Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
J Cell Physiol. 2020 Sep;235(9):6204-6217. doi: 10.1002/jcp.29549. Epub 2020 Feb 3.
Vascular resident endothelial progenitor cells (VR-EPCs) have a certain ability to differentiate into endothelial cells (ECs) and participate in the process of angiogenesis. Glycolysis and mitochondrial fission and fusion play a pivotal role in angiogenesis. Pyruvate kinase muscle isoenzyme 2 (PKM2), which mediates energy metabolism and mitochondrial morphology, is regarded as the focus of VR-EPCs angiogenesis in our study. VR-EPCs were isolated from the hearts of 12-weeks-old Sprague-Dawley rats. The role of PKM2 on angiogenesis was evaluated by tube formation assay, wound healing assay, transwell assay, and chick chorioallantoic membrane assay. Western blot analysis, flow cytometry, mitochondrial membrane potential detection, reactive oxygen species (ROS) detection, immunofluorescence staining, and quantitative real-time polymerase chain reaction were used to investigate the potential mechanism of PKM2 for regulating VR-EPCs angiogenesis. We explored the function of PKM2 on the angiogenesis of VR-EPCs. DASA-58 (the activator of PKM2) promoted VR-EPCs proliferation and PKM2 activity, it also could promote angiogenic differentiation. At the same time, DASA-58 significantly enhanced glycolysis, mitochondrial fusion, slightly increased mitochondrial membrane potential, and maintained ROS at a low level. C3k, an inhibitor of PKM2, inhibited PKM2 activity, expression of angiogenesis-related genes and tube formation. Besides, C3k drastically reduced glycolysis and mitochondrial membrane potential while significantly promoting mitochondrial fission and ROS level. Activation of PKM2 could promote VR-EPCs angiogenesis through modulating glycolysis, mitochondrial fission and fusion. By contrast, PKM2 inhibitor has opposite effects.
血管驻留内皮祖细胞 (VR-EPCs) 具有一定的分化为内皮细胞 (ECs) 的能力，并参与血管生成过程。糖酵解和线粒体分裂与融合在血管生成中起着关键作用。丙酮酸激酶肌肉同工酶 2 (PKM2) 介导能量代谢和线粒体形态，被认为是我们研究中 VR-EPCs 血管生成的焦点。从 12 周龄 Sprague-Dawley 大鼠心脏中分离出 VR-EPCs。通过管形成试验、划痕愈合试验、Transwell 试验和鸡胚绒毛尿囊膜试验评估 PKM2 对血管生成的作用。Western blot 分析、流式细胞术、线粒体膜电位检测、活性氧 (ROS) 检测、免疫荧光染色和实时定量聚合酶链反应用于研究 PKM2 调节 VR-EPCs 血管生成的潜在机制。我们探讨了 PKM2 对 VR-EPCs 血管生成的功能。DASA-58（PKM2 的激活剂）促进 VR-EPCs 的增殖和 PKM2 活性，也能促进血管生成分化。同时，DASA-58 显著增强糖酵解和线粒体融合，略微增加线粒体膜电位，并将 ROS 维持在低水平。PKM2 的抑制剂 C3k 抑制 PKM2 活性、血管生成相关基因的表达和管形成。此外，C3k 显著降低糖酵解和线粒体膜电位，同时显著促进线粒体分裂和 ROS 水平。激活 PKM2 可通过调节糖酵解、线粒体分裂与融合来促进 VR-EPCs 血管生成。相比之下，PKM2 抑制剂则有相反的效果。
