Shindo Atsushi, Nakamura Takehiro, Matsumoto Yoshihito, Kawai Nobuyuki, Okano Hideyuki, Nagao Seigo, Itano Toshifumi, Tamiya Takashi
Department of Neurological Surgery, Kagawa University Faculty of Medicine, Kagawa.
Neurol Med Chir (Tokyo). 2010;50(2):98-105; disucussion 105-6. doi: 10.2176/nmc.50.98.
Embryonic stem cells (ES cells) differentiate into multiple cell lineages including neural cells. The present study optimized the method to induce differentiation of gamma-aminobutyric acid-producing neurons (GABAergic neurons) from ES cell-derived neural stem/progenitor cells (NS/PCs), and transplanted these ES cell-derived GABAergic neurons producing neural progenitors into kindled epileptic mice, and analyzed the morphological and functional recovery from epilepsy. The response of kindling was evaluated by the modified Racine scale. Following stage 5 kindling, the mice were divided into two groups. Group 1 received NS/PCs derived from the ES cells ubiquitously expressing green fluorescent protein transplanted into the dorsal hippocampal area. Group 2 received microinjections of only the medium. After transplantation, the recovery of seizures was evaluated by the modified Racine scale again. All mice were perfused and fixed for immunohistochemical analysis after finishing the kindling experiment. In Group 1, one mouse was classified as stage 0, five as stage 3, and one as stage 4 recovering from stage 5 at 6 weeks after transplantation. In Group 2, all mice remained in stage 5. The transplanted cells were examined immunohistochemically using neuronal and GABAergic markers. In the transplanted mice, substantial hippocampal GABAergic re-innervation and seizure-suppressing effects were observed. NS/PCs derived from ES cells have high potential for use in transplantation therapy for clinically intractable epilepsies.
胚胎干细胞（ES细胞）可分化为包括神经细胞在内的多种细胞谱系。本研究优化了从ES细胞来源的神经干/祖细胞（NS/PCs）诱导分化产生γ-氨基丁酸能神经元（GABA能神经元）的方法，并将这些产生神经祖细胞的ES细胞来源的GABA能神经元移植到点燃癫痫小鼠体内，分析癫痫的形态学和功能恢复情况。通过改良的Racine量表评估点燃反应。在5期点燃后，将小鼠分为两组。第1组接受将普遍表达绿色荧光蛋白的ES细胞来源的NS/PCs移植到背侧海马区。第2组仅接受培养基的微量注射。移植后，再次通过改良的Racine量表评估癫痫发作的恢复情况。在完成点燃实验后，对所有小鼠进行灌注固定以进行免疫组织化学分析。在第1组中，1只小鼠在移植后6周被分类为0期，5只为3期，1只为从5期恢复的4期。在第2组中，所有小鼠仍处于5期。使用神经元和GABA能标记物对移植细胞进行免疫组织化学检查。在移植小鼠中，观察到大量海马GABA能神经再支配和癫痫抑制作用。ES细胞来源的NS/PCs在临床上难治性癫痫的移植治疗中具有很高的应用潜力。
