Hypoxia reduces the osteogenic differentiation of peripheral blood mesenchymal stem cells by upregulating Notch-1 expression
Hypoxia reduces the osteogenic differentiation of peripheral blood mesenchymal stem cells by upregulating Notch-1 expression
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缺氧通过上调Notch-1表达减少外周血间充质干细胞的成骨分化
DOI：
10.1080/03008207.2019.1611792
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发表时间：
2019-05-29
影响因子：
2.9
通讯作者：
Zhu, Lixin
中科院分区：
文献类型：
--
作者：
Yang, Minsheng;Liu, Haixin;Zhu, Lixin
摘要目的：骨髓间充质干细胞(MSCs)种植于生物相容性支架上，具有骨修复的潜能。然而，由于支架中缺乏血管，MSCs会受到缺氧和营养不足的影响。在此，我们探讨了低氧对MSC分化的影响，以阐明这些机制。方法：外周血间充质干细胞(PBMSCs)在氧气浓度分别为1%、9%、21%的小室内培养。用细胞计数试剂盒8法检测细胞增殖，用活/死实验测定细胞存活率。采用划痕试验评价细胞迁移情况。通过钙沉积/矿化实验、逆转录实时定量聚合酶链式反应和Western blotting检测细胞的成骨分化情况。慢病毒转染的PBMSCs下调NOTCH1的表达，观察Notch1基因敲除对成骨基因和蛋白表达的影响。结果：在无血清条件下，PBMSCs在低氧(1%O2)条件下增殖加快，迁移增加，存活率降低。尽管9%的氧气促进了成骨分化，但1%O2显著降低了PBMSCs的成骨分化，这种作用与Notch1的表达增加有关。使用小干扰RNA降低Notch1的表达可显著恢复PBMSCs的成骨分化。结论：低氧通过增加Notch1的表达促进细胞增殖，增加迁移，减少PBMSC向成骨细胞的分化。这些发现可能有助于发展合适的细胞培养或体内移植条件，以保持PBMSCs的全部成骨潜力。
ABSTRACT Purpose: Mesenchymal stem cells (MSCs) seeded on biocompatible scaffolds have therapeutic potential for bone defect repair. However, MSCs can be affected by hypoxia and nutritional deficiency due to a lack of blood vessels in the scaffolds. Here, we explored the effects of hypoxia on MSC differentiation to clarify these mechanisms. Methods: Peripheral blood mesenchymal stem cells (PBMSCs) were cultured in small individual chambers with oxygen concentrations of 1%, 9%, and 21%. Cell proliferation was evaluated by Cell Counting Kit 8 assays, and cell survival was determined using live/dead assays. Scratch assays were performed to evaluate cell migration. Ca2+ deposition/mineralization experiments, reverse transcription quantitative real-time polymerase chain reaction, and Western blotting were performed to assess the osteogenic differentiation of cells. Notch1 expression was downregulated by lentivirus-transfected PBMSCs to observe the effects of Notch1 knockdown on osteogenic gene and protein expression. Results: PBMSCs exposed to hypoxia (1% O2) demonstrated accelerated proliferation, increased migration, and reduced survival in the absence of serum. Although 9% oxygen promoted osteogenic differentiation, the osteogenic differentiation of PBMSCs was significantly reduced by 1% O2, and this effect was associated with increased Notch1 expression. Reducing Notch1 expression using small interfering RNA significantly restored the osteogenic differentiation of PBMSCs. Conclusions: Hypoxia accelerated proliferation, increased migration, and reduced PBMSC differentiation into osteoblasts by increasing Notch1 expression. These findings may contribute to the development of appropriate cell culture or in vivo transplantation conditions to maintain the full osteogenic potential of PBMSCs.