VEGF/NRP1和Sema3s/Plexin-D1信号通过共受体交互调控脑梗死后新生血管极性和血管成熟

Angiogenesis is one of the most promising treatment targets for stroke. Our study found that compensatory angiogenesis exist in cerebral infarction area after rat pMCAO, but new blood vessels distribute disorderly and disappear gradually, maybe regulated by crosstalking of VEGF/NRP1 and Sema3s/Plexin-D1 signaling pathway through co-receptors. In present project, we will study the several aspects with immunofluorescence, western blot, RT-PCR, radionuclide imaging, and other methods. Firstly, we will explore the spatio-temporal dynamic characteristics of Sema3s/Plexin-D1 and VEGF/NRP1 signals. Secondly, we will detect the effect of dynamic VEGF/Sema3s ratio on controlling both tip cells and angiogenesis after stroke, we will also try to illuminate the detail molecular mechanisms. Thirdly, by regulating the key factors such as Sema3A, Sema3E and VEGF, we will determine the best therapeutic time point and targets to induce orderly and infarctional angiogenesis after stroke.

血管新生是脑缺血最有前途的治疗目标之一, 我们研究发现：大鼠脑梗死区域虽有自发的异常的血管新生，但分布无序且会消退, 与Sema3s/Plexin-D1信号和VEGF/NRP1信号的交互调控密切相关。本项目拟在invivo和invitro情况下，采用免疫荧光、免疫印迹、RT-PCR、核素显像等多种技术结合的试验方法，探讨脑梗死后缺血脑组织Sema3s/Plexin-D1信号和VEGF/NRP1信号的动态时空变化规律，彼此调控关系，确定VEGF/Sema3s比值时空动态变化规律及其对tip 细胞和血管新生的影响，讨论二者交互调控时空二维失衡后诱导在脑梗死后异常血管新生中的具体分子机制，并通过对其中关键分子如：Sema3A、Sema3E、VEGF等在不同时间点进行定向干预，以诱导缺血区域形成持续的血管新生，向梗死区域生长的血管极性，重建血运，为脑梗死的治疗提供新的靶点和理论依据。

缺血性脑梗塞后，脑组织存活和功能恢复很大程度上取决于缺血区的脑血流灌注水平和功能性侧支循环重建。轴突引导分子Semaphorin 3E（Sema3E）近年来被证实能够引导血管形成，并通过其受体PlexinD1与疾病进展密切相关，基因水平敲除Sema3E或其受体PlexinD1可促进血管生长。在研究中，我们发现用慢病毒介导的PlexinD1-shRNA可特异性阻断Sema3E / PlexinD1信号通路，从而明显改善缺血半暗带的组织存活和功能恢复。 Sema3E / PlexinD1抑制不仅增加了梗死侧大脑皮质的血流灌注，而且改善了脑梗塞后血脑屏障破坏。在机制方面，我们进一步证明了Sema3E可通过抑制DLL4/Notch信号通路抑制内皮细胞增殖和血管生成能力。更重要的是，Sema3E / PlexinD1信号通过减少内皮细胞分泌PDGF-BB从而抑制周细胞的募集。综上所述，我们的研究显示，在缺血半暗带中抑制Sema3E / PlexinD1信号传导（其增加内皮血管生成能力和周细胞募集）有助于脑梗塞后的功能性新血管形成和血脑屏障的完整性。我们的研究结果表明Sema3E / PlexinD1信号是改善缺血性脑卒中后脑组织存活和功能恢复的新型治疗靶点。