Laboratory of Experimental Neurology, Université Libre de Bruxelles (ULB), 1070, Brussels, Belgium.
Laboratory of Neurophysiology, Université Libre de Bruxelles (ULB), 1070, Brussels, Belgium.
Sci Rep. 2020 May 8;10(1):7752. doi: 10.1038/s41598-020-64831-6.
Human induced pluripotent stem cells (iPSCs) are used to generate models of human diseases that recapitulate the pathogenic process as it occurs in affected cells. Many differentiated cell types can currently be obtained from iPSCs, but no validated protocol is yet available to specifically generate primary proprioceptive neurons. Proprioceptors are affected in a number of genetic and acquired diseases, including Friedreich ataxia (FRDA). To develop a cell model that can be applied to conditions primarily affecting proprioceptors, we set up a protocol to differentiate iPSCs into primary proprioceptive neurons. We modified the dual-SMAD inhibition/WNT activation protocol, previously used to generate nociceptor-enriched cultures of primary sensory neurons from iPSCs, to favor instead the generation of proprioceptors. We succeeded in substantially enriching iPSC-derived primary sensory neuron cultures for proprioceptors, up to 50% of finally differentiated neurons, largely exceeding the proportion of 7.5% normally represented by these cells in dorsal root ganglia. We also showed that almost pure populations of proprioceptors can be purified from these cultures by fluorescence-activated cell sorting. Finally, we demonstrated that the protocol can be used to generate proprioceptors from iPSCs from FRDA patients, providing a cell model for this genetic sensory neuronopathy.
人诱导多能干细胞（iPSCs）用于生成人类疾病模型，这些模型再现了受影响细胞中发生的致病过程。目前可以从 iPSCs 中获得许多分化的细胞类型，但尚未有经过验证的方案可专门生成原发性本体感受神经元。本体感受器会受到多种遗传和获得性疾病的影响，包括弗里德里希共济失调（FRDA）。为了开发一种可应用于主要影响本体感受器的疾病的细胞模型，我们建立了一个将 iPSCs 分化为原发性本体感受神经元的方案。我们修改了双 SMAD 抑制/WNT 激活方案，该方案先前用于从 iPSCs 中生成富含伤害感受器的初级感觉神经元培养物，转而有利于生成本体感受器。我们成功地将 iPSC 衍生的初级感觉神经元培养物中本体感受器的含量大大富集，达到最终分化神经元的 50%，大大超过这些细胞在背根神经节中通常占 7.5%的比例。我们还表明，可以通过荧光激活细胞分选从这些培养物中纯化出几乎纯的本体感受器群体。最后，我们证明该方案可用于从 FRDA 患者的 iPSCs 中生成本体感受器，为这种遗传性感觉神经元病提供了一种细胞模型。
