, in situ immunofluorescence.

The cytoplasmic calcium concentration variations seemed to adapt to the expected in vivo mechanical stimulation at the successive stages of cell differentiation. The bone-on-chip produced an ECM of which the strength was nearly a quarter of native bone at 109 days and that contained type I collagen at 256 days and calcium minerals at 39 days [1]. The cells were organized in network connected by processes and the newly-formed matrix characterized by in situ immuno-fluorescence showed the presence of E11 and sclerostin at 547 days.

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