Human Stem Cell Derived Osteocytes in Bone-on-Chip

Abstract

Human mesenchymal stem cells were reseeded in decellularized human bone subject to a controlled mechanical loading to create a bone-on-chip that was cultured for over 26 months. The cell morphology and their secretome were characterized using immunohistochemistry and in situ immunofluorescence under confocal microscopy. The presence of stem cell derived osteocytes was confirmed at 547 days. Different cell populations were identified. Some cells were connected by long processes and formed a network. Comparison of the MSCs in vitro reorganization and calcium response to in situ mechanical stimulation were compared to MLOY4 cells reseeded on human bone. The bone-on-chip produced an ECM of which the strength was nearly a quarter of native bone after 109 days and that contained calcium minerals at 39 days and type I collagen at 256 days. The cytoplasmic calcium concentration variations seemed to adapt to the expected in vivo mechanical load at the successive stages of cell differentiation in agreement with studies using fluid shear flow stimulation. Some degree of bone-like formation over a long period of time with the formation of a newly formed matrix was observed.

Chips for Biomaterials and Biomaterials for Chips: Recent Advances at the Interface between Microfabrication and Biomaterials Research

“…Similarly, in bone‐on‐chip studies, relatively soft hydrogels are a poor mimic of native bone matrix. Therefore, different biomaterials have been used to mimic the hard, mineralized ECM of bone, including fibrin incorporating hydroxyapatite, [ 237 ] mineralized collagen, [ 227 ] and tightly packed calcium phosphate microbeads, [ 194 ] as well as decellularized human [ 238 ] or animal [ 239 ] bone. These models have been used to investigate cancer metastasis into bone [ 227,239 ] or bone angiogenesis, [ 193 ] to investigate the mechanotransduction of osteocytes, [ 194 ] and to study the effect of mechanical stimulation on bone formation over timescales of multiple years.…”

“…These models have been used to investigate cancer metastasis into bone [ 227,239 ] or bone angiogenesis, [ 193 ] to investigate the mechanotransduction of osteocytes, [ 194 ] and to study the effect of mechanical stimulation on bone formation over timescales of multiple years. [ 238 ]…”

Towards More Predictive, Physiological and Animal-free In Vitro Models: Advances in Cell and Tissue Culture 2020 Conference Proceedings

“…These systems displayed multi-level spatial reorganisation of the cells, and the collagen fibres. 29 Hence, this bone-on-chip system makes it possible to simultaneously study the effects of mechanical stimulations and age on bone cell differentiation, their calcium mechanobiology and the quality of the bone they form over very long periods of time (at least over two years).…”

Osteoblast-Induced Collagen Alignment in a 3Din vitroBone Model

“…In recent years, multiple 3D in vitro cell culture models for bone have been developed to study variable aspects of bone development 7, 8 9 10 , which significantly improved the physiological relevance of bone cell cultures compared to previously used monolayer systems. These studies showed that mechanical stimulation 9,10 and matrix stiffness 7 are critical influencers of bone formation with regard to differentiation and bone matrix maturation. Remarkably, some of these 3D models showed that inducing longitudinal strain by fixation of a 3D cell culture on opposite sides was enough to induce matrix alignment and aligned mineralization.…”

“…In recent years, multiple 3D in vitro cell culture models for bone have been developed to study variable aspects of bone development (Akiva et al, 2021; Budyn et al, 2018; Nasello et al, 2020; Thrivikraman et al, 2019), which significantly improved the physiological relevance of bone cell cultures compared to previously used monolayer systems. These studies showed that mechanical stimulation (Akiva et al, 2021; Budyn et al, 2018) and matrix stiffness (Thrivikraman et al, 2019) are critical influencers of bone formation with regard to differentiation and bone matrix maturation. Remarkably, some of these 3D models showed that inducing longitudinal strain by fixation of a 3D cell culture on opposite sides was enough to induce matrix alignment and aligned mineralization (Iordachescu et al, 2018; Sasaki et al, 2010; Sasaki et al, 2015).…”