Stem Cell Institute, KU Leuven, 3000 Leuven, Belgium; Stem Cell Biology and Embryology Unit, Department of Development and Regeneration, KU Leuven, 3000 Leuven, Belgium.
Leuven Research Institute for Neuroscience and Disease (LIND), KU Leuven, 3000 Leuven, Belgium; Research Group Experimental Neurology, Department of Neurosciences, KU Leuven, 3000 Leuven, Belgium; Laboratory of Neurobiology, VIB Vesalius Research Center, 3000 Leuven, Belgium.
Stem Cell Reports. 2015 Jan 13;4(1):16-24. doi: 10.1016/j.stemcr.2014.12.001. Epub 2014 Dec 31.
To understand how haploinsufficiency of progranulin (PGRN) causes frontotemporal dementia (FTD), we created induced pluripotent stem cells (iPSCs) from patients carrying the GRN(IVS1+5G > C) mutation (FTD-iPSCs). FTD-iPSCs were fated to cortical neurons, the cells most affected in FTD. Although generation of neuroprogenitors was unaffected, their further differentiation into CTIP2-, FOXP2-, or TBR1-TUJ1 double-positive cortical neurons, but not motorneurons, was significantly decreased in FTD-neural progeny. Zinc finger nuclease-mediated introduction of GRN cDNA into the AAVS1 locus corrected defects in cortical neurogenesis, demonstrating that PGRN haploinsufficiency causes inefficient cortical neuron generation. RNA sequencing analysis confirmed reversal of the altered gene expression profile following genetic correction. We identified the Wnt signaling pathway as one of the top defective pathways in FTD-iPSC-derived neurons, which was reversed following genetic correction. Differentiation of FTD-iPSCs in the presence of a WNT inhibitor mitigated defective corticogenesis. Therefore, we demonstrate that PGRN haploinsufficiency hampers corticogenesis in vitro.
为了了解颗粒体蛋白前体（PGRN）单倍不足如何导致额颞叶痴呆（FTD），我们从携带 GRN(IVS1+5G > C)突变的患者中创建了诱导多能干细胞（iPSC）（FTD-iPSC）。FTD-iPSC 被定向为皮质神经元，这是 FTD 中受影响最严重的细胞。尽管神经祖细胞的生成不受影响，但它们进一步分化为 CTIP2-、FOXP2- 或 TBR1-TUJ1 双阳性皮质神经元，但不是运动神经元，在 FTD 神经祖细胞中显著减少。锌指核酸酶介导的将 GRN cDNA 引入 AAVS1 基因座纠正了皮质神经发生的缺陷，证明 PGRN 单倍不足导致皮质神经元生成效率低下。RNA 测序分析证实了遗传纠正后改变的基因表达谱的逆转。我们确定 Wnt 信号通路是 FTD-iPSC 衍生神经元中缺陷最多的通路之一，该通路在遗传纠正后得到逆转。在存在 WNT 抑制剂的情况下分化 FTD-iPSC 可减轻皮质生成缺陷。因此，我们证明 PGRN 单倍不足会阻碍体外皮质发生。
