转录因子CREB对人干细胞稳态和衰老的调控及机制研究

The people all over the world have been confronted with the challenges from population aging, while how to achieve a healthy aging world is still a social and scientific problem. Stem cell exhaustion is a hallmark of individual aging. It has been proved that cAMP-responsive element-binding protein (CREB) involves in a variety of biological processes, such as energy metabolism, neural development, tissue differentiation and lifespan regulation in worms and mouse, indicating that CREB may play an important role in regulating homeostasis and senescence of human stem cells. Investigating the role of CREB in human stem cells can help to further understand the function of CREB in regulating human development, aging or diseases. Here, we aim to generate CREB-null human embryonic stem cells (hESCs) by using CRISPR/Cas9-based gene editing technique, explore the role of CREB on pluripotency maintenance and the differentiation ability towards neural stem cells, mesenchymal stem cells, vascular endothelial cells as well as other cell types. By combination of transcriptomic, epigenomic, proteomic and genomic analyses, we expect to reveal the molecular signaling networks regulated by CREB in different types of human stem cells. This project will contribute to a better understanding of the role of CREB in human stem cell aging, and further provide novel therapeutic targets for aging-related diseases.

当前，世界各国都面临着人口老龄化的严峻挑战。研究表明，干细胞的耗竭是机体衰老的重要因素之一。CREB在啮齿类和低等动物的能量代谢、神经发育、组织分化、寿命调控等诸多生物学过程中扮演着重要的角色，预示着其可能在人干细胞稳态维持及衰老调控中也发挥着重要的作用，因此，揭示CREB在人干细胞稳态维持中的作用及分子机制可以帮助我们进一步了解CREB在人细胞水平的功能和机制，从而为阐明CREB信号通路在人类发育、衰老和疾病中的作用提供线索。本项目将利用CRISPR/Cas9介导的基因编辑技术产生CREB缺失的人胚胎干细胞系，以此探究CREB对于人多能干细胞干性维持及向神经干细胞、间充质干细胞和血管内皮细胞等细胞类型分化能力的影响，同时结合转录组、表观遗传组、蛋白质组等多组学分析方法，阐明CREB对人干细胞稳态及衰老的调控作用及其分子机制，进而为临床干预衰老或者衰老相关疾病的发生提供理论基础。

当前，世界各国都面临着人口老龄化的严峻挑战。研究表明，干细胞的耗竭是机体衰老的重要因素之一。CREB在啮齿类和低等动物的能量代谢、神经发育、组织分化、寿命调控等诸多生物学过程中扮演着重要的角色，预示着其可能在人干细胞稳态维持及衰老调控中也发挥着重要的作用。因此，揭示CREB在人干细胞稳态维持中的作用及分子机制可以帮助我们进一步了解CREB在人细胞水平的功能和机制，从而为阐明CREB信号通路在人类发育、衰老和疾病中的作用提供线索。本项目中，我们首先利用CRISPR/Cas9基因编辑技术成功构建了CREB缺失的hESCs，并将其成功地定向诱导分化为hMSCs。通过对CREB缺失hMSCs的单克隆形成能力检测、SA-β-gal染色分析、衰老相关基因表达检测，我们发现CREB缺失的hMSCs表现出了衰老相关的缺陷。同时我们也将CREB缺失的hESCs成功分化为hNSCs，通过对CREB缺失的hNeurons的特异性标志物 Tuj1、MAP2以及hNSC的标志物PAX6和Nestin的免疫荧光染色发现，CREB缺失的hNSCs无显著的衰老相关缺陷，同时CREB的缺失对端脑类器官的分化也没有显著性的影响。未来，我们将进一步探究CREB对人干细胞稳态及衰老的调控作用及其分子机制，进而为临床干预衰老或者衰老相关疾病的发生提供理论基础。