College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China; Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A&F University, Yangling 712100, Shaanxi, China.
Exp Cell Res. 2013 Oct 15;319(17):2684-99. doi: 10.1016/j.yexcr.2013.08.027. Epub 2013 Sep 7.
Embryonic stem cells (ESCs) can proliferate indefinitely in vitro and differentiate into cells of all three germ layers. These unique properties make them exceptionally valuable for drug discovery and regenerative medicine. However, the practical application of ESCs is limited because it is difficult to derive and culture ESCs. It has been demonstrated that CHIR99021 (CHIR) promotes self-renewal and enhances the derivation efficiency of mouse (m)ESCs. However, the downstream targets of CHIR are not fully understood. In this study, we identified CHIR-regulated genes in mESCs using microarray analysis. Our microarray data demonstrated that CHIR not only influenced the Wnt/β-catenin pathway by stabilizing β-catenin, but also modulated several other pluripotency-related signaling pathways such as TGF-β, Notch and MAPK signaling pathways. More detailed analysis demonstrated that CHIR inhibited Nodal signaling, while activating bone morphogenetic protein signaling in mESCs. In addition, we found that pluripotency-maintaining transcription factors were up-regulated by CHIR, while several developmental-related genes were down-regulated. Furthermore, we found that CHIR altered the expression of epigenetic regulatory genes and long intergenic non-coding RNAs. Quantitative real-time PCR results were consistent with microarray data, suggesting that CHIR alters the expression pattern of protein-encoding genes (especially transcription factors), epigenetic regulatory genes and non-coding RNAs to establish a relatively stable pluripotency-maintaining network.
胚胎干细胞（ESCs）在体外可无限增殖并分化为三个胚层的细胞。这些独特的性质使它们在药物发现和再生医学中具有极高的价值。然而，由于难以获得和培养 ESCs，其实际应用受到限制。已经证明 CHIR99021（CHIR）可促进自我更新并提高小鼠（m）ESC 的衍生效率。然而，CHIR 的下游靶点尚未完全阐明。在本研究中，我们使用微阵列分析鉴定了 mESC 中 CHIR 调节的基因。我们的微阵列数据表明，CHIR 不仅通过稳定 β-catenin 影响 Wnt/β-catenin 通路，还调节了其他几个与多能性相关的信号通路，如 TGF-β、Notch 和 MAPK 信号通路。更详细的分析表明，CHIR 抑制了 Nodal 信号通路，同时激活了 mESC 中的骨形态发生蛋白信号通路。此外，我们发现 CHIR 上调了多能性维持转录因子，而下调了几个发育相关基因。此外，我们发现 CHIR 改变了表观遗传调节基因和长链非编码 RNA 的表达。实时定量 PCR 结果与微阵列数据一致，表明 CHIR 通过改变蛋白质编码基因（尤其是转录因子）、表观遗传调节基因和非编码 RNA 的表达模式，建立了一个相对稳定的多能性维持网络。
