Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, Republic of Korea; Personalized Genomic Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, 34141, Republic of Korea.
Asan Institute for Life Sciences, Asan Medical Center, Seoul, 05505, Republic of Korea; Disease Target Structure Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, 34141, Republic of Korea; Department of Functional Genomics, KRIBB School of Bioscience, University of Science and Technology (UST), Daejeon, 34113, Republic of Korea.
Redox Biol. 2020 Oct;37:101716. doi: 10.1016/j.redox.2020.101716. Epub 2020 Sep 9.
The use of human mesenchymal stem cells (hMSCs) in clinical applications requires large-scale cell expansion prior to administration. However, the prolonged culture of hMSCs results in cellular senescence, impairing their proliferation and therapeutic potentials. To understand the role of microRNAs (miRNAs) in regulating cellular senescence in hMSCs, we globally depleted miRNAs by silencing the DiGeorge syndrome critical region 8 (DGCR8) gene, an essential component of miRNA biogenesis. DGCR8 knockdown hMSCs exhibited severe proliferation defects and senescence-associated alterations, including increased levels of reactive oxygen species (ROS). Transcriptomic analysis revealed that the antioxidant gene superoxide dismutase 2 (SOD2) was significantly downregulated in DGCR8 knockdown hMSCs. Moreover, we found that DGCR8 silencing in hMSCs resulted in hypermethylation in CpG islands upstream of SOD2. 5-aza-2'-deoxycytidine treatment restored SOD2 expression and ROS levels. We also found that these effects were dependent on the epigenetic regulator DNA methyltransferase 3 alpha (DNMT3A). Using computational and experimental approaches, we demonstrated that DNMT3A expression was regulated by miR-29a-3p and miR-30c-5p. Overexpression of miR-29a-3p and/or miR-30c-5p reduced ROS levels in DGCR8 knockdown hMSCs and rescued proliferation defects, mitochondrial dysfunction, and premature senescence. Our findings provide novel insights into hMSCs senescence regulation by the miR-29a-3p/miR-30c-5p/DNMT3A/SOD2 axis.
人骨髓间充质干细胞（hMSCs）在临床应用中需要在给药前进行大规模细胞扩增。然而，hMSCs 的长期培养会导致细胞衰老，损害其增殖和治疗潜力。为了了解 microRNAs（miRNAs）在调节 hMSCs 细胞衰老中的作用，我们通过沉默 miRNA 生物发生的必需组成部分 DiGeorge 综合征关键区域 8（DGCR8）基因来全局耗尽 miRNAs。DGCR8 敲低 hMSCs 表现出严重的增殖缺陷和衰老相关改变，包括活性氧物种（ROS）水平增加。转录组分析显示，DGCR8 敲低 hMSCs 中抗氧化基因超氧化物歧化酶 2（SOD2）的表达显著下调。此外，我们发现 hMSCs 中的 DGCR8 沉默导致 SOD2 上游 CpG 岛的过度甲基化。5-氮杂-2'-脱氧胞苷处理可恢复 SOD2 表达和 ROS 水平。我们还发现这些效应依赖于表观遗传调节剂 DNA 甲基转移酶 3α（DNMT3A）。通过计算和实验方法，我们证明了 DNMT3A 表达受 miR-29a-3p 和 miR-30c-5p 的调控。miR-29a-3p 和/或 miR-30c-5p 的过表达可降低 DGCR8 敲低 hMSCs 中的 ROS 水平，并挽救增殖缺陷、线粒体功能障碍和过早衰老。我们的研究结果为 miR-29a-3p/miR-30c-5p/DNMT3A/SOD2 轴调节 hMSCs 衰老提供了新的见解。
