Location of Neutrophils in Different Compartments of the Damaged Mouse Brain After Severe Ischemia/Reperfusion
Location of Neutrophils in Different Compartments of the Damaged Mouse Brain After Severe Ischemia/Reperfusion
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DOI：
10.1161/strokeaha.118.023837
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发表时间：
2019-06-01
期刊：
影响因子：
8.3
通讯作者：
Planas, Anna M.
中科院分区：
文献类型：
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作者：
Otxoa-de-Amezaga, Amaia;Gallizioli, Mattia;Planas, Anna M.
背景和目的缺血可吸引中性粒细胞进入受损的脑组织。然而，中性粒细胞的位置和进入受损脑组织的途径尚未完全了解。我们的目的是调查中性粒细胞的位置在小鼠模型的脑缺血/reperfusion.Methods成年雄性C57 BL/6小鼠（n=52）接受45分钟的管腔内大脑中动脉闭塞，然后再灌注14，24，48，或96小时。对假手术小鼠（n=9）进行整个外科手术。我们使用野生型小鼠和Catchup（IVM）小鼠，在中性粒细胞中表达红色荧光蛋白。此外，在缺血后，将从报道DsRed（discosoma红色荧光蛋白）小鼠获得的荧光嗜中性粒细胞静脉内转移到野生型小鼠。小鼠接受经心多聚甲醛灌注，大脑冷冻保护，冷冻，冷冻切片进行了研究，免疫荧光和共聚焦microscopic.Results缺血诱导的时间依赖性增加，脑中性粒细胞数量与假手术。我们在软脑膜、脑室、毛细血管腔、血管周围空间和梗死核心内的实质中检测到中性粒细胞。大多数缺血性小鼠在软脑膜和血管周围空间中显示中性粒细胞，而在缺血性小鼠中，实质中中性粒细胞的存在和数量是可变的。在前24小时内，仅少数小鼠显示实质中性粒细胞，但在48和96小时，实质中显示中性粒细胞的小鼠频率和中性粒细胞数量增加。我们还检测到基底膜破坏的迹象，偶尔中性粒细胞脱颗粒和中性粒细胞胞外traps.Conclusions形成提示缺血/再灌注后，中性粒细胞聚集在柔脑膜和血管周围的空间，并最终可以达到梗死的脑实质。
Background and Purpose Ischemia attracts neutrophils to the injured brain. However, neutrophil location and access to the damaged brain tissue is not yet entirely understood. We aimed to investigate neutrophil location in a mouse model of cerebral ischemia/reperfusion.Methods Adult male C57BL/6 mice (n=52) received 45-minute intraluminal middle cerebral artery occlusion followed by 14, 24, 48, or 96 hours of reperfusion. Sham-operated mice (n=9) were subjected to the entire surgical procedure. We used wild-type mice and Catchup(IVM) mice expressing a red fluorescent protein in neutrophils. In addition, fluorescent neutrophils obtained from reporter DsRed (discosoma red fluorescent protein) mice were transferred intravenously to wild-type mice after ischemia. Mice received transcardial paraformaldehyde perfusion, the brain was cryoprotected, frozen, and cryostat sections were studied by immunofluorescence and confocal microscopy.Results Ischemia induced a time-dependent increase in brain neutrophil numbers versus sham operation. We detected neutrophils in the leptomeninges, ventricles, capillary lumen, perivascular spaces, and parenchyma within the infarcted core. Most ischemic mice showed neutrophils in the leptomeninges and perivascular spaces, whereas the presence and number of neutrophils in the parenchyma was variable among ischemic mice. During the first 24 hours, only a few mice showed parenchymal neutrophils, but the frequency of mice showing neutrophils in the parenchyma and neutrophil numbers increased at 48 and 96 hours. We also detected signs of basement membrane disruption and hints of occasional neutrophil degranulation and formation of neutrophil extracellular traps.Conclusions After ischemia/reperfusion, neutrophils accumulate in the leptomeninges and perivascular spaces, and eventually can reach the infarcted brain parenchyma.