Table_1_Wnt/β-Catenin Signaling Promotes Differentiation of Ischemia-Activated Adult Neural Stem/Progenitor Cells to Neuronal Precursors.xlsx

Modulating endogenous regenerative processes may represent a suitable treatment for central nervous system (CNS) injuries, such as stroke or trauma. Neural stem/progenitor cells (NS/PCs), which naturally reside in the subventricular zone (SVZ) of the adult brain, proliferate and differentiate to other cell types, and therefore may compensate the negative consequences of ischemic injury. The fate of NS/PCs in the developing brain is largely influenced by Wingless/Integrated (Wnt) signaling; however, its role in the differentiation of adult NS/PCs under ischemic conditions is still enigmatic. In our previous study, we identified the Wnt/β-catenin signaling pathway as a factor promoting neurogenesis at the expense of gliogenesis in neonatal mice. In this study, we used adult transgenic mice in order to assess the impact of the canonical Wnt pathway modulation (inhibition or hyper-activation) on NS/PCs derived from the SVZ, and combined it with the middle cerebral artery occlusion (MCAO) to disclose the effect of focal cerebral ischemia (FCI). Based on the electrophysiological properties of cultured cells, we first identified three cell types that represented in vitro differentiated NS/PCs – astrocytes, neuron-like cells, and precursor cells. Following FCI, we detected fewer neuron-like cells after Wnt signaling inhibition. Furthermore, the immunohistochemical analysis revealed an overall higher expression of cell-type-specific proteins after FCI, indicating increased proliferation and differentiation rates of NS/PCs in the SVZ. Remarkably, Wnt signaling hyper-activation increased the abundance of proliferating and neuron-like cells, while Wnt pathway inhibition had the opposite effect. Finally, the expression profiling at the single cell level revealed an increased proportion of neural stem cells and neuroblasts after FCI. These observations indicate that Wnt signaling enhances NS/PCs-based regeneration in the adult mouse brain following FCI, and supports neuronal differentiation in the SVZ.

调节内源性再生过程可能成为治疗中枢神经系统（CNS）损伤，如中风或创伤的适宜治疗方案。神经干细胞/祖细胞（NS/PCs），这些细胞自然存在于成年大脑的室下区（SVZ），能够增殖并分化为其他细胞类型，因此可能补偿缺血性损伤的负面影响。发育中大脑中NS/PCs的命运在很大程度上受Wingless/Integrated（Wnt）信号通路的影响；然而，其在缺血条件下对成年NS/PCs分化的作用仍是一个谜。在我们之前的研究中，我们确定了Wnt/β-catenin信号通路是促进新生小鼠神经发生而非胶质发生的因素。在本研究中，我们使用了成年转基因小鼠，以评估经典Wnt通路调节（抑制或超活化）对SVZ来源的NS/PCs的影响，并将其与大脑中动脉闭塞（MCAO）相结合，以揭示局部脑缺血（FCI）的影响。基于培养细胞的电生理特性，我们首先确定了代表体外分化NS/PCs的三个细胞类型——星形胶质细胞、神经元样细胞和前体细胞。在FCI之后，我们检测到Wnt信号抑制后神经元样细胞数量减少。此外，免疫组化分析显示FCI后细胞类型特异性蛋白的整体表达更高，这表明SVZ中NS/PCs的增殖和分化速率增加。值得注意的是，Wnt信号超活化增加了增殖和神经元样细胞的数量，而Wnt通路抑制则产生相反的效果。最后，单细胞水平的表达谱分析显示FCI后神经干细胞和神经母细胞的比例增加。这些观察结果表明，Wnt信号通路增强了成年小鼠脑中基于NS/PCs的再生，并支持SVZ中的神经元分化。