Murata Hidetoshi, Tajima Nobuyoshi, Nagashima Yoji, Yao Masahiro, Baba Masaya, Goto Masayuki, Kawamoto Susumu, Yamamoto Isao, Okuda Kenji, Kanno Hiroshi
Departments of Neurosurgery, Yokohama City University School of Medicine, Yokohama 236-0004, Japan.
Cancer Res. 2002 Dec 1;62(23):7004-11.
Von Hippel-Lindau (VHL) tumor suppressor protein is expressed in neurons of the central nervous system and plays an important role during the neuronal differentiation of central nervous system progenitor cells. To elucidate the neuronal differentiating potential of VHL protein in neuroblastoma cells, we overexpressed or inhibited VHL protein in human neuroblastoma cells (SY-SH5Y), and examined the morphological change, expressions of neuronal markers, and electrophysiological functions. Here we show that with VHL gene transduction SY-SH5Y cells stably expressing the VHL protein had neurite-like processes with varicosities, showed the distinct expression of the neuronal markers neuropeptide Y and neurofilament 200, acquired regulated neurosecretion competence in response to depolarizing and cholinergic stimuli, and had large voltage-gated fast sodium currents and delayed rectifier potassium (Kv) currents compatible with those of functional neurons. In addition, they displayed inactivated ether-á-go-go potassium channels related to the promotion of the cell cycle and to the termination of differentiation. Also, by treatment with retinoic acid, they rapidly underwent cell death related to apoptosis. These findings suggest that the induction of neuronal function by VHL protein is associated with down-regulation of the cell cycle. In contrast, the inhibition of endogenous expression of VHL protein with antisense-orientated VHL gene transduction reduced such neuronal properties inherent to these cells, including the capacity for activation of ether-á-go-go channels. In conclusion, VHL protein has a neuronal differentiating potential to transform neuroblastoma cells into functional neuron-like cells. Our finding of the neuronal differentiation of neuroblastoma cells under the control of the VHL gene may contribute to the development of clinical techniques for neuronal regeneration in the case of intractable neuronal diseases and for differentiation therapy against neuroblastomas.
冯·希佩尔-林道（VHL）肿瘤抑制蛋白在中枢神经系统的神经元中表达，并在中枢神经系统祖细胞的神经元分化过程中发挥重要作用。为了阐明VHL蛋白在神经母细胞瘤细胞中的神经元分化潜能，我们在人神经母细胞瘤细胞（SY-SH5Y）中过表达或抑制VHL蛋白，并检测其形态变化、神经元标志物的表达及电生理功能。我们发现，通过VHL基因转导，稳定表达VHL蛋白的SY-SH5Y细胞具有带有膨体的神经突样突起，显示出神经肽Y和神经丝200这两种神经元标志物的明显表达，在去极化和胆碱能刺激下获得了调节性神经分泌能力，并且具有与功能性神经元相符的大电压门控快速钠电流和延迟整流钾（Kv）电流。此外，它们表现出与细胞周期促进和分化终止相关的失活的醚-á-去极化钾通道。而且，经视黄酸处理后，它们迅速经历与凋亡相关的细胞死亡。这些发现表明，VHL蛋白诱导的神经元功能与细胞周期的下调有关。相反，用反义方向的VHL基因转导抑制VHL蛋白的内源性表达降低了这些细胞固有的神经元特性，包括激活醚-á-去极化通道的能力。总之，VHL蛋白具有将神经母细胞瘤细胞转化为功能性神经元样细胞的神经元分化潜能。我们在VHL基因控制下发现神经母细胞瘤细胞的神经元分化，可能有助于开发针对难治性神经元疾病的神经元再生临床技术以及针对神经母细胞瘤的分化疗法。
