文章摘要
韩肖华 ，陈潞婷， ，李娟 ，李丹萍 ，陈红.2种不同诱导方法体外诱导人多潜能干细胞分化成大脑基底内侧神经节隆起区神经前体细胞的差异[J].神经损伤功能重建,2021,16(10):565-568
2种不同诱导方法体外诱导人多潜能干细胞分化成大脑基底内侧神经节隆起区神经前体细胞的差异
Comparison of Inducing Human Pluripotent Stem Cells to Differentiate into Medial GanglionicEminence Neural Progenitor Cells in vitro with Two Culture Methods
  
DOI：
中文关键词: 单层贴壁培养法  拟胚体悬浮培养法  人多潜能干细胞  大脑基底内侧神经节隆起区  神经前体细 胞  SAG浓度
英文关键词: monolayer adherent culture method  EB culture method  human pluripotent stem cells  medial ganglionic eminence  neural progenitor cells  concentrations of SAG
基金项目:国家自然科学基 金（No. 8177440 4）
作者单位
韩肖华1 ，陈潞婷1，2 ，李娟1 ，李丹萍1 ，陈红1 1. 华中科技大学 同济医学院附属 同济医院康复医 学科 2. 长江航运总医 院康复医学科 
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中文摘要:
      目的：比较单层贴壁培养法和拟胚体（EB）悬浮培养法诱导人多潜能干细胞分化为大脑基底内侧神经 节隆起区（MGE）神经前体细胞分化的效率，并探讨不同浓度的SHH通路激动剂SAG对分化为MGE神经前 体细胞纯度的影响，为细胞移植治疗探索优化的分化方案。方法：分别采用单层贴壁诱导法和EB悬浮法诱 导人多潜能干细胞分化为神经上皮细胞，通过加入不同浓度的SAG使其腹侧化，最终诱导成表达NKX2.1的 MGE前体细胞并进行计数。比较2种不同的分化方案和不同浓度的SAG对MGE神经前体细胞纯度的影 响。结果：在单层贴壁诱导法中，当SAG浓度为0 μM、0.1 μM、0.5 μM、1 μM、2 μM和3 μM时，NKX2.1阳性细 胞率分别为0%、（48.3±11.5）%、（78.6±2.7）%、（77.6±4.0）%、（69.0±7.5）%和（67.7±7.9）%；SAG浓度为0.5 μM和 1 μM 时诱导分化获得的 MGE 神经前体细胞纯度显著高于其他浓度组（P＜0.001 或 0.05）。当 SAG 浓度为 0.5 μM时，在EB悬浮培养法中NKX2.1的阳性细胞率为（74.8±6.5）%，与单层贴壁诱导法差异无统计学意义 （P＞0.05）。采用单层贴壁诱导法时，神经球和神经元出现的时间点均要早于采用EB悬浮培养法。结论：采 用单层贴壁诱导法，0.5 μM的SAG可能是人多潜能干细胞分化培养为MGE神经前体细胞的更优化方案。
英文摘要:
      To compare the different effects of inducing human pluripotent stem cells to differentiate into medial ganglionic eminence (MGE) neural progenitor cells with monolayer adherent culture and embryoid bodies (EB) culture methods and to explore the effect of different concentrations of SAG (SHH signaling agonist) on the purity of neural progenitor cells differentiated into MGE. Methods: Human pluripotent stem cells were differentiated into neuro epithelial cells using monolayer adherent culture and EB culture methods. Varying concentrations of SAG were used to induce ventralization. The cells were ultimately induced to become NKX2.1-expressing MGE neuro progenitor cells and then counted. The effect of the two culture methods and the different concentrations of SAG on the purity of MGE neural progenitor cells were investigated. Results: In the monolayer adherent culture method, the positive expression of NKX2.1 was 0% in 0 μM SAG, (48.3±11.5)% in 0.1 μM SAG, (78.6± 2.7)% in 0.5 μM SAG, (77.6±4.0)% in 1 μM SAG, (69.0±7.5)% in 2 μM SAG, and (67.7±7.9)% in 3 μM SAG. The purity of differentiated MGE neural progenitor cells induced by 0.5μM and 1μM SAG was superior to those induced by other concentrations (P<0.001 or 0.05). Using the EB culture method with 0.5 μM SAG, the positive expression of NKX2.1 was (74.8±6.5)% and showed no significant difference from the monolayer adherent culture method (P>0.05). Neurosphere and neuron formation occurred earlier when using the monolayer adherent culture method than when using the EB culture method. Conclusion: To obtain MGE neural progenitor cells from human pluripotent stem cells for transplantation treatment, the monolayer adherent culture method with 0.5 μM SAG may be optimal.
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