Türeyen Kudret, Vemuganti Raghu, Sailor Kurt A, Bowen Kellie K, Dempsey Robert J
Department of Neurological Surgery and Cardiovascular Research Center, University of Wisconsin, Madison, Wisconsin 53792, USA.
J Neurosurg. 2004 Nov;101(5):799-805. doi: 10.3171/jns.2004.101.5.0799.
Throughout the life of a mammal, new neurons are produced each day from resident progenitor cells located in the hippocampal dentate gyrus (DG). The availability of transgenic and knockout mice enables the evaluation of specific molecular mediators of this phenomenon. To facilitate such studies the authors characterized the proliferation, survival, and maturation of progenitor cells in the DG of adult mice following transient focal cerebral ischemia.
Anesthesia was induced in adult C57BL/6 mice by administering halothane. The middle cerebral artery (MCA) was then occluded for 120 minutes by applying an endovascular suture. The marker used to detect the presence of proliferating cells, 5-bromodeoxyuridine (BrdU; 50 mg/kg) was administered intraperitoneally twice daily on Days 2 through 6 after the MCA occlusion. Cohorts of mice were killed on Days 7 and 21, after which their brains were sectioned and BrdU-positive cells were detected using immunohistochemical analysis. The phenotype of the BrdU-positive cells was identified by fluorescent triple labeling by using antibodies specific for neuronal and astroglial markers together with anti-BrdU antibodies. The infarction was confirmed by applying cresyl violet staining. Compared with sham-operated control animals, there was a 4.6-fold (p < 0.05) increase in BrdU-positive cells in the ipsilateral DG at Day 7 postischemia. Twenty-one percent of the newly proliferated cells survived to Day 21 postischemia. At this time, the newly proliferated cells expressed the immature and mature neuron markers doublecortin and NeuN, respectively, but none expressed the astroglial marker glial fibrillary acidic protein.
Focal ischemia induces neurogenesis in the DG of the mouse brain; this may be critical for postischemic brain repair.
在哺乳动物的整个生命过程中，每天都有新的神经元由位于海马齿状回（DG）的常驻祖细胞产生。转基因和基因敲除小鼠的可得性使得评估这一现象的特定分子介质成为可能。为便于此类研究，作者对成年小鼠短暂局灶性脑缺血后DG中祖细胞的增殖、存活和成熟情况进行了表征。
通过给予氟烷对成年C57BL/6小鼠进行麻醉。然后通过应用血管内缝线将大脑中动脉（MCA）闭塞120分钟。用于检测增殖细胞存在的标记物5-溴脱氧尿苷（BrdU；50mg/kg）在MCA闭塞后的第2天至第6天每天腹腔注射两次。在第7天和第21天处死小鼠队列，之后将其大脑切片，并使用免疫组织化学分析检测BrdU阳性细胞。通过使用针对神经元和星形胶质细胞标记物的特异性抗体与抗BrdU抗体进行荧光三重标记来鉴定BrdU阳性细胞的表型。通过应用甲酚紫染色确认梗死。与假手术对照动物相比，缺血后第7天同侧DG中BrdU阳性细胞增加了4.6倍（p<0.05）。21%的新增殖细胞存活至缺血后第21天。此时，新增殖的细胞分别表达未成熟和成熟神经元标记物双皮质素和NeuN，但均未表达星形胶质细胞标记物胶质纤维酸性蛋白。
局灶性缺血诱导小鼠脑海马齿状回中的神经发生；这可能对缺血后大脑修复至关重要。
