Department of Chemical and Biological Engineering, University of Ottawa, Ottawa, Canada.
Lunenfeld-Tanenbaum Research Institute, Mt Sinai Hospital, Toronto, Ontario, Canada.
Acta Biomater. 2014 Sep;10(9):3899-910. doi: 10.1016/j.actbio.2014.06.008. Epub 2014 Jun 16.
Elasmobranchs (e.g. sharks and rays), like all fishes, grow continuously throughout life. Unlike other vertebrates, their skeletons are primarily cartilaginous, comprising a hyaline cartilage-like core, stiffened by a thin outer array of mineralized, abutting and interconnected tiles called tesserae. Tesserae bear active mineralization fronts at all margins and the tesseral layer is thin enough to section without decalcifying, making this a tractable but largely unexamined system for investigating controlled apatite mineralization, while also offering a potential analog for endochondral ossification. The chemical mechanism for tesserae mineralization has not been described, but has been previously attributed to spherical precursors, and alkaline phosphatase (ALP) activity. Here, we use a variety of techniques to elucidate the involvement of phosphorus-containing precursors in the formation of tesserae at their mineralization fronts. Using Raman spectroscopy, fluorescence microscopy and histological methods, we demonstrate that ALP activity is located with inorganic phosphate polymers (polyP) at the tessera-uncalcified cartilage interface, suggesting a potential mechanism for regulated mineralization: inorganic phosphate (Pi) can be cleaved from polyP by ALP, thus making Pi locally available for apatite biomineralization. The application of exogenous ALP to tissue cross-sections resulted in the disappearance of polyP and the appearance of Pi in uncalcified cartilage adjacent to mineralization fronts. We propose that elasmobranch skeletal cells control apatite biomineralization by biochemically controlling polyP and ALP production, placement and activity. Previous identification of polyP and ALP shown previously in mammalian calcifying cartilage supports the hypothesis that this mechanism may be a general regulating feature in the mineralization of vertebrate skeletons.
软骨鱼（例如鲨鱼和鳐鱼）与所有鱼类一样，在其整个生命周期中都在不断生长。与其他脊椎动物不同，它们的骨骼主要由软骨组成，由透明软骨样核心组成，由薄的外部排列的矿化、相邻和相互连接的瓦片（称为瓦片）加固。瓦片在所有边缘都具有活跃的矿化前沿，并且瓦片层足够薄，可以不经脱钙就进行切片，这使得这个系统易于处理，但在很大程度上尚未被研究，可用于研究受控磷灰石矿化，同时也为软骨内骨化提供了潜在的模拟。瓦片矿化的化学机制尚未描述，但以前归因于球形前体和碱性磷酸酶（ALP）活性。在这里，我们使用多种技术阐明含磷前体在瓦片矿化前沿形成中的参与。使用拉曼光谱、荧光显微镜和组织学方法，我们证明 ALP 活性位于瓦片-未钙化软骨界面处的无机磷酸盐聚合物（polyP）中，这表明了一种潜在的调节矿化机制：ALP 可以从 polyP 中裂解无机磷酸盐（Pi），从而使 Pi 在局部可用于磷灰石生物矿化。将外源性 ALP 应用于组织切片导致 polyP 的消失和紧邻矿化前沿的未钙化软骨中 Pi 的出现。我们提出，软骨鱼骨骼细胞通过生化控制 polyP 和 ALP 的产生、位置和活性来控制磷灰石生物矿化。以前在哺乳动物钙化软骨中发现的 polyP 和 ALP 的鉴定支持了这样一种假设，即该机制可能是脊椎动物骨骼矿化的一个普遍调节特征。
