Houston Dean A, Staines Katherine A, MacRae Vicky E, Farquharson Colin
Developmental Biology, The Roslin Institute and R(D)SVS, The University of Edinburgh;
Developmental Biology, The Roslin Institute and R(D)SVS, The University of Edinburgh.
J Vis Exp. 2016 Dec 3(118):54978. doi: 10.3791/54978.
The fundamental process of endochondral ossification is under tight regulation in the healthy individual so as to prevent disturbed development and/or longitudinal bone growth. As such, it is imperative that we further our understanding of the underpinning molecular mechanisms involved in such disorders so as to provide advances towards human and animal patient benefit. The mouse metatarsal organ explant culture is a highly physiological ex vivo model for studying endochondral ossification and bone growth as the growth rate of the bones in culture mimic that observed in vivo. Uniquely, the metatarsal organ culture allows the examination of chondrocytes in different phases of chondrogenesis and maintains cell-cell and cell-matrix interactions, therefore providing conditions closer to the in vivo situation than cells in monolayer or 3D culture. This protocol describes in detail the intricate dissection of embryonic metatarsals from the hind limb of E15 murine embryos and the subsequent analyses that can be performed in order to examine endochondral ossification and longitudinal bone growth.
在健康个体中,软骨内成骨的基本过程受到严格调控,以防止发育紊乱和/或纵向骨生长异常。因此,我们必须进一步了解这些疾病背后的分子机制,以便为人类和动物患者带来更多益处。小鼠跖骨器官外植体培养是一种高度生理学的体外模型,用于研究软骨内成骨和骨生长,因为培养中骨骼的生长速度与体内观察到的相似。独特的是,跖骨器官培养允许检查软骨形成不同阶段的软骨细胞,并维持细胞间和细胞与基质的相互作用,因此提供了比单层或三维培养中的细胞更接近体内情况的条件。本方案详细描述了从E15小鼠胚胎后肢精细解剖胚胎跖骨以及随后为检查软骨内成骨和纵向骨生长而可进行的分析。