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Enhanced Osteogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells in Ossification of the Posterior Longitudinal Ligament Through Activation of the BMP2-Smad1/5/8 Pathway
Stem Cells and Development ( IF 4 ) Pub Date : 2020-12-09 , DOI: 10.1089/scd.2020.0117
Zhaopeng Cai 1 , Boyang Wu 1 , Guiwen Ye 2 , Wenjie Liu 1 , Keng Chen 1 , Peng Wang 1 , Zhongyu Xie 1 , Jinteng Li 1 , Guan Zheng 1 , Wenhui Yu 1 , Zepeng Su 1 , Jiajie Lin 1 , Yanfeng Wu 3 , Huiyong Shen 1, 2
Affiliation  

Ossification of the posterior longitudinal ligament (OPLL) is characterized by ectopic OPLL. To date, the specific molecular pathogenesis of OPLL has not been clearly elucidated. In this study, bone marrow-derived mesenchymal stem cells obtained from healthy donors (HD-MSCs) and patients with OPLL (OPLL-MSCs) were cultured in osteogenic differentiation medium for 21 days. The osteogenic differentiation capacity was determined by alizarin red S (ARS) and alkaline phosphatase (ALP) assays. Gene expression levels of osteoblastic markers were measured by quantitative reverse transcription–polymerase chain reaction. Protein levels of related genes and the activation of related signaling pathways were measured by western blotting. LDN193189 was used to inhibit the Smad1/5/8 pathway, and small interfering RNA was used to regulate BMP2 expression. Our results showed that the OPLL-MSCs had stronger ARS staining and ALP activity and higher expression of RUNX2, Osterix, and OCN than the HD-MSCs. During osteogenic differentiation, the Smad1/5/8 pathway was overactivated in the OPLL-MSCs, and LDN193189 inhibition reversed the enhanced osteogenic ability of these cells. Besides, BMP2 was upregulated in the OPLL-MSCs. After BMP2 knockdown, the abnormal osteogenic differentiation of OPLL-MSCs was rescued. Thus, abnormal activation of the BMP2-Smad1/5/8 pathway induces enhanced osteogenic differentiation of OPLL-MSCs compared with HD-MSCs. These findings reveal a mechanism of pathological osteogenesis in OPLL and provide a new perspective on inhibiting pathological osteogenesis by regulating BMP2.

中文翻译:

通过激活 BMP2-Smad1/5/8 通路增强人骨髓间充质干细胞在后纵韧带骨化中的成骨分化

后纵韧带 (OPLL) 的骨化以异位 OPLL 为特征。迄今为止,OPLL 的具体分子发病机制尚未明确阐明。在这项研究中,从健康供体 (HD-MSCs) 和 OPLL 患者 (OPLL-MSCs) 获得的骨髓间充质干细胞在成骨分化培养基中培养了 21 天。成骨分化能力由茜素红 S (ARS) 和碱性磷酸酶 (ALP) 测定确定。通过定量逆转录聚合酶链反应测量成骨细胞标志物的基因表达水平。通过蛋白质印迹法测量相关基因的蛋白质水平和相关信号通路的激活。LDN193189用于抑制Smad1/5/8通路,小干扰RNA用于调节BMP2表达。我们的结果表明,与 HD-MSCs 相比,OPLL-MSCs 具有更强的 ARS 染色和 ALP 活性以及更高的 RUNX2、Osterix 和 OCN 表达。在成骨分化过程中,OPLL-MSCs 中 Smad1/5/8 通路被过度激活,LDN193189 抑制逆转了这些细胞增强的成骨能力。此外,BMP2 在 OPLL-MSC 中上调。BMP2敲低后,OPLL-MSCs的异常成骨分化得以挽救。因此,与 HD-MSCs 相比,BMP2-Smad1/5/8 通路的异常激活诱导 OPLL-MSCs 的成骨分化增强。这些发现揭示了 OPLL 病理性成骨的机制,并为通过调节 BMP2 抑制病理性成骨提供了新的视角。Osterix 和 OCN 高于 HD-MSC。在成骨分化过程中,OPLL-MSCs 中 Smad1/5/8 通路被过度激活,LDN193189 抑制逆转了这些细胞增强的成骨能力。此外,BMP2 在 OPLL-MSC 中上调。BMP2敲低后,OPLL-MSCs的异常成骨分化得以挽救。因此,与 HD-MSCs 相比,BMP2-Smad1/5/8 通路的异常激活诱导 OPLL-MSCs 的成骨分化增强。这些发现揭示了 OPLL 病理性成骨的机制,并为通过调节 BMP2 抑制病理性成骨提供了新的视角。Osterix 和 OCN 高于 HD-MSC。在成骨分化过程中,OPLL-MSCs 中 Smad1/5/8 通路被过度激活,LDN193189 抑制逆转了这些细胞增强的成骨能力。此外,BMP2 在 OPLL-MSC 中上调。BMP2敲低后,OPLL-MSCs的异常成骨分化得以挽救。因此,与 HD-MSCs 相比,BMP2-Smad1/5/8 通路的异常激活诱导 OPLL-MSCs 的成骨分化增强。这些发现揭示了 OPLL 病理性成骨的机制,并为通过调节 BMP2 抑制病理性成骨提供了新的视角。此外,BMP2 在 OPLL-MSC 中上调。BMP2敲低后,OPLL-MSCs的异常成骨分化得以挽救。因此,与 HD-MSCs 相比,BMP2-Smad1/5/8 通路的异常激活诱导 OPLL-MSCs 的成骨分化增强。这些发现揭示了 OPLL 病理性成骨的机制,并为通过调节 BMP2 抑制病理性成骨提供了新的视角。此外,BMP2 在 OPLL-MSC 中上调。BMP2敲低后,OPLL-MSCs的异常成骨分化得以挽救。因此,与 HD-MSCs 相比,BMP2-Smad1/5/8 通路的异常激活诱导 OPLL-MSCs 的成骨分化增强。这些发现揭示了 OPLL 病理性成骨的机制,并为通过调节 BMP2 抑制病理性成骨提供了新的视角。
更新日期:2020-12-11
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