Yan Zhao, Ruan Banjun, Zhu Zheng, Cao Xiaorui, Lu Zifan
Department of Anatomy and K.K. Leung Brain Research Centre, Fourth Military Medical University, Xi'an, 710032, China.
Institute of Orthopedic Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an, 710032, People's Republic of China.
Hum Cell. 2025 Mar 20;38(3):73. doi: 10.1007/s13577-025-01192-0.
The reciprocal fate decision of mesenchymal stem cells (MSCs) to either bone or adipocytes is determined by Wnt-related signaling and the glucagon-like peptide-1 receptor (GLP-1R). Azoramide, an ER stress alleviator, was reported to have an antidiabetic effect. In this study, we investigated the function of azoramide in regulating the lineage determination of MSCs for either adipogenic or osteogenic differentiation. Microcomputed tomography and histological analysis on bone morphogenetic protein (BMP)2-induced parietal periosteum bone formation assays, C3H10T1/2 and mouse bone marrow MSC-derived bone formation and adipogenesis assays, and specific staining for bone tissue and lipid droplets were used to evaluate the role of azoramide on the lineage determination of MSC differentiation. Cells were harvested for Western blot and quantitative real-time polymerase chain reaction (PCR), and immunofluorescence staining was used to explore the potential mechanism of azoramide for regulating MSC differentiation. Based on MSC-derived bone formation assays both in vivo and in vitro, azoramide treatment displayed a cell fate determining ability in favor of adipogenesis over osteogenesis. Further mechanistic characterizations disclosed that both the GLP-1R agonist peptide exendin-4 (Ex-4) and GLP-1R small interfering (si)RNA abrogated azoramide dual effects. Moreover, cAMP-protein kinase A (PKA)-mediated nuclear β-catenin activity was responsible for the negative function of azoramide on bone formation in favor of adipogenesis. These data provide the first evidence to show that azoramide may serve as an inhibitor against GLP-1R in MSC lineage determination.
间充质干细胞（MSC）向骨细胞或脂肪细胞的相互命运决定由Wnt相关信号传导和胰高血糖素样肽-1受体（GLP-1R）决定。据报道，内质网应激缓解剂阿佐酰胺具有抗糖尿病作用。在本研究中，我们研究了阿佐酰胺在调节MSC成脂或成骨分化的谱系决定中的功能。利用微计算机断层扫描和组织学分析对骨形态发生蛋白（BMP）2诱导的顶骨骨膜骨形成试验、C3H10T1/2和小鼠骨髓MSC来源的骨形成及脂肪生成试验，以及对骨组织和脂滴的特异性染色来评估阿佐酰胺在MSC分化谱系决定中的作用。收获细胞用于蛋白质免疫印迹和定量实时聚合酶链反应（PCR），并使用免疫荧光染色来探索阿佐酰胺调节MSC分化的潜在机制。基于体内和体外的MSC来源的骨形成试验，阿佐酰胺处理显示出一种细胞命运决定能力，有利于脂肪生成而非骨生成。进一步的机制表征表明，GLP-1R激动剂肽艾塞那肽-4（Ex-4）和GLP-1R小干扰（si）RNA均消除了阿佐酰胺的双重作用。此外，cAMP-蛋白激酶A（PKA）介导的核β-连环蛋白活性是阿佐酰胺对骨形成产生负向作用从而有利于脂肪生成的原因。这些数据提供了首个证据表明阿佐酰胺在MSC谱系决定中可能作为GLP-1R的抑制剂。
