Hematology Department, AP-HP, Georges Pompidou European Hospital, Paris, France; Université Paris-Cité, INSERM UMR-S 1140, Innovative Therapies in Haemostasis, Paris, France.
Université Paris-Cité, INSERM UMR-S 1140, Innovative Therapies in Haemostasis, Paris, France.
J Thromb Haemost. 2023 Dec;21(12):3640-3648. doi: 10.1016/j.jtha.2023.08.029. Epub 2023 Sep 9.
The involvement of thrombin receptor PAR1 in blood vessel development has been largely demonstrated in knockout mice; however, its implication in adult mouse angiogenesis seems very moderate.
We aimed to explore the potential relationships between PAR1, stemness, and angiogenic properties of human endothelial colony-forming cells (ECFCs).
PAR1 activation on ECFCs using the selective PAR1-activating peptide induced a significant decrease in CD133 expression (RTQ-PCR analysis). In line, silencing of PAR1 gene expression with siRNA increased CD133 mRNA as well as intracellular CD133 protein expression. To confirm the link between CD133 and PAR1, we explored the association between PAR1 and CD133 levels in fast and slow fibroblasts prone to reprogramming. An imbalance between PAR1 and CD133 levels was evidenced, with a decreased expression of PAR1 in fast reprogramming fibroblasts expressing a high CD133 level. Regarding in vitro ECFC angiogenic properties, PAR1 silencing with specific siRNA induced cell proliferation evidenced by the overexpression of Ki67. However, it did not impact migration properties nor ECFC adhesion on smooth muscle cells or human arterial endothelial cells. In a mouse model of hind-limb ischemia, PAR1 silencing in ECFCs significantly increased postischemic revascularization compared to siCtrl-ECFCs along with a significant increase in cutaneous blood flows (P < .0001), microvessel density (P = .02), myofiber regeneration (P < .0001), and human endothelial cell incorporation in muscle (P < .0001).
In conclusion, our work describes for the first time a link between PAR1, stemness, and vasculogenesis in human ECFCs.
血栓酶受体 PAR1 在血管发育中的作用已在基因敲除小鼠中得到广泛证实；然而，其在成年小鼠血管生成中的作用似乎非常有限。
我们旨在探讨 PAR1 与人类内皮祖细胞（ECFC）的干性和血管生成特性之间的潜在关系。
用选择性 PAR1 激活肽激活 ECFC 上的 PAR1，导致 CD133 表达显著下降（RTQ-PCR 分析）。与此一致，用 siRNA 沉默 PAR1 基因表达会增加 CD133 mRNA 以及细胞内 CD133 蛋白表达。为了证实 CD133 和 PAR1 之间的联系，我们研究了快速和慢速成纤维细胞中 PAR1 和 CD133 水平之间的关联，这些成纤维细胞容易重编程。证据表明 PAR1 和 CD133 水平之间存在不平衡，快速重编程的成纤维细胞中 PAR1 的表达降低，而 CD133 水平较高。关于体外 ECFC 的血管生成特性，用特异性 siRNA 沉默 PAR1 会诱导细胞增殖，这表现为 Ki67 的过度表达。然而，它不会影响迁移特性，也不会影响 ECFC 对平滑肌细胞或人动脉内皮细胞的黏附。在小鼠后肢缺血模型中，与 siCtrl-ECFC 相比，PAR1 在 ECFC 中的沉默显著增加了缺血后的再血管化，同时皮肤血流量（P <.0001）、微血管密度（P =.02）、肌纤维再生（P <.0001）和肌肉中人类内皮细胞的掺入（P <.0001）也显著增加。
总之，我们的工作首次描述了 PAR1、干性和人类 ECFC 血管生成之间的联系。
