Yuan Cheng, Ni Lihua, Zhang Changjiang, Xia Hao, Wu Xiaoyan
Department of Gynecological Oncology, Zhongnan Hospital of Wuhan University, Wuhan, China.
Department of Nephrology, Zhongnan Hospital of Wuhan University, Wuhan, China.
Cardiovasc Diagn Ther. 2020 Aug;10(4):778-785. doi: 10.21037/cdt-20-299.
Previous study revealed that high glucose (HG) induced endothelial cell (EC) damage via endothelial-to-mesenchymal transition (EndMT). Recent studies suggested the role of Ephrin B2 in mediate ECs damage. However, the underlying mechanism remains unclear. The aim of the present study was to investigate whether Ephrin B2 mediates HG-induced EndMT in human aortic ECs (HAECs) and to determine the possible downstream signaling effector.
Primary HAECs were exposed to normal glucose (NG, 5.5 mM), HG (30 mM) and HG+Ephrin B2 small interfering RNA (siRNA), respectively. The pathological changes were investigated by light microscope and confocal microscopy. To study the effects of focal adhesion kinase (FAK) activation on Ephrin B2 in HAECs, cells were incubated with FAK siRNA in HG group. The expression of EndMT-related markers (CD31 and FSP1), Ephrin B2 and FAK were detected by qRT-PCR and western blot.
The results showed that HG significantly inhibited the expression of CD31 and increased FSP1 compared with NG group. Moreover, Ephrin B2 was increased after HG incubation. Ephrin B2 siRNA attenuated HG-induced expression of EndMT-related markers. Furthermore, HG increased the expression of FAK and phosphorylated FAK (pho-FAK) in HAECs. In contrast, blocking Ephrin B2 could partially attenuate HG-induced FAK activation. And FAK siRNA further inhibited the EndMT-related markers in HAECs treated with HG.
HG-induced EndMT in HAECs might be partially mediated by Ephrin B2 and the downstream FAK pathway.
先前的研究表明，高糖（HG）通过内皮-间充质转化（EndMT）诱导内皮细胞（EC）损伤。近期研究提示Ephrin B2在介导EC损伤中发挥作用。然而，其潜在机制仍不清楚。本研究旨在探讨Ephrin B2是否介导HG诱导的人主动脉内皮细胞（HAECs）的EndMT，并确定可能的下游信号效应器。
原代HAECs分别暴露于正常葡萄糖（NG，5.5 mM）、HG（30 mM）和HG+Ephrin B2小干扰RNA（siRNA）。通过光学显微镜和共聚焦显微镜观察病理变化。为研究黏着斑激酶（FAK）激活对HAECs中Ephrin B2的影响，在HG组中用FAK siRNA孵育细胞。通过qRT-PCR和蛋白质印迹法检测EndMT相关标志物（CD31和FSP1）、Ephrin B2和FAK的表达。
结果显示，与NG组相比，HG显著抑制CD31的表达并增加FSP1的表达。此外，HG孵育后Ephrin B2增加。Ephrin B2 siRNA减弱了HG诱导的EndMT相关标志物的表达。此外，HG增加了HAECs中FAK和磷酸化FAK（pho-FAK）的表达。相反，阻断Ephrin B2可部分减弱HG诱导的FAK激活。并且FAK siRNA进一步抑制了HG处理的HAECs中的EndMT相关标志物。
HG诱导的HAECs的EndMT可能部分由Ephrin B2及其下游的FAK途径介导。
