Zhang Da-Wei, Chen Tao, Li Jin-Xiang, Wang Hong-Gang, Huang Zong-Wen, Lv Hai
Department of Spine Surgery, The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, Guangdong Province, 519000, PR China.
Department of Orthopaedic and Microsurgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, 510080, PR China.
Regen Ther. 2021 Sep 29;18:391-400. doi: 10.1016/j.reth.2021.09.004. eCollection 2021 Dec.
Osteoporosis, a common skeletal disorder mainly affecting postmenopausal women, is characterized by the imbalance between osteogenesis and osteoclastogenesis. Circ_0134944 has been recently found to be upregulated in postmenopausal osteoporosis (PMOP) patients. However, its role in osteogenesis remains unknown. Here we aimed to explore the role of circ_0134944 in osteogenesis and reveal the underlying mechanism.
qRT-PCR was used to determine the expression of circ_0134944, miR-127-5p, PDX1 and SPHK1 in the blood mononuclear cells (BMCs) of PMOP patients. Bone marrow mesenchymal stem cells (BMSCs) were used as the cellular model. Western blotting and qRT-PCR were used to determine the expression of osteogenesis-related genes (Runx2, OPN, OCN). ALP and Alizarin Red S staining were performed to evaluate osteogenic differentiation. The interactions between circ_0134944 and miR-127-5p, miR-127-5p and PDX1, PDX1 and SPHK1 were determined by dual-luciferase reporter and ChIP assay.
Circ_0134944, PDX1 and SPHK1 were upregulated while miR-127-5p was downregulated in PMOP patients. Enhanced expression of circ_0134944 suppressed osteogenesis, which was then reversed by miR-127-5p overexpression. The binding between circ_0134944 and miR-127-5p, PDX1 and miR-127-5p were confirmed by dual-luciferase reporter assay. Moreover, PDX1 was enriched in the promoter region of SPHK1, and SPHK1 overexpression prevented the promotion of osteogenesis induced by miR-127-5p overexpression.
Taken together, these results demonstrate that circ_0134944 inhibit osteogenesis via miR-127-5p/PDX1/SPHK1 axis. Thus, the present study offered evidence that circ_0134944/miR-127-5p/PDX1/SPHK1 axis could be a potential therapeutic target for PMOP.
骨质疏松症是一种常见的骨骼疾病，主要影响绝经后女性，其特征是成骨与破骨细胞生成之间的失衡。最近发现circ_0134944在绝经后骨质疏松症（PMOP）患者中表达上调。然而，其在成骨过程中的作用尚不清楚。在此，我们旨在探讨circ_0134944在成骨中的作用，并揭示其潜在机制。
采用qRT-PCR检测PMOP患者血液单核细胞（BMC）中circ_0134944、miR-127-5p、PDX1和SPHK1的表达。以骨髓间充质干细胞（BMSC）作为细胞模型。采用蛋白质免疫印迹法和qRT-PCR检测成骨相关基因（Runx2、OPN、OCN）的表达。进行碱性磷酸酶（ALP）和茜素红S染色以评估成骨分化。通过双荧光素酶报告基因和染色质免疫沉淀（ChIP）试验确定circ_0134944与miR-127-5p、miR-127-5p与PDX1、PDX1与SPHK1之间的相互作用。
PMOP患者中circ_0134944、PDX1和SPHK1表达上调，而miR-127-5p表达下调。circ_0134944表达增强抑制了成骨，而miR-127-5p过表达可逆转这种抑制作用。双荧光素酶报告基因试验证实了circ_0134944与miR-127-5p、PDX1与miR-127-5p之间的结合。此外，PDX1在SPHK1启动子区域富集，SPHK1过表达可阻止miR-127-5p过表达诱导的成骨促进作用。
综上所述，这些结果表明circ_013４944通过miR-127-5p/PDX1/SPHK1轴抑制成骨。因此，本研究提供了证据表明circ_0134944/miR-127-5p/PDX1/SPHK1轴可能是PMOP的潜在治疗靶点。
