Niemann-Pick disease type C1 (NPC1) is a rare progressive neurodegenerative disorder caused by mutations in the NPC1 gene, resulting mainly in the accumulation of cholesterol and the ganglioside GM2. Recently, we described accumulations of these lipids in neuronal differentiated cells derived from NPC1 patient-specific induced pluripotent stem cells (iPSCs). As these lipids are essential for proper cell membrane composition, we were interested in the expression and function of voltage-gated ion channels and excitatory AMPA receptors (AMPARs) in neurons derived from three patient-specific iPSC lines. By means of patch clamp recordings and microfluorimetric measurements of calcium (Ca2+), we examined the expression of voltage-gated ion channels and AMPARs. Cells of the three used cell lines carrying the c.1836A>C/c.1628delC, the c.1180T>C or the c.3182T>C mutation demonstrated a significantly reduced AMPA-induced Ca2+-influx, suggesting an altered expression profile of these receptors. RT-qPCR revealed a significant upregulation of mRNA for the AMPA receptor subunits GluA1 and GluA2 and western blot analysis showed increased protein level of GluA2. Thus, we conclude that the observed reduced Ca2+-influx is based on an increase of GluA2 containing Ca2+-impermeable AMPARs. An attenuated function of GluRs in neurons potentially contributes to the progressive neurodegeneration observed in NPC1 and might represent an objective in regard of the development of new therapeutic approaches in NPC1.

中文翻译：

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Niemann-Pick C1型患者特异性iPSC来源的神经元中钙通量的降低是由于钙不可渗透的AMPA受体数量增加所致。

Niemann-Pick疾病C1型（NPC1）是一种罕见的进行性神经退行性疾病，由NPC1基因的突变引起，主要导致胆固醇和神经节苷脂GM2的积累。最近，我们描述了这些脂质在NPC1患者特异性诱导多能干细胞（iPSC）衍生的神经元分化细胞中的积累。由于这些脂质对于适当的细胞膜组成至关重要，因此我们对电压门控离子通道和兴奋性AMPA受体（AMPAR）在源自三种患者特异性iPSC系的神经元中的表达和功能感兴趣。通过膜片钳记录和钙（Ca2 +）的微荧光测量，我们检查了电压门控离子通道和AMPAR的表达。三种使用过的细胞系中的细胞携带c.1836A> C / c.1628delC，c.1180T> C或c.3182T> C突变表明AMPA诱导的Ca2 +内流明显减少，表明这些受体的表达谱发生了改变。RT-qPCR显示AMPA受体亚基GluA1和GluA2的mRNA显着上调，蛋白质印迹分析显示GluA2的蛋白水平升高。因此，我们得出结论，观察到的减少的Ca2 +流入是基于含Ca2 +不可渗透AMPAR的GluA2的增加。GluRs在神经元中的功能减弱可能会导致在NPC1中观察到的进行性神经退行性变，并且可能代表了开发NPC1中新治疗方法的目标。RT-qPCR显示AMPA受体亚基GluA1和GluA2的mRNA明显上调，western blot分析显示GluA2的蛋白水平升高。因此，我们得出结论，观察到的减少的Ca2 +流入是基于含Ca2 +不可渗透AMPAR的GluA2的增加。GluRs在神经元中的功能减弱可能会导致在NPC1中观察到的进行性神经退行性变，并且可能代表了开发NPC1中新治疗方法的目标。RT-qPCR显示AMPA受体亚基GluA1和GluA2的mRNA明显上调，western blot分析显示GluA2的蛋白水平升高。因此，我们得出结论，观察到的减少的Ca2 +流入是基于含Ca2 +不可渗透AMPAR的GluA2的增加。GluRs在神经元中的功能减弱可能会导致在NPC1中观察到的进行性神经退行性变，并且可能代表了开发NPC1中新治疗方法的目标。