Endothelial Cell Cycle State and Cell Fate

建立有功能的血管网络是胚胎发育的限速步骤，修复 受伤的组织，以及组织替换的工程。尽管我们在辨认 促进内皮细胞增殖和发芽的因素，我们缺乏对如何适当控制的了解 血管重塑过程中内皮细胞的生长和表型特化，这造成了显著的 这是临床治疗、组织工程和再生医学的障碍。尽管有多个信令 动脉-静脉网络形成的调节涉及多种途径，包括血流诱导。 机械转导与Notch信号--这些信号协调调节的机制 内皮细胞的生长抑制和身份鉴定尚不清楚。我们最近的研究表明，重塑 血管丛受制于全身血液循环，不同大小的剪应力促进 不同的生长反应和基因表达。也就是说，动脉/小动脉切应力水平促进了Notch 信号，以及下游p27诱导的晚期G1期停滞，使动脉基因表达(Fang 2017)。 相反，静脉/小静脉典型的流量大小会导致早期G1期停滞，并使静脉上调。 基因。有趣的是，出生后动脉和静脉似乎维持着不同的内皮细胞周期状态。 我们对细胞周期控制在决定内皮细胞命运或差异信号中的作用知之甚少。 血管特定流量所诱导的通路，以及它们如何协调地诱导和维持 内皮细胞周期状态和特性。我们研究的科学前提是内皮细胞周期 适当的动脉和静脉规范需要控制，这样当内皮细胞处于不同的 在细胞周期状态下，它们表现出不同的动脉和静脉基因表达倾向。支持这一想法 来自对胚胎干细胞的研究，表明处于早期和晚期G1期的细胞有 中胚层/内胚层vs.外胚层命运(Paulkin 2014)。因此，我们的假设是这种差异 动脉和静脉中的流动力诱导不同的细胞内信号通路，促进不同的 内皮细胞周期状态，为动脉与血管的调节创造了不同的机会窗口。 静脉基因表达。为了确保科学的严谨性，我们将在体内动脉模型中测试这一假说。 静脉网络的形成和修复，以及在体外允许血流的人内皮细胞培养系统 操纵。我们将定义血管特异性流量调节内皮细胞的机制 周期状态，决定不同的内皮细胞周期状态如何实现不同的表型特化 (动脉与静脉)，并确定对内皮细胞周期状态的操纵是否可以预防或纠正 动-静脉畸形和加强损伤后的血管修复。对这一假设的评估将产生 关于血管形成和再生的新的基本见解，可以用来创造人类 体外微血管和治疗血管病变。

Establishing a functional vascular network is a rate-limiting step in embryonic development, the repair of injured tissues, and the engineering of tissue replacements. Although we have made progress in identifying factors that promote endothelial cell proliferation and sprouting, we lack understanding of how to properly control endothelial cell growth and phenotypic specialization during vascular remodeling, which has created a significant roadblock for clinical therapies, tissue engineering and regenerative medicine. Although multiple signaling pathways have been implicated in the regulation of arterial-venous network formation, including flow-induced mechanotransduction and Notch signaling, the mechanisms by which these signals coordinately regulate endothelial cell growth suppression and identity were unclear. Our recent studies revealed that remodeling vascular plexi are subject to systemic blood circulation, and that shear stress of different magnitudes promotes differential growth responses and gene expression. That is, arterial/arteriolar shear stress levels promote Notch signaling, and downstream p27-induced late G1 phase arrest that enables arterial gene expression (Fang 2017). Conversely, flow magnitudes typical of veins/venules induce early G1 arrest, and enables upregulation of venous genes. Interestingly, distinct endothelial cell cycle states appear to be maintained in arteries vs. veins postnatally. We know very little about the role of cell cycle control in endothelial cell fate decisions, or the differential signaling pathways induced by vessel-specific flow magnitudes, and how they may coordinately induce and maintain endothelial cell cycle state and identity. The scientific premise of our research is that endothelial cell cycle control is required for proper arterial and venous specification, such that when endothelial cells are in different cell cycle states, they exhibit different propensity for arterial vs. venous gene expression. Support for this idea comes from studies in embryonic stem cells that show cells in early vs. late G1 phase have a propensity for mesoderm/endoderm vs. ectoderm fate, respectively (Paulkin 2014). Thus, our hypothesis is that differential flow forces in arteries and veins induce different intracellular signaling pathways that promote distinct endothelial cell cycle states, creating distinct windows of opportunity for the regulation of arterial vs. venous gene expression. To ensure scientific rigor, we will test this hypothesis in vivo in models of arterial- venous network formation and repair, and in vitro in human endothelial cell culture systems that allow flow manipulation. We will define mechanisms by which vessel-specific flow magnitudes modulate endothelial cell cycle state, determine how distinct endothelial cell cycle states enable differential phenotypic specialization (artery vs. vein), and determine whether manipulation of endothelial cell cycle state can prevent or correct arterial-venous malformations and enhance post-injury vascular repair. Evaluation of this hypothesis will yield novel fundamental insights into blood vessel formation and regeneration that can be used to create human microvasculature ex vivo and treat vascular pathologies.