Neural patterning of human induced pluripotent stem cells in 3-D cultures for studying biomolecule-directed differential cellular responses.
Neural patterning of human induced pluripotent stem cells in 3-D cultures for studying biomolecule-directed differential cellular responses.
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DOI：
10.1016/j.actbio.2016.06.027
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发表时间：
2016-09
影响因子：
9.7
通讯作者：
Yuanwei Yan;Julie Bejoy;Junfei Xia;J. Guan;Yi Zhou;Yan Li
中科院分区：
文献类型：
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作者：
Yuanwei Yan;Julie Bejoy;Junfei Xia;J. Guan;Yi Zhou;Yan Li
适当的人类诱导多能干细胞（hiPSCs）的神经模式对于产生特定的神经细胞/组织甚至是与模拟神经系统疾病生理相关的迷你大脑至关重要。然而，体外调节三维神经组织模式的信号因子的能力以及由此产生的神经群体对各种生物分子的差异反应尚未完全了解。方法通过使用靶向sonic hedgehog （SHH）信号的小分子调节hipsc的神经模式，本研究生成了主要由皮质谷氨酸能神经元或运动神经元组成的不同3d神经元培养物。结果经SHH信号拮抗剂环巴胺处理后，谷氨酸能神经元丰富，而经SHH信号拮抗剂氨基吗啡处理后，表达胰岛-1和hb9的运动神经元丰富。在来自不同神经模式因子的神经元中，全细胞膜片钳记录显示相似的电压门控Na+/K+电流、去极化诱发动作电位和自发兴奋突触后电流。此外，这些不同的神经元群体对三类生物分子表现出不同的反应，包括(1)影响细胞外基质重塑的基质金属蛋白酶抑制剂；(2) n -甲基-d-天冬氨酸引起全身神经毒性；(3)引起神经元亚型特异性神经毒性的β淀粉样蛋白（1-42）低聚物。结论本研究将促进我们对hiPSC自组织和神经组织发育的理解，并为神经疾病建模和药物开发提供一种革命性的3d模型建立方法。适当的人类诱导多能干细胞（hiPSCs）的神经模式对于生成与模拟神经系统疾病生理相关的特定神经细胞、组织甚至迷你大脑至关重要。然而，与音刺猬相关的小分子在hipsc的三维分化中调节不同神经元亚型的能力以及区域特异性神经元亚型对各种生物分子的差异细胞反应尚未得到充分研究。通过使用靶向超音刺猬信号的小分子调节hipsc的神经模式，本研究提供了hipsc衍生的区域特异性神经元亚型在细胞外基质重塑和神经毒性中对不同生物分子的差异敏感性的知识。这一发现对于理解hipsc的三维神经模式在脑类器官形成、神经系统疾病建模和药物发现中的应用具有重要意义。
IntroductionAppropriate neural patterning of human induced pluripotent stem cells (hiPSCs) is critical to generate specific neural cells/tissues and even mini-brains that are physiologically relevant to model neurological diseases. However, the capacity of signaling factors that regulate 3-D neural tissue patterningin vitroand differential responses of the resulting neural populations to various biomolecules have not yet been fully understood.MethodsBy tuning neural patterning of hiPSCs with small molecules targeting sonic hedgehog (SHH) signaling, this study generated different 3-D neuronal cultures that were mainly comprised of either cortical glutamatergic neurons or motor neurons.ResultsAbundant glutamatergic neurons were observed following the treatment with an antagonist of SHH signaling, cyclopamine, while Islet-1 and HB9-expressing motor neurons were enriched by an SHH agonist, purmorphamine. In neurons derived with different neural patterning factors, whole-cell patch clamp recordings showed similar voltage-gated Na+/K+currents, depolarization-evoked action potentials and spontaneous excitatory post-synaptic currents. Moreover, these different neuronal populations exhibited differential responses to three classes of biomolecules, including (1) matrix metalloproteinase inhibitors that affect extracellular matrix remodeling; (2) N-methyl-d-aspartate that induces general neurotoxicity; and (3) amyloid β (1–42) oligomers that cause neuronal subtype-specific neurotoxicity.ConclusionsThis study should advance our understanding of hiPSC self-organization and neural tissue development and provide a transformative approach to establish 3-D models for neurological disease modeling and drug discovery.Statement of SignificanceAppropriate neural patterning of human induced pluripotent stem cells (hiPSCs) is critical to generate specific neural cells, tissues and even mini-brains that are physiologically relevant to model neurological diseases. However, the capability of sonic hedgehog-related small molecules to tune different neuronal subtypes in 3-D differentiation from hiPSCs and the differential cellular responses of region-specific neuronal subtypes to various biomolecules have not been fully investigated. By tuning neural patterning of hiPSCs with small molecules targeting sonic hedgehog signaling, this study provides knowledge on the differential susceptibility of region-specific neuronal subtypes derived from hiPSCs to different biomolecules in extracellular matrix remodeling and neurotoxicity. The findings are significant for understanding 3-D neural patterning of hiPSCs for the applications in brain organoid formation, neurological disease modeling, and drug discovery.