Kato Hiroshi, Ochiai-Shino Hiromi, Onodera Shoko, Saito Akiko, Shibahara Takahiko, Azuma Toshifumi
Department of Oral and Maxillofacial Surgery, Tokyo Dental College, Tokyo, Japan.
Department of Biochemistry, Tokyo Dental College, Tokyo, Japan.
Open Biol. 2015 Feb;5(2):140201. doi: 10.1098/rsob.140201.
We recently reported a new method to purify the induced pluripotent stem (iPS)-derived osteoprogenitors (iPSop). In this paper, we optimized the procedure and characterized cells at each process step. We observed that 10 days of treatment with FGF-2, IGF-1 and TGF-β (FIT) resulted in early-phase osteoblasts and 14 days of treatment resulted in late-phase osteoblasts. We found that treatment with 1,25(OH)2 vitamin D3 increased expression of osteocalcin and decreased expression of tissue-non-specific alkaline phosphatase and runt-related transcription factor 2 (RUNX2) in iPSop-day14 cells (cells treated with FIT for 14 days). Therefore, iPSop-day14 cells were promoted to mature osteoblasts by 1,25(OH)2 vitamin D3 treatment. In addition, we found that 1,25(OH)2 vitamin D3 treatment for 14 days enhanced not only mineralization but also expression of osteocyte markers, including dentin matrix protein-1 and fibroblast growth factor-23, in iPSop cells. Therefore, 1,25(OH)2 vitamin D3 is a potent promoter of osteoblast-osteocyte transition. The results of this study suggest that it is possible to evaluate both early- and late-phase osteoblasts and to apply cells to drug screening for anabolic drugs that stimulate bone formation.
我们最近报道了一种纯化诱导多能干细胞（iPS）来源的骨祖细胞（iPSop）的新方法。在本文中，我们优化了该程序，并对每个步骤的细胞进行了表征。我们观察到，用FGF-2、IGF-1和TGF-β（FIT）处理10天可产生早期成骨细胞，处理14天可产生晚期成骨细胞。我们发现，用1,25(OH)₂维生素D₃处理可增加iPSop-day14细胞（用FIT处理14天的细胞）中骨钙素的表达，并降低组织非特异性碱性磷酸酶和 runt相关转录因子2（RUNX2）的表达。因此，1,25(OH)₂维生素D₃处理可促进iPSop-day14细胞向成熟成骨细胞分化。此外，我们发现，用1,25(OH)₂维生素D₃处理14天不仅增强了iPSop细胞的矿化能力，还增强了包括牙本质基质蛋白-1和成纤维细胞生长因子-23在内的骨细胞标志物的表达。因此，1,25(OH)₂维生素D₃是成骨细胞向骨细胞转变的有效促进剂。本研究结果表明，评估早期和晚期成骨细胞以及将细胞应用于刺激骨形成的合成代谢药物的药物筛选是可行的。
