Transcription profiling by array of forebrain from mice expressing VEGF120 and deficient for VEGF164

This work was designed to determine the role of the vascular endothelial growth factor A (VEGF) isoforms during early neuroepithelial development in the mammalian central nervous system (CNS), specifically in the forebrain. An emerging model of interdependence between neural and vascular systems includes VEGF, with its dual roles as a potent angiogenesis factor and neural regulator. Although a number of studies have implicated VEGF in CNS development, little is known about the role that the different VEGF isoforms play in early neurogenesis. We used a mouse model of disrupted VEGF isoform expression that eliminates the predominant brain isoform, VEGF164, and expresses only the diffusible form, VEGF120. We tested the hypothesis that VEGF164 plays a key role in controlling neural precursor populations in developing cortex. We used microarray analysis to compare gene expression differences between wild type and VEGF120 mice at E9.5, the primitive stem cell stage of the neuroepithelium. We quantified changes in PHH3-positive nuclei, neural stem cell markers (Pax6 and nestin) and the Tbr2-positive intermediate progenitors at E11.5 when the neural precursor population is expanding rapidly. Absence of VEGF164 (and VEGF188) leads to reduced proliferation without an apparent effect on the number of Tbr2-positive cells. There is a corresponding reduction in the number of mitotic spindles that are oriented parallel to the ventricular surface relative to those with a vertical or oblique angle. These results support a role for the VEGF isoforms in supporting the neural precursor population of the early neuroepithelium. Four samples each of E9.5 wildtype or VEGF120 mouse forebrain were analyzed with the Mouse 430 2.0 Affymetrix GeneChip本研究旨在明确血管内皮生长因子A（vascular endothelial growth factor A, VEGF）亚型在哺乳动物中枢神经系统（central nervous system, CNS），尤其是前脑的早期神经上皮发育过程中的作用。神经与血管系统间相互依存的新兴研究模型中包含VEGF，其兼具强效血管生成因子与神经调节因子的双重功能。尽管已有多项研究证实VEGF参与中枢神经系统发育，但目前对于不同VEGF亚型在早期神经发生中所发挥的具体作用仍知之甚少。本研究使用了VEGF亚型表达紊乱的小鼠模型，该模型剔除了大脑中主要的VEGF亚型VEGF164，仅表达可扩散型亚型VEGF120。我们提出假说：VEGF164在调控发育中大脑皮层的神经前体细胞群中发挥关键作用。本研究采用微阵列分析技术，对比了胚胎发育第9.5天（E9.5，神经上皮的原始干细胞阶段）野生型与VEGF120小鼠的基因表达差异。同时，我们定量分析了E11.5时期（神经前体细胞群快速扩增阶段）PHH3阳性细胞核、神经干细胞标志物（Pax6与巢蛋白（nestin））以及Tbr2阳性中间前体细胞的数量变化。实验结果显示，VEGF164（及VEGF188）缺失会导致细胞增殖能力下降，但对Tbr2阳性细胞的数量无明显影响。与垂直或倾斜角度的纺锤体相比，平行于脑室表面的有丝分裂纺锤体数量相应减少。上述结果证实，VEGF亚型在维持早期神经上皮的神经前体细胞群中具有重要作用。本研究共采集4份E9.5野生型小鼠前脑样本与4份E9.5 VEGF120小鼠前脑样本，采用Affymetrix Mouse 430 2.0基因芯片完成检测分析。