Design and evaluation of EphrinA1 mutants with cerebral protective effect.
Design and evaluation of EphrinA1 mutants with cerebral protective effect.
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EphrinA1脑保护作用突变体的设计与评价
DOI：
10.1038/s41598-017-02091-7
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发表时间：
2017-05-15
影响因子：
4.6
通讯作者：
Wang Y
中科院分区：
文献类型：
--
作者：
Zhu Y;Gao Y;Zheng D;Shui M;Yu K;Liu X;Lin Y;Su L;Yang W;Wang Y
EphA2受体被其天然配体EPhinA1激活导致血脑屏障功能障碍，而EphA2失活可减轻缺血性卒中的BBB损伤。因此，EphA2靶向拮抗剂可能作为神经保护剂。我们分别设计了四个突变体，分别是EPhinA1、EM1、EM2、EM3和EM4。计算分析表明，这四个突变体都能与EphA2相互作用。在小鼠局灶性脑缺血/再灌注(I/R)模型上观察其潜在的神经保护作用。EM2表现出强大的神经保护作用，包括缩小脑梗塞体积，神经元凋亡，脑水肿，并改善神经功能评分。EM2介导的保护作用与相对减少血脑屏障渗漏、炎性渗透以及紧密连接蛋白如闭锁带-1和闭锁蛋白的高表达有关。EM2可下调缺血再灌注后Rho相关蛋白激酶2(ROCK2)的高表达。此外，EM2还减少了激动剂多沙唑嗪诱导的PC3细胞EphA2的磷酸化和细胞圆形，表明EphA2拮抗EM2的活性。这些发现为EphA2拮抗剂的神经保护作用提供了证据，为缺血性卒中的治疗提供了新的途径。这些结果还表明，受体激动剂可以通过取代一个或多个相关残基而转换为拮抗剂。
The activation of EphA2 receptor by its natural ligand EphrinA1 causes blood brain barrier dysfunction, and inactivation of EphA2 reduces BBB damage in ischemic stroke. Thus, EphA2 targeted antagonists may serve as neuroprotective agents. We engineered four mutants of EphrinA1, EM1, EM2, EM3 and EM4, respectively. The computational analysis showed that these four mutants were capable of interacting with EphA2. Their potential neuroprotective effects were examined in mouse focal ischemia/reperfusion (I/R) model. EM2 exhibited strong neuroprotective effects, including reduced brain infarct volume, neuronal apoptosis, cerebral edema, and improved neurological scores. The EM2-mediated protection was associated with a comparative decrease in BBB leakage, inflammatory infiltration, and higher expression levels of tight junction proteins, such as zonula occludens-1 and Occludin. I/R-induced high expression of Rho-associated protein kinase 2 (ROCK2) was down-regulated after EM2 treatment. Moreover, EM2 reduced agonist doxazosin-induced EphA2 phosphorylation and cells rounding in PC3 cells, indicating EphA2-antagonizing activity of EM2. These finding provided evidences of the neuroprotection of EphA2 antagonist and a novel approach for ischemic stroke treatment. These results also suggested that a receptor agonist can be switched to an antagonist by substituting one or more relevant residues.