IRF4 通过转录激活 miR-636/DOCK9 轴抑制 BM-MSCs 的成骨分化。
IRF4 suppresses osteogenic differentiation of BM-MSCs by transcriptionally activating miR-636/DOCK9 axis.
机构信息
Orthopedics, The First Affiliated Hospital of Jinzhou Medical University, China.
Dental Department, The Second Affiliated Hospital of Jinzhou Medical University, China.
出版信息
Clinics (Sao Paulo). 2022 Apr 6;77:100019. doi: 10.1016/j.clinsp.2022.100019. eCollection 2022.
OBJECTIVES
Osteoblasts are derived from Bone Marrow-derived Mesenchymal Stem Cells (BM-MSCs), which play an indispensable role in bone formation. In this study, the authors aim to investigate the role of IRF4 in the osteogenic differentiation of BM-MSCs and its potential molecular mechanism.
METHODS
The authors used lentivirus infection to overexpress IRF4 in BM-MSCs. The expression of IRF4 and osteogenesis-related genes were detected by qRT-PCR and western blot analysis. The osteogenic differentiation of BM-MSCs was evaluated by Alkaline Phosphatase (ALP) activity, Alizarin red staining, and Alkaline Phosphatase (ALP) staining. Chromatin Immunoprecipitation (ChIP), Dual-Luciferase reporter assay and RNA Immunoprecipitation Assay were applied to confirm the regulatory mechanism between IRF4, miR-636 and DOCK9.
RESULTS
The authors found IRF4 was down-regulated during the osteogenic differentiation of BM-MSCs, and IRF4 overexpression could decrease the osteogenic differentiation of BM-MSCs by specifically promoting the reduction of Alkaline Phosphatase (ALP) activity and down-regulating osteogenic indicators, including OCN, OPN, Runx2 and CollA1. Mechanistically, IRF4 activated microRNA-636 (miR-636) expression via binding to its promoter region, and Dedicator of Cytokinesis 9 (DOCK9) was identified as the target of miR-636 in BM-MSCs. Moreover, the damage in the capacity of osteogenic differentiation of BM-MSCs induced by IRF4 overexpression could be rescued by miR-636 inhibition.
CONCLUSIONS
In summary, this paper proposed that IRF4/miR-636/DOCK9 may be considered as targets for the treatment of osteoporosis (OP).
目的
成骨细胞来源于骨髓间充质干细胞（BM-MSCs），在骨形成中起着不可或缺的作用。本研究旨在探讨 IRF4 在 BM-MSCs 成骨分化中的作用及其潜在的分子机制。
方法
作者通过慢病毒感染过表达 BM-MSCs 中的 IRF4。通过 qRT-PCR 和 Western blot 分析检测 IRF4 和成骨相关基因的表达。通过碱性磷酸酶（ALP）活性、茜素红染色和碱性磷酸酶（ALP）染色评估 BM-MSCs 的成骨分化。应用染色质免疫沉淀（ChIP）、双荧光素酶报告基因检测和 RNA 免疫沉淀assay 来验证 IRF4、miR-636 和 DOCK9 之间的调控机制。
结果
作者发现，在 BM-MSCs 的成骨分化过程中，IRF4 表达下调，IRF4 过表达可通过特异性降低碱性磷酸酶（ALP）活性和下调成骨标志物，包括 OCN、OPN、Runx2 和 CollA1，从而减少 BM-MSCs 的成骨分化。机制上，IRF4 通过结合其启动子区域激活 microRNA-636（miR-636）的表达，并且在 BM-MSCs 中鉴定出细胞分裂蛋白 9（DOCK9）是 miR-636 的靶标。此外，IRF4 过表达诱导的 BM-MSCs 成骨分化能力损伤可通过 miR-636 抑制得到挽救。
结论
综上所述，本研究提出 IRF4/miR-636/DOCK9 可作为骨质疏松症（OP）治疗的靶点。
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