Department of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
Department of Biology, Skeletal Research Center, Case Western Reserve University, Cleveland, Ohio, USA.
Tissue Eng Part A. 2022 Mar;28(5-6):254-269. doi: 10.1089/ten.TEA.2021.0112. Epub 2021 Oct 25.
The production of a clinically useful engineered cartilage is an outstanding and unmet clinical need. High-throughput RNA sequencing provides a means of characterizing the molecular phenotype of populations of cells and can be leveraged to better understand differences among source cells, derivative engineered tissues, and target phenotypes. In this study, small RNA sequencing is utilized to comprehensively characterize the microRNA transcriptomes (miRNomes) of native human neonatal articular cartilage and human bone marrow-derived mesenchymal stem cells (hBM-MSCs) differentiating into cartilage organoids, contrasting the microRNA regulation of engineered cartilage with that of a promising target phenotype. Five dominant microRNAs are upregulated during cartilage organoid differentiation and disproportionately regulate transcription factors: miR-148a-3p, miR-140-3p, miR-27b-3p, miR-140-5p, and miR-181a-5p. Two microRNAs that dominate the miRNomes of hBM-MSCs, miR-21-5p and miR-143-3p, persist throughout the differentiation process and may limit the ability of these cells to differentiate into an engineered cartilage resembling target native articular cartilage. By using predictive bioinformatics tools and antagomir inhibition, these persistent microRNAs are shown to destabilize the mRNA of genes with known or potential roles in cartilage biology including , , , , , , , , and These results shed light on the extent to which only a few microRNAs contribute to the complex regulatory environment of hBM-MSCs for engineered tissues. Impact statement MicroRNAs are emerging as important controlling elements in the differentiation of human bone marrow-derived mesenchymal stem cells (hBM-MSCs). By using a robust bioinformatic approach and further validation , here we provide a comprehensive characterization of the microRNA transcriptomes (miRNomes) of a commonly studied and clinically promising source of multipotent cells (hBM-MSCs), a gold standard model of chondrogenesis (hBM-MSC-derived cartilage organoids), and an attractive target phenotype for clinically useful engineered cartilage (neonatal articular cartilage). These analyses highlighted a specific set of microRNAs involved in the chondrogenic program that could be manipulated to acquire a more robust articular cartilage-like phenotype. This characterization provides researchers in the cartilage tissue engineering field a useful atlas with which to contextualize microRNA involvement in complex differentiation pathways.
临床上急需一种有实际用途的工程化软骨，高通量 RNA 测序为细胞群体的分子表型特征提供了一种手段，可借此更好地了解起始细胞、衍生工程组织和目标表型之间的差异。在本研究中，我们利用小 RNA 测序全面分析了天然人新生儿关节软骨和骨髓间充质干细胞（hBM-MSCs）分化为软骨类器官过程中的 microRNA 转录组（miRNome），对比了工程化软骨和有前景的目标表型的 microRNA 调控。在软骨类器官分化过程中，有 5 种主要 microRNA 上调，不成比例地调控转录因子：miR-148a-3p、miR-140-3p、miR-27b-3p、miR-140-5p 和 miR-181a-5p。2 种在 hBM-MSCs 中占主导地位的 microRNA，miR-21-5p 和 miR-143-3p，在整个分化过程中持续存在，可能限制这些细胞分化为类似于目标天然关节软骨的工程化软骨的能力。通过使用预测性生物信息学工具和反义寡核苷酸抑制，这些持续存在的 microRNA 被证明可以使已知或潜在参与软骨生物学的基因的 mRNA 不稳定，包括、、、、、、、和。这些结果揭示了只有少数 microRNA 对 hBM-MSCs 用于工程组织的复杂调控环境有多大影响。 影响陈述 microRNA 正在成为人类骨髓间充质干细胞（hBM-MSCs）分化的重要调控因子。通过使用稳健的生物信息学方法和进一步验证，在这里，我们对一种常用且具有临床应用前景的多能细胞来源（hBM-MSCs）、软骨发生的金标准模型（hBM-MSC 衍生的软骨类器官）以及有吸引力的临床有用工程化软骨的目标表型（新生儿关节软骨）的 microRNA 转录组（miRNome）进行了全面描述。这些分析突出了一组特定的 microRNA 参与软骨形成程序，这些 microRNA 可被操纵以获得更类似关节软骨的表型。这种描述为软骨组织工程领域的研究人员提供了一个有用的图谱，有助于了解 microRNA 在复杂分化途径中的作用。
