微纤维相关蛋白 5 通过调节 Wnt/β-连环蛋白和 AMPK 信号通路来调节成骨分化。
Microfibrillar-associated protein 5 regulates osteogenic differentiation by modulating the Wnt/β-catenin and AMPK signaling pathways.
机构信息
Department of Orthopedics, Shanghai Fifth People's Hospital, Fudan University, Shanghai, China.
出版信息
Mol Med. 2021 Dec 5;27(1):153. doi: 10.1186/s10020-021-00413-0.
BACKGROUND
Dysfunctional osteogenesis of bone marrow mesenchymal stem cells (BMSCs) plays an important role in osteoporosis occurrence and development. However, the molecular mechanisms of osteogenic differentiation remain unclear. This study explored whether microfibrillar-associated protein 5 (MFAP5) regulated BMSCs osteogenic differentiation.
METHODS
We used shRNA or cDNA to knock down or overexpress MFAP5 in C3H10 and MC3T3-E1 cells. AR-S- and ALP-staining were performed to quantify cellular osteogenic differentiation. The mRNA levels of the classical osteogenic differentiation biomarkers Runx2, Col1α1, and OCN were quantified by qRT-PCR. Finally, we employed Western blotting to measure the levels of Wnt/β-catenin and AMPK signaling proteins.
RESULTS
At days 0, 3, 7, and 14 after osteogenic induction, AR-S- and ALP-staining was lighter in MFAP5 knockdown compared to control cells, as were the levels of Runx2, Col1α1 and OCN. During osteogenesis, the levels of β-catenin, p-GSK-3β, AMPK, and p-AMPK were upregulated, while that of GSK-3β was downregulated, indicating that Wnt/β-catenin and AMPK signaling were activated. The relevant molecules were expressed at lower levels in the knockdown than control group; the opposite was seen for overexpressing cell lines.
CONCLUSIONS
MFAP5 regulates osteogenesis via Wnt/β‑catenin- and AMPK-signaling; MFAP5 may serve as a therapeutic target in patients with osteoporosis.
背景
骨髓间充质干细胞（BMSCs）的成骨功能障碍在骨质疏松症的发生和发展中起着重要作用。然而，其成骨分化的分子机制尚不清楚。本研究探讨了微纤维相关蛋白 5（MFAP5）是否调节 BMSCs 的成骨分化。
方法
我们使用 shRNA 或 cDNA 敲低或过表达 C3H10 和 MC3T3-E1 细胞中的 MFAP5。通过 AR-S 和 ALP 染色来定量细胞的成骨分化。通过 qRT-PCR 定量经典成骨分化生物标志物 Runx2、Col1α1 和 OCN 的 mRNA 水平。最后，我们通过 Western blot 测定 Wnt/β-catenin 和 AMPK 信号蛋白的水平。
结果
在成骨诱导后第 0、3、7 和 14 天，MFAP5 敲低组的 AR-S 和 ALP 染色比对照组浅，Runx2、Col1α1 和 OCN 的水平也较低。在成骨过程中，β-catenin、p-GSK-3β、AMPK 和 p-AMPK 的水平上调，而 GSK-3β 的水平下调，表明 Wnt/β-catenin 和 AMPK 信号被激活。在敲低组中，相关分子的表达水平低于对照组；而过表达细胞系则相反。
结论
MFAP5 通过 Wnt/β-catenin-和 AMPK 信号通路调节成骨；MFAP5 可能成为骨质疏松症患者的治疗靶点。
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