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Abstract：
Human embryonic stem cells (hESCs) are advantaged sources for large-scale and homogeneous mesenchymal stem/stromal cells (MSCs) generation. However, due to the limitations in high-efficiency procedures for hESC-MSCs induction, the systematic and detailed information of mesengenesis and early MSC development are largely obscure. In this study, we took advantage of the well-established twist-related protein 1 (TWIST1)-overexpressing hESCs and two small molecular cocktails (CHIR99021, decitabine) for high-efficient MSC induction. To assess the multidimensional biological and transcriptomic characteristics, we turned to cellular and molecular methods, such as flow cytometry (FCM), quantitative reverse transcription-polymerase chain reaction (qRT-PCR), in vitro tri-lineage differentiation, cytokine secretion analysis, in vivo transplantation for acute liver injury (ALI) management, and bioinformatics analyses (eg, gene ontology-biological processes [GO-BP], Kyoto Encyclopedia of Genes and Genomes [KEGG], HeatMap, and principal component analysis [PCA]). By combining TWIST1 overexpression (denoted as T) and the indicated small molecular cocktails (denoted as S), hESCs high-efficiently differentiated into MSCs (denoted as TS-MSCs, induced by T and S combination) within 2 weeks. TS-MSCs satisfied the criteria for MSC definition and revealed comparable tri-lineage differentiation potential and ameliorative efficacy upon ALI mice. According to RNA-sequencing (SEQ) analysis, we originally illuminated the gradual variations in gene expression pattern and the concomitant biofunctions of the programmed hESC-MSCs. Overall, our data indicated the feasibility of high-efficient generation of hESC-MSCs by TWIST1 and cocktail-based programming. The generated hESC-MSCs revealed multifaceted in vivo and in vitro biofunctions as adult BM-MSCs, which collectively suggested promising prospects in ALI management in future.
摘要：
人胚胎干细胞（hESC）是大规模和均质的间充质干细胞/基质细胞（MSC）产生的有利来源。然而,由于hESC-MSCs诱导的高效程序的局限性，有关中膜发生和早期MSC发育的系统和详细信息在很大程度上是模糊的。在这项研究中,我们利用已确立的扭曲相关蛋白1(TWIST1)过表达hESCs和两种小分子混合物(CHIR99021,地西他滨)进行高效MSC诱导.为了评估多维生物学和转录组特征，我们转向细胞和分子方法，如流式细胞术(FCM)，定量逆转录聚合酶链反应（qRT-PCR），体外三谱系分化，细胞因子分泌分析，体内移植治疗急性肝损伤(ALI),和生物信息学分析(例如，基因本体论-生物过程[GO-BP]，京都基因和基因组百科全书[KEGG]，HeatMap,和主成分分析[PCA])。通过组合TWIST1过表达（表示为T）和所示的小分子混合物（表示为S），hESCs高效分化成MSCs(表示为TS-MSCs，由T和S联合诱导）在2周内。TS-MSC满足MSC定义的标准，并显示出相当的三谱系分化潜力和对ALI小鼠的改善功效。根据RNA测序（SEQ）分析，我们最初阐明了基因表达模式的逐渐变化和程序性hESC-MSCs伴随的生物功能。总的来说,我们的数据表明通过TWIST1和基于混合物的编程高效生成hESC-MSCs的可行性.产生的hESC-MSCs在体内和体外表现出多方面的生物功能，作为成年BM-MSCs，这共同暗示了未来ALI管理的前景。
