Qin Haotian, Weng Jian, Zhou Bo, Zhang Weifei, Li Guoqing, Chen Yingqi, Qi Tiantian, Zhu Yuanchao, Yu Fei, Zeng Hui
Department of Bone & Joint Surgery, Peking University Shenzhen Hospital, Shenzhen, 518036, China.
National & Local Joint Engineering Research Center of Orthopaedic Biomaterials, Peking University Shenzhen Hospital, Shenzhen, 518036, China.
Biol Trace Elem Res. 2023 Jun;201(6):2823-2842. doi: 10.1007/s12011-022-03364-7. Epub 2022 Jul 23.
Bone defects are often caused by trauma or surgery and can lead to delayed healing or even bone nonunion, thereby resulting in impaired function of the damaged site. Magnesium ions and related metallic materials play a crucial role in repairing bone defects, but the mechanism remains unclear. In this study, we induced the angiogenic differentiation of bone marrow stromal cells (BMSCs) with different concentrations of magnesium ions. The mechanism was investigated using γ-secretase inhibitor (DAPT) at different time points (7 and 14 days). Angiogenesis, differentiation, migration, and chemotaxis were detected using the tube formation assay, wound-healing assay, and Transwell assay. Besides, we analyzed mRNA expression and the angiogenesis-related protein levels of genes by RT-qPCR and western blot. We discovered that compared with other concentrations, the 5 mM magnesium ion concentration was more conducive to forming tubes. Additionally, hypoxia-inducible factor 1 alpha (Hif-1α) and endothelial nitric oxide (eNOS) expression both increased (p < 0.05). After 7 and 14 days of induction, 5 mM magnesium ion group tube formation, migration, and chemotaxis were enhanced, and the expression of Notch pathway genes increased. Moreover, expression of the Notch target genes hairy and enhancer of split 1 (Hes1) and Hes5 (hairy and enhancer of split 5), as well as the angiogenesis-related genes Hif-1α and eNOS, were enhanced (p < 0.05). However, these trends did not occur when DAPT was applied. This indicates that 5 mM magnesium ion is the optimal concentration for promoting the angiogenesis and differentiation of BMSCs in vitro. By activating the Notch signaling pathway, magnesium ions up-regulate the downstream genes Hes1 and Hes5 and the angiogenesis-related genes Hif-1α and eNOS, thereby promoting the angiogenesis differentiation of BMSCs. Additionally, magnesium ion-induced differentiation enhances the migration and chemotaxis of BMSCs. Thus, we can conclude that magnesium ions and related metallic materials promote angiogenesis to repair bone defects. This provides the rationale for developing artificial magnesium-containing bone materials through tissue engineering.
骨缺损常由创伤或手术引起，可导致愈合延迟甚至骨不连，从而致使受损部位功能受损。镁离子及相关金属材料在骨缺损修复中起关键作用，但其机制尚不清楚。在本研究中，我们用不同浓度的镁离子诱导骨髓间充质干细胞（BMSCs）的血管生成分化。在不同时间点（7天和14天）使用γ-分泌酶抑制剂（DAPT）研究其机制。采用管形成实验、伤口愈合实验和Transwell实验检测血管生成、分化、迁移和趋化性。此外，我们通过RT-qPCR和蛋白质免疫印迹法分析基因的mRNA表达和血管生成相关蛋白水平。我们发现，与其他浓度相比，5 mM镁离子浓度更有利于形成血管管腔。此外，缺氧诱导因子1α（Hif-1α）和内皮型一氧化氮（eNOS）的表达均增加（p < 0.05）。诱导7天和14天后，5 mM镁离子组的管形成、迁移和趋化性增强，Notch信号通路基因的表达增加。此外，Notch靶基因毛状分裂增强子1（Hes1）和毛状分裂增强子5（Hes5）以及血管生成相关基因Hif-1α和eNOS的表达均增强（p < 0.05）。然而，应用DAPT时未出现这些趋势。这表明5 mM镁离子是体外促进BMSCs血管生成和分化的最佳浓度。通过激活Notch信号通路，镁离子上调下游基因Hes1和Hes5以及血管生成相关基因Hif-1α和eNOS，从而促进BMSCs的血管生成分化。此外，镁离子诱导的分化增强了BMSCs的迁移和趋化性。因此，我们可以得出结论，镁离子及相关金属材料通过促进血管生成来修复骨缺损。这为通过组织工程开发含镁人工骨材料提供了理论依据。
