Hara Emilio Satoshi, Okada Masahiro, Kuboki Takuo, Nakano Takayoshi, Matsumoto Takuya
Department of Biomaterials, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama-shi, Okayama-ken, 700-8525, Okayama, Japan.
J Mater Chem B. 2018 Oct 14;6(38):6153-6161. doi: 10.1039/c8tb01544a. Epub 2018 Sep 11.
Bone is a sophisticated organic-inorganic hybrid material, whose formation involves a complex spatio-temporal sequence of events regulated by the cells. A deeper understanding of the mechanisms behind bone mineralization at different size scales, and using a multidisciplinary approach, may uncover novel pathways for the design and fabrication of functional bone tissue in vitro. The objectives of this study were first to investigate the environmental factors that prime initial mineralization using the secondary ossification center as an in vivo model, and then to apply the obtained knowledge for rapid in vitro synthesis of bone-like tissue. First, the direct and robust measurement of pH showed that femur epiphysis is alkaline (pH ≅ 8.5) at the initial mineral stage at post-natal day 6. We showed that the alkaline milieu is decisive not only for alkaline phosphatase activity, which precedes mineral formation at P6, but also for determining initial mineral precipitation and spherical morphology. Next, engineering approaches were used to synthesize bone-like tissue based on alkaline milieu and artificial chondrocyte membrane nanofragments, previously shown to be the nucleation site for mineral formation. Interestingly, mineralization using artificial cell membrane nanofragments was achieved in just 1 day. Finally, ex vivo culture of femur epiphysis in alkaline pH strongly induced chondrocyte burst, which was previously shown to be the origin of chondrocyte membrane nanofragments, and also enhanced mineral formation. Taken together, these findings not only shed more light on the microenvironmental conditions that prime initial bone formation in vivo, but they also show that alkaline milieu can be used as an important factor for enhancing methods for in vitro synthesis of bone tissue.
骨骼是一种复杂的有机 - 无机复合材料，其形成涉及由细胞调节的复杂时空事件序列。采用多学科方法，更深入地了解不同尺度下骨矿化背后的机制，可能会揭示体外功能性骨组织设计与制造的新途径。本研究的目的首先是利用次级骨化中心作为体内模型，研究引发初始矿化的环境因素，然后将获得的知识应用于体外快速合成类骨组织。首先，对pH的直接且可靠的测量表明，出生后第6天股骨骨骺在初始矿化阶段呈碱性（pH≈8.5）。我们发现，碱性环境不仅对出生后第6天在矿化形成之前的碱性磷酸酶活性起决定性作用，而且对确定初始矿物质沉淀和球形形态也起决定性作用。接下来，采用工程方法基于碱性环境和人工软骨细胞膜纳米片段合成类骨组织，人工软骨细胞膜纳米片段先前已被证明是矿物质形成的成核位点。有趣的是，使用人工细胞膜纳米片段仅在1天内就实现了矿化。最后，在碱性pH条件下对股骨骨骺进行离体培养强烈诱导软骨细胞破裂，软骨细胞破裂先前已被证明是软骨细胞膜纳米片段的来源，并且还增强了矿物质形成。综上所述，这些发现不仅更清楚地揭示了体内引发初始骨形成的微环境条件，而且还表明碱性环境可作为增强骨组织体外合成方法的重要因素。
