Department of Orthopedics, Jingzhou First People's Hospital, The First Affiliated Hospital of Yangtze University, Jingzhou, China.
Eur Rev Med Pharmacol Sci. 2019 Sep;23(18):7766-7774. doi: 10.26355/eurrev_201909_18986.
The aim of this study was to explore the exact role of miRNA-365a-3p in the progression of osteoporosis, as well as its function in regulating osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs).
The serum level of miRNA-365a-3p in osteoporosis patients and normal controls was determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). After transfection of miRNA-365a-3p mimics, miRNA-365a-3p inhibitor or si-RUNX2 in hBMSCs, the relative expression levels of miRNA-365a-3p, osteocalcin (OCN), osteopontin (OPN) and collagen I were determined by qRT-PCR. Western blot was conducted to examine the protein expression of RUNX2 influenced by miRNA-365a-3p. Subsequently, the regulatory effects of miRNA-365a-3p and RUNX2 on osteogenic differentiation and capability of mineralization were evaluated by alkaline phosphatase (ALP) determination and alizarin red staining, respectively. Furthermore, the binding relationship between miRNA-365a-3p and RUNX2 was predicted and verified by miRanda and Dual-Luciferase reporter gene assay, respectively.
MiRNA-365a-3p was highly expressed in osteoporosis patients. The expression of miRNA-365a-3p in hBMSCs decreased gradually with the prolongation of osteogenic differentiation. The subsequent results showed that RUNX2 could bind to miRNA-365a-3p, which was negatively regulated by miRNA-365a-3p in hBMSCs. Down-regulation of miRNA-365a-3p significantly decreased the expression levels of OCN, OPN and collagen I. Furthermore, overexpression of miRNA-365a-3p markedly weakened the capability of mineralization of hBMSCs, whereas was further reversed by transfection of si-RUNX2.
MiRNA-365a-3p negatively regulates osteogenic differentiation of hBMSCs by targeting RUNX2, thus promoting the progression of osteoporosis.
本研究旨在探讨 miRNA-365a-3p 在骨质疏松症进展中的确切作用，以及其在调节人骨髓间充质干细胞（hBMSCs）成骨分化中的功能。
采用实时定量聚合酶链反应（qRT-PCR）检测骨质疏松症患者和正常对照者血清中 miRNA-365a-3p 的水平。转染 miRNA-365a-3p 模拟物、miRNA-365a-3p 抑制剂或 si-RUNX2 后，采用 qRT-PCR 检测 hBMSCs 中 miRNA-365a-3p、骨钙素（OCN）、骨桥蛋白（OPN）和Ⅰ型胶原的相对表达水平。采用 Western blot 检测 miRNA-365a-3p 对 RUNX2 蛋白表达的影响。随后，通过碱性磷酸酶（ALP）测定和茜素红染色分别评估 miRNA-365a-3p 和 RUNX2 对成骨分化和矿化能力的调节作用。此外，通过 miRanda 和双荧光素酶报告基因检测分别预测和验证 miRNA-365a-3p 和 RUNX2 之间的结合关系。
miRNA-365a-3p 在骨质疏松症患者中高表达。hBMSCs 中成骨分化时间延长，miRNA-365a-3p 的表达逐渐降低。随后的结果表明，RUNX2 可与 miRNA-365a-3p 结合，miRNA-365a-3p 在 hBMSCs 中受到负调控。下调 miRNA-365a-3p 显著降低 OCN、OPN 和胶原 I 的表达水平。此外，miRNA-365a-3p 的过表达显著削弱了 hBMSCs 的矿化能力，而转染 si-RUNX2 则进一步逆转了这种能力。
miRNA-365a-3p 通过靶向 RUNX2 负调控 hBMSCs 的成骨分化，从而促进骨质疏松症的进展。
