Meesuk Ladda, Kheolamai Pakpoom, Tantrawatpan Chairat, Suwanprateeb Jintamai, Manochantr Sirikul
Division of Cell Biology, Department of Preclinical Sciences, Faculty of Medicine, Thammasat University, Pathumthani, 12120, Thailand.
Center of Excellence in Stem Cell Research and Innovation, Thammasat University, Pathumthani, 12120, Thailand.
Sci Rep. 2025 Aug 5;15(1):28485. doi: 10.1038/s41598-025-13093-1.
Umbilical cord-derived human mesenchymal stem cells (UC-hMSCs) are multipotent stem cells with great potential for treating bone diseases. Although they can be easily isolated from umbilical cord tissue, their osteogenic differentiation is less efficient than differentiation of bone marrow-derived hMSCs (BM-hMSCs). Improving osteogenic differentiation of UC-hMSCs is essential for their clinical use. This study identified specific microRNAs (miRNAs) that inhibit osteogenic differentiation and explored their regulatory mechanisms to improve the osteogenic potential of UC-hMSCs. High-throughput miRNA expression analysis was performed to identify miRNAs involved in osteogenic differentiation. Quantitative real-time RT-PCR confirmed the expression levels of these miRNAs during osteogenic differentiation. The effects of specific anti-miRNAs on osteogenic differentiation were evaluated using alkaline phosphatase (ALP) activity, Alizarin Red S staining, and osteogenic gene expression assays. Analysis revealed significant differential expression of 806 miRNAs in high-osteogenic UC-hMSCs and 760 miRNAs in low-osteogenic UC-hMSCs. Four miRNAs-miR-21, miR-27b, miR-29a, and let-7b-were significantly down-regulated during osteogenic differentiation in high-osteogenic UC-hMSCs but remained elevated in low-osteogenic UC-hMSCs. Inhibition of these miRNAs using specific anti-miRs significantly increased osteogenic gene expression, ALP activity, and matrix mineralization. These effects could be partially mediated by modulation of the PI3K/Akt and Wnt/β-catenin signaling pathways, which led to the up-regulation of RUNX2 expression in UC-hMSCs. Our findings indicate that miR-21, miR-27b, miR-29a, and let-7b are important regulators of osteogenic differentiation in UC-hMSCs. Targeting these miRNAs could enhance osteogenic differentiation by modulating the PI3K/Akt and Wnt/β-catenin signaling pathways, leading to increased RUNX2 expression. These findings provide valuable insights into the role of specific miRNAs in regulating osteogenic differentiation of UC-hMSCs and highlight potential therapeutic strategies for bone regeneration through miRNA modulation.
脐带源人间充质干细胞（UC-hMSCs）是具有治疗骨疾病巨大潜力的多能干细胞。尽管它们可以很容易地从脐带组织中分离出来，但其成骨分化效率低于骨髓源hMSCs（BM-hMSCs）的分化效率。提高UC-hMSCs的成骨分化能力对其临床应用至关重要。本研究鉴定了抑制成骨分化的特定微小RNA（miRNA），并探索了它们的调控机制以提高UC-hMSCs的成骨潜能。进行高通量miRNA表达分析以鉴定参与成骨分化的miRNA。定量实时RT-PCR证实了这些miRNA在成骨分化过程中的表达水平。使用碱性磷酸酶（ALP）活性、茜素红S染色和成骨基因表达测定评估了特异性抗miRNA对成骨分化的影响。分析显示，高成骨UC-hMSCs中有806个miRNA和低成骨UC-hMSCs中有760个miRNA存在显著差异表达。在高成骨UC-hMSCs的成骨分化过程中，四种miRNA——miR-21、miR-27b、miR-29a和let-7b——显著下调，但在低成骨UC-hMSCs中仍保持升高。使用特异性抗miR抑制这些miRNA可显著增加成骨基因表达、ALP活性和基质矿化。这些作用可能部分通过PI3K/Akt和Wnt/β-连环蛋白信号通路的调节介导，这导致UC-hMSCs中RUNX2表达上调。我们的研究结果表明，miR-21、miR-27b、miR-29a和let-7b是UC-hMSCs成骨分化的重要调节因子。靶向这些miRNA可通过调节PI3K/Akt和Wnt/β-连环蛋白信号通路增强成骨分化，导致RUNX2表达增加。这些发现为特定miRNA在调节UC-hMSCs成骨分化中的作用提供了有价值的见解，并突出了通过miRNA调节进行骨再生的潜在治疗策略。
