BACKGROUND Mesenchymal stem cells (MSCs) play a crucial role in maintaining the dynamic balance of bone metabolism. Melatonin may have a regulatory effect on bone metabolism by regulating the lineage commitment and differentiation signalling pathways of MSCs. Among the BMP families, the osteogenesis of BMP9 is considered to be one of the strongest in MSCs. Here, we explored whether melatonin and BMP9 act synergistically on MSC osteogenic differentiation. METHODS The C3H10T1/2 osteogenic differentiation function induced by melatonin synergizes with BMP9, as detected by the expression of osteogenic markers at different periods. The result was further confirmed by foetal limb explant culture and in vivo stem cell implantation experiments. The effects of the AMPK/β-catenin pathway on the osteogenic differentiation of C3H10T1/2 cells were evaluated by Western blotting. RESULTS Melatonin combined with BMP9 significantly enhanced the expression of osteogenic markers at different periods in C3H10T1/2 cells, effectively enhancing BMP9-induced bone formation in cultured foetal explants and ectopic bone formation in vivo in stem cell transplantation experiments. Melatonin increases the expression of BMP9 in C3H10T1/2 cells and induces Smad1/5/8 translocation from the cytoplasm to the nucleus. In addition, melatonin and BMP9 synergistically promote AMPK and β-catenin phosphorylation, which can be largely eliminated by AMPK siRNA pretreatment. CONCLUSIONS Melatonin and BMP9 in C3H10T1/2 cells synergistically promote osteogenic differentiation at least in part by activating the AMPK/β-catenin signalling pathway.

中文翻译：

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褪黑素通过激活AMPK /β-catenin信号通路来促进BMP9诱导的间充质干细胞的成骨分化。

背景技术间充质干细胞（MSC）在维持骨代谢的动态平衡中起关键作用。褪黑激素可能通过调节MSC的血统承诺和分化信号通路而对骨代谢产生调节作用。在BMP家族中，BMP9的成骨作用被认为是MSC中最强的成骨作用之一。在这里，我们探讨了褪黑激素和BMP9是否协同作用于MSC成骨分化。方法褪黑激素诱导的C3H10T1 / 2成骨分化功能与BMP9协同作用，通过不同时期成骨标记物的表达来检测。胎儿肢体外植体培养和体内干细胞植入实验进一步证实了该结果。通过蛋白质印迹法评估了AMPK /β-catenin途径对C3H10T1 / 2细胞成骨分化的影响。结果褪黑素与BMP9结合可显着增强C3H10T1 / 2细胞在不同时期成骨标记的表达，在干细胞移植实验中有效增强BMP9诱导的培养的外植体中骨形成和体内异位骨形成。褪黑素增加C3H10T1 / 2细胞中BMP9的表达，并诱导Smad1 / 5/8从细胞质转移到细胞核。此外，褪黑激素和BMP9协同促进AMPK和β-catenin磷酸化，这可以通过AMPK siRNA预处理大大消除。