3D mechanically-loaded bone cell models for drug screening

Abstract: Osteoarthritis affects 8.5 million in the UK and cannot be treated. There is an urgent need for relevant human models to screen new drugs for osteoarthritis. Although abnormal joint loading is the major risk factor for osteoarthritis, the underlying mechanisms are poorly understood. We have developed a custom loading device where 3D bone models are housed in a multiwell flexible plate, and drugs screened under mechanical loading by measuring readouts of joint destruction, swelling and pain (Vazquez et al. 2014). We have compared the effects of an AMPA/kainate GluR antagonist, NBQX, in our bone model and animal models of osteoarthritis.

Mechanical loading of MLO-Y4 osteocytes embedded in type I collagen gels and overlayed with osteoblasts increased type I collagen mRNA and protein expression and reduced RANKL: OPG mRNA ratios, revealing an osteogenic response. NBQX treatment of unloaded co-cultures appeared to mimic these effects of mechanical loading.

Osteocytes differentiated from a human MSC cell line, responded to mechanical loading by reducing OPG protein release, and reducing protein expression of inflammatory mediators. Load-induced reduction in OPG release was inhibited by NBQX, consistent with its anti-anabolic effects in arthritis in vivo. NBQX also reduced osteocyte release of IL-6, IL-8, IP-10, MCP-1 and RANTES release, mimicking some of the load-induced effects on osteocytes.

To determine whether the high glutamate concentrations found in arthritic joints influenced responses to loading, we loaded 3D bone models in physiological and pathophysiological concentrations of glutamate. We have discovered that IL-6, OPG, RANKL and RANKL:OPG mRNA exression in osteocytes all respond to glutamate concentrations. High glutamate concentrations appeared to increase RANKL expression under pathophysiological loading, indicative of increased bone resorption.

Screening NBQX in our 3D bone model has revealed anti-anabolic and anti-inflammatory effects consistent with in vivo data (Bonnet et al. 2015). The interactions between mechanical load, and glutamate concentration is starting to reveal the mechanism of action of this drug.

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