PDF(Pubmed)
Abstract：
BACKGROUND: Neural stem cell (NSC) proliferation and differentiation in the mammalian brain decreases to minimal levels postnatally. Nevertheless, neurogenic niches persist in the adult cortex and hippocampus in rodents, primates and humans, with adult NSC differentiation sharing key regulatory mechanisms with development. Adult neurogenesis impairments have been linked to Alzheimer\'s disease (AD) pathology. Addressing these impairments by using neurotrophic factors is a promising new avenue for therapeutic intervention based on neurogenesis. However, this possibility has been hindered by technical difficulties of using in-vivo models to conduct screens, including working with scarce NSCs in the adult brain and differences between human and mouse models or ethical limitations.
METHODS: Here, we use a combination of mouse and human stem cell models for comprehensive in-vitro characterization of a novel neurogenic compound, focusing on the brain-derived neurotrophic factor (BDNF) pathway. The ability of ENT-A011, a steroidal dehydroepiandrosterone derivative, to activate the tyrosine receptor kinase B (TrkB) receptor was tested through western blotting in NIH-3T3 cells and its neurogenic and neuroprotective action were assessed through proliferation, cell death and Amyloid-β (Aβ) toxicity assays in mouse primary adult hippocampal NSCs, mouse embryonic cortical NSCs and neural progenitor cells (NPCs) differentiated from three human induced pluripotent stem cell lines from healthy and AD donors. RNA-seq profiling was used to assess if the compound acts through the same gene network as BDNF in human NPCs.
RESULTS: ENT-A011 was able to increase proliferation of mouse primary adult hippocampal NSCs and embryonic cortical NSCs, in the absence of EGF/FGF, while reducing Aβ-induced cell death, acting selectively through TrkB activation. The compound was able to increase astrocytic gene markers involved in NSC maintenance, protect hippocampal neurons from Αβ toxicity and prevent synapse loss after Aβ treatment. ENT-A011 successfully induces proliferation and prevents cell death after Aβ toxicity in human NPCs, acting through a core gene network shared with BDNF as shown through RNA-seq.
CONCLUSIONS: Our work characterizes a novel BDNF mimetic with preferable pharmacological properties and neurogenic and neuroprotective actions in Alzheimer\'s disease via stem cell-based screening, demonstrating the promise of stem cell systems for short-listing competitive candidates for further testing.
METHODS: Here, we use a combination of mouse and human stem cell models for comprehensive in-vitro characterization of a novel neurogenic compound, focusing on the brain-derived neurotrophic factor (BDNF) pathway. The ability of ENT-A011, a steroidal dehydroepiandrosterone derivative, to activate the tyrosine receptor kinase B (TrkB) receptor was tested through western blotting in NIH-3T3 cells and its neurogenic and neuroprotective action were assessed through proliferation, cell death and Amyloid-β (Aβ) toxicity assays in mouse primary adult hippocampal NSCs, mouse embryonic cortical NSCs and neural progenitor cells (NPCs) differentiated from three human induced pluripotent stem cell lines from healthy and AD donors. RNA-seq profiling was used to assess if the compound acts through the same gene network as BDNF in human NPCs.
RESULTS: ENT-A011 was able to increase proliferation of mouse primary adult hippocampal NSCs and embryonic cortical NSCs, in the absence of EGF/FGF, while reducing Aβ-induced cell death, acting selectively through TrkB activation. The compound was able to increase astrocytic gene markers involved in NSC maintenance, protect hippocampal neurons from Αβ toxicity and prevent synapse loss after Aβ treatment. ENT-A011 successfully induces proliferation and prevents cell death after Aβ toxicity in human NPCs, acting through a core gene network shared with BDNF as shown through RNA-seq.
CONCLUSIONS: Our work characterizes a novel BDNF mimetic with preferable pharmacological properties and neurogenic and neuroprotective actions in Alzheimer\'s disease via stem cell-based screening, demonstrating the promise of stem cell systems for short-listing competitive candidates for further testing.
摘要：
背景：哺乳动物脑中神经干细胞（NSC）的增殖和分化在出生后降低到最低水平。然而，在啮齿动物的成年皮质和海马中存在神经源性生态位，灵长类动物和人类,成人NSC分化与发展共享关键的监管机制。成人神经发生损伤与阿尔茨海默病(AD)病理有关。通过使用神经营养因子来解决这些损伤是基于神经发生的治疗干预的有希望的新途径。然而,使用体内模型进行筛查的技术困难阻碍了这种可能性，包括使用成人大脑中稀缺的神经干细胞以及人类和小鼠模型之间的差异或伦理限制。
方法：这里，我们使用小鼠和人类干细胞模型的组合，对一种新的神经源性化合物进行全面的体外表征，关注脑源性神经营养因子（BDNF）通路。ENT-A011，一种甾体脱氢表雄酮衍生物，通过蛋白质印迹在NIH-3T3细胞中测试酪氨酸受体激酶B（TrkB）受体的激活，并通过增殖评估其神经源性和神经保护作用，小鼠原代成年海马神经干细胞的细胞死亡和淀粉样β（Aβ）毒性测定，小鼠胚胎皮质神经干细胞和神经祖细胞（NPC）从三个人诱导多能干细胞系从健康和AD供体分化。RNA-seq谱分析用于评估化合物是否通过与人NPC中的BDNF相同的基因网络起作用。
结果：ENT-A011能够增加小鼠原代成年海马神经干细胞和胚胎皮质神经干细胞的增殖，在没有EGF/FGF的情况下，同时减少Aβ诱导的细胞死亡，有选择地通过TrkB激活。该化合物能够增加参与NSC维持的星形细胞基因标记，保护海马神经元免受Αβ毒性并防止Aβ治疗后的突触丢失。ENT-A011成功诱导增殖并防止人NPC中Aβ毒性后的细胞死亡，通过与BDNF共享的核心基因网络起作用，如通过RNA-seq所示。
结论：我们的工作表征了一种新型BDNF模拟物，通过基于干细胞的筛选，在阿尔茨海默病中具有较好的药理特性和神经源性和神经保护作用。证明干细胞系统有望入围竞争候选人进行进一步测试。
方法：这里，我们使用小鼠和人类干细胞模型的组合，对一种新的神经源性化合物进行全面的体外表征，关注脑源性神经营养因子（BDNF）通路。ENT-A011，一种甾体脱氢表雄酮衍生物，通过蛋白质印迹在NIH-3T3细胞中测试酪氨酸受体激酶B（TrkB）受体的激活，并通过增殖评估其神经源性和神经保护作用，小鼠原代成年海马神经干细胞的细胞死亡和淀粉样β（Aβ）毒性测定，小鼠胚胎皮质神经干细胞和神经祖细胞（NPC）从三个人诱导多能干细胞系从健康和AD供体分化。RNA-seq谱分析用于评估化合物是否通过与人NPC中的BDNF相同的基因网络起作用。
结果：ENT-A011能够增加小鼠原代成年海马神经干细胞和胚胎皮质神经干细胞的增殖，在没有EGF/FGF的情况下，同时减少Aβ诱导的细胞死亡，有选择地通过TrkB激活。该化合物能够增加参与NSC维持的星形细胞基因标记，保护海马神经元免受Αβ毒性并防止Aβ治疗后的突触丢失。ENT-A011成功诱导增殖并防止人NPC中Aβ毒性后的细胞死亡，通过与BDNF共享的核心基因网络起作用，如通过RNA-seq所示。
结论：我们的工作表征了一种新型BDNF模拟物，通过基于干细胞的筛选，在阿尔茨海默病中具有较好的药理特性和神经源性和神经保护作用。证明干细胞系统有望入围竞争候选人进行进一步测试。
