Effects of Hypothermia on Subventricular Neural Stem Cells Following Focal Cerebral Ischemia in Adult Rats

低温对成年大鼠局灶性脑缺血后室下神经干细胞的影响

• 批准号: 15591641
• 负责人: YANO Toshiyuki
• 金额: $ 2.24万
• 依托单位: Kumamoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15591641/
• 关键词: hypothermia; neural stem cells; subventricular zone; focal cerebral ischemia; immunohistochemistry; DiI; rat; ependymal cell; cerebral ischemia; stem cell

Background : Recently, neural stem cells have been identified in the subventricular zone(SVZ) and the hippocampal dentate gyrus of adult rodents. Hypothermia is well known to provide neuroprotection following various brain insults, In this study, we examined if hypothermia activated neural stem cell of the SVZ exposed to focal cerebral ischemia.Materials and Methods : Adult male Wistar rats were anesthetized, intubated and ventilated with isoflurane and N_2O/O_2. DiI(0.2%,10 μL) was injected into the left lateral ventricle. After preparation the animals were assigned to the following groups : group 1(hypothermia 32.5±0.5℃), group 2(ischemia/hypothermia 32.5±0.5℃), and group 3(ischemia/normothermia 37.5±0.5℃. Ischemia(120 min) was induced by right middle cerebral occlusion with a 4-0 nylon suture and confirmed by the decreased regional cerebral blood flow using laser Doppler flowmetry. Hypothermia for 6 hours was applied immediately after the initiation of ischemia. Seven days after hyp … More othermia or ischemia, bromodeoxyuridine(BrdU) was administered intraperitoneally 10 and 2 hours before sacrifice and the brains were removed, fixed with 4% paraformadlehyde in PBS, cryoprotected with graded sucrose solutions, frozen and cut for DiI fluolescence detection and fluolescent immunohistochemistry. Separately, the brains were stained with triphenyltetrazolium chloride for assessment of infarct area.Results : In the group 3,DiI-positive cells migrated from the SVZ toward the striatum on days 7 and 14. Infarct area on day 7 was much larger in the group 3 than in the group 2. Hypothermia did not facilitate the migration of DiI positive cells from the SVZ. BrdU and doublecortin were expressed in the SVZ and the rostral migratory stream in all groups and into the infarct region in the groups 2 and 3. Nestih that is expressed in neural precursors was developed in the whole ischemic hemisphere but not in the contralateral hemisphere. Migrating DiI-positive cells occasionally expressed NeuN or GFAP. Howerver, hypothermia did not influence the expression of BrdU, doublecortin, nestin, NeuN, and GFAP.Conclusion : Hypothermia did not activate the subventricular neural stem cells to prolirerate, migrate, and difierentiate into neurons or glia in the injured striatum following focal cerebral ischemia. Less

背景：最近在成年啮齿类动物的脑室下区(SVZ)和海马齿状回中发现了神经干细胞。在本研究中，我们观察了低温对局灶性脑缺血大鼠的神经干细胞的激活作用。材料和方法：成年雄性Wistar大鼠麻醉，气管插管，吸入异氟醚和N_2O/O_2。左侧脑室注射0.2%DiI(0.2%，10μL)。实验结束后，动物分为3组：低温组(32.5±0.5℃)、缺血/低温组(32.5±0.5℃)、缺血/常温组(37.5±0.5)℃。用4-0尼龙线结扎右侧大脑中叶造成脑缺血(120min)，激光多普勒血流仪证实局部脑血流量减少。缺血开始后立即进行6h的低温治疗。Hyp…后七天处死前10小时和2小时分别经腹腔注射溴脱氧尿苷(BrdU)，取脑，用4%对甲醛PBS固定，分级蔗糖液冷冻保护，冰冻切片进行DiI荧光检测和荧光免疫组织化学染色。结果：第3组DII阳性细胞在第7天和第14天从SVZ向纹状体迁移，第7天梗塞面积明显大于第2组。低温不利于DII阳性细胞从SVZ向纹状体迁移。BrdU和Doublecortin在各组的SVZ和吻侧迁移流处均有表达，并进入梗死区。神经前体表达的Nestih在整个缺血侧大脑半球均有表达，而对侧半球则不表达。迁移性DiI阳性细胞偶见NeuN或GFAP表达。但低温不影响BrdU、Doublecortin、Nestin、NeuN和GFAP的表达。结论：低温不能激活室下神经干细胞在局灶性脑缺血后向神经元或神经胶质细胞的增殖、迁移和分化。较少