Vissapragada Ravi, Contreras Mauricio A, da Silva Cleide G, Kumar Vivek A, Ochoa Angelica, Vasudevan Anju, Selim Magdy H, Ferran Christiane, Thomas Ajith J
Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, United States of America.
Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, United States of America; Division of Vascular Surgery, Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, United States of America.
Brain Res. 2014 May 27;1565:8-17. doi: 10.1016/j.brainres.2014.03.018. Epub 2014 Mar 24.
Interactions between neural progenitor cells (NPC) and endothelial cells (EC) from adult vascular beds have been well explored previously. However, the factors and signaling mechanisms that regulate neurogenesis and angiogenesis are most prevalent during embryonic development. This study aimed to determine whether embryonic brain endothelial cells from the periventricular region (PVEC) present an advantage over adult brain EC in supporting NPC growth and differentiation. PVEC were isolated from E15 mouse brains, processed, and sorted with immunomagnetic beads using antibodies against CD31/PECAM. On immunofluorescence (IF) staining, nearly all cells were positive for EC markers CD31 and CD144/VE-Cadherin. In proliferation studies, NPC proliferation was highest in transwell co-culture with PVEC, approximately 2.3 fold increase compared to baseline versus 1.4 fold increase when co-cultured with adult brain endothelial cells (ABEC). These results correlated with the PVEC mediated delay in NPC differentiation, evidenced by high expression of progenitor marker Nestin evaluated by IF staining. Upon further characterization of PVEC in an angiogenesis assay measuring cord length, PVEC exhibited a high capacity to form cords in basal conditions compared to ABEC. This was enhanced in the presence of NPC, with both cell types displaying a preferential structural alignment resembling neurovascular networks. PVEC also expressed high Vegfa levels at baseline in comparison to NPC and ABEC. Vegfa levels increased when co-cultured with NPC. We demonstrate that PVEC and NPC co-cultures act synergistically to promote the formation of a neurovascular unit through dynamic and reciprocal communication. Our results suggest that PVEC/NPC could provide promising neuro-regenerative therapies for patients suffering brain injuries.
成年血管床中的神经祖细胞（NPC）与内皮细胞（EC）之间的相互作用此前已得到充分研究。然而，调节神经发生和血管生成的因子及信号机制在胚胎发育过程中最为普遍。本研究旨在确定来自脑室周围区域的胚胎脑内皮细胞（PVEC）在支持NPC生长和分化方面是否比成年脑EC具有优势。从E15小鼠脑中分离出PVEC，进行处理，并用抗CD31/PECAM抗体的免疫磁珠进行分选。免疫荧光（IF）染色显示，几乎所有细胞均为EC标志物CD31和CD144/VE-钙黏蛋白阳性。在增殖研究中，NPC与PVEC的Transwell共培养中增殖最高，与基线相比增加约2.3倍，而与成年脑内皮细胞（ABEC）共培养时增加1.4倍。这些结果与PVEC介导的NPC分化延迟相关，IF染色评估显示祖细胞标志物巢蛋白高表达证明了这一点。在测量索长度的血管生成试验中对PVEC进行进一步表征时，与ABEC相比，PVEC在基础条件下形成索的能力较高。在存在NPC的情况下这种能力增强，两种细胞类型均显示出类似于神经血管网络的优先结构排列。与NPC和ABEC相比，PVEC在基线时也表达高水平的Vegfa。与NPC共培养时，Vegfa水平升高。我们证明PVEC和NPC共培养通过动态和相互的通讯协同作用促进神经血管单元的形成。我们的结果表明，PVEC/NPC可为脑损伤患者提供有前景的神经再生疗法。
