Abstract
Ex vivo explant culture models are powerful tools in bone research. They allow investigation of bone and cartilage responses to specific stimuli in a controlled manner that closely mimics the in vivo processes. Because of limitations in obtaining healthy human bone samples the explant growth of animal tissue serves as a platform to study the complex physico-chemical properties of the bone. Moreover, these models enable preserving important cell–cell and cell–matrix interactions in order to better understand the behaviour of cells in their natural three-dimensional environment. Thus, the use of bone ex vivo explant cultures can frequently be of more physiological relevance than the use of two-dimensional primary cells grown in vitro. Here, we describe isolation and ex vivo growth of different animal bone explant models including metatarsals, femoral heads, calvaria, mandibular slices and trabecular cores. We also describe how these explants are utilised to study bone development, cartilage and bone metabolism, cancer-induced bone diseases, stem cell-driven bone repair and mechanoadaptation. These techniques can be directly used to understand mechanisms linked with bone physiology or bone-associated diseases.
Highlights
The bone is a complex and specialised connective tissue in a state of dynamic equilibrium that, together with cartilage, forms the skeletal system
MEMa medium with 10% fetal bovine serum (FBS) and antibiotics (100 mg ml À 1 streptomycin, 100 U ml À 1 penicillin); PBS; 0.05 and 0.25% trypsin-ethylenediaminetetraacetic acid disodium salt (EDTA); WD-40 (San Diego, CA, USA); LIVE/DEAD Viability/Cytotoxicity kit (Invitrogen, Waltham, MA, USA); equipment: diamond-tipped coring tool, 7 mm diameter (Starlite Industries, Rosemont, PA, USA); 60 ml sterile syringe and needles (18 and 30 gauge); Roccal-D Plus detergent (Zoetis); custom cell seeder, small magnetic stirrer; large pronged forceps, scalpel and blades; hand drill with locking bit and key; small vise; Interplak water jet (Conair, Stamford, CT, USA); IsoMet low-speed saw with high carbon (HC) wafering blade (Buehler, Lake Bluff, IL, USA); magnetic stir plate; mechanical testing device such as Bose Electroforce (Bose, Eden Prairie, MN, USA)
Ex vivo growth of metatarsal and femoral head explants are grown in the absence of serum, which eliminates the need of serum batch testing and improves reproducibility
Summary
Silvia Marino[1], Katherine Ann Staines[2], Genevieve Brown[3], Rachel Anne Howard-Jones[4] and Magdalena Adamczyk[1,4]. Because of limitations in obtaining healthy human bone samples the explant growth of animal tissue serves as a platform to study the complex physico-chemical properties of the bone. These models enable preserving important cell–cell and cell–matrix interactions in order to better understand the behaviour of cells in their natural three-dimensional environment. We describe how these explants are utilised to study bone development, cartilage and bone metabolism, cancer-induced bone diseases, stem cell-driven bone repair and mechanoadaptation These techniques can be directly used to understand mechanisms linked with bone physiology or bone-associated diseases. BoneKEy Reports 5, Article number: 818 (2016) | doi:10.1038/bonekey.2016.49; published online 29 June 2016
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