Kameda M, Shingo T, Takahashi K, Muraoka K, Kurozumi K, Yasuhara T, Maruo T, Tsuboi T, Uozumi T, Matsui T, Miyoshi Y, Hamada H, Date I
Department of Neurological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho Okayama, Okayama, 700-8558, Japan.
Eur J Neurosci. 2007 Sep;26(6):1462-78. doi: 10.1111/j.1460-9568.2007.05776.x.
Adult neural stem and progenitor cells (NSPCs) are important autologous transplantation tools in regenerative medicine, as they can secrete factors that protect the ischemic brain. We investigated whether adult NSPCs genetically modified to secrete more glial cell line-derived neurotrophic factor (GDNF) could protect against transient ischemia in rats. NSPCs were harvested from the subventricular zone of adult Wistar rats and cultured for 3 weeks in the presence of epidermal growth factor. The NSPCs were treated with fibre-mutant Arg-Gly-Asp adenovirus containing the GDNF gene (NSPC-GDNF) or enhanced green fluorescent protein (EGFP) gene (NSPC-EGFP; control group). In one experiment, cultured cells were transplanted into the right ischemic boundary zone of Wistar rat brains. One week later, animals underwent 90 min of intraluminal right middle cerebral artery occlusion followed by magnetic resonance imaging and behavioural tests. The NSPC-GDNF group had higher behavioural scores and lesser infarct volume than did controls at 1, 7 and 28 days postocclusion. In the second experiment, we transplanted NSPCs 3 h after ischemic insult. Compared to controls, rats receiving NSPC-GDNF had decreased infarct volume and better behavioural assessments at 7 days post-transplant. Animals were killed on day 7 and brains were collected for GDNF ELISA and morphological assessment. Compared to controls, more GDNF was secreted, more NSPC-GDNF cells migrated toward the ischemic core and more NSPC-GDNF cells expressed immature neuronal marker. Moreover, the NSPC-GDNF group showed more effective inhibition of microglial invasion and apoptosis. These findings suggest that NSPC-GDNF may be useful in treatment of cerebral ischemia.
成体神经干细胞和祖细胞（NSPCs）是再生医学中重要的自体移植工具，因为它们能分泌保护缺血性脑的因子。我们研究了经基因改造以分泌更多胶质细胞源性神经营养因子（GDNF）的成体NSPCs是否能保护大鼠免受短暂性缺血损伤。从成年Wistar大鼠的脑室下区采集NSPCs，并在表皮生长因子存在的情况下培养3周。用含有GDNF基因的纤维突变型精氨酸-甘氨酸-天冬氨酸腺病毒（NSPC-GDNF）或增强型绿色荧光蛋白（EGFP）基因（NSPC-EGFP；对照组）处理NSPCs。在一项实验中，将培养的细胞移植到Wistar大鼠脑的右侧缺血边界区。一周后，对动物进行90分钟的右侧大脑中动脉腔内闭塞，随后进行磁共振成像和行为测试。在闭塞后1天、7天和28天，NSPC-GDNF组的行为评分高于对照组，梗死体积小于对照组。在第二项实验中，我们在缺血损伤后3小时移植NSPCs。与对照组相比，接受NSPC-GDNF的大鼠在移植后7天梗死体积减小，行为评估更好。在第7天处死动物并收集大脑进行GDNF ELISA和形态学评估。与对照组相比，分泌的GDNF更多，更多的NSPC-GDNF细胞向缺血核心迁移，更多的NSPC-GDNF细胞表达未成熟神经元标记物。此外，NSPC-GDNF组对小胶质细胞浸润和凋亡的抑制更有效。这些发现表明NSPC-GDNF可能对治疗脑缺血有用。
