In vivo three-dimensional evaluation of leukocyte behavior in retinal microcirculation of mice
In vivo three-dimensional evaluation of leukocyte behavior in retinal microcirculation of mice
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DOI：
10.1167/iovs.04-0192
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发表时间：
2004-11-01
影响因子：
4.4
通讯作者：
Yoshimura, N
中科院分区：
文献类型：
--
作者：
Miyahara, S;Kiryu, J;Yoshimura, N
目的.评价新的生理和三维方法监测白细胞在小鼠视网膜中的行为。通过在小鼠足垫注射脂多糖（LPS），建立内毒素诱导的葡萄膜炎（EIU）模型。用吖啶橙子（AO）标记白细胞。用AO数字荧光成像评价视网膜微循环中白细胞滚动。在平铺的视网膜中计数迁移的白细胞的数量。白细胞迁移的行为进行了三维观察时，迁移的高峰。用AO标记白细胞后，用罗丹明标记的刀豆球蛋白A凝集素灌注小鼠血管内皮，染色。然后用共聚焦显微镜三维监测白细胞向视网膜的迁移，并测量迁移速度。白细胞滚动和迁移均在注射LPS后48小时达到峰值。观察到白细胞从较深的毛细血管层渗出并向视网膜外层迁移。视网膜组织中渗出的白细胞的移动速度为2.0+/-0.1 mum/h。新的方法已被证明是三维和定量评价白细胞在小鼠视网膜的行为。
Purpose. To evaluate new physiologic and three-dimensional methods for monitoring leukocyte behavior in mouse retina.Methods. Endotoxin-induced uveitis (EIU) was produced in mice by footpad injection of lipopolysaccharide (LPS). Leukocytes were labeled with acridine orange (AO). Leukocyte rolling in the retinal microcirculation was evaluated in vivo with AO digital fluorography. The number of migrated leukocytes was counted in flatmounted retina. The behavior of leukocyte migration was observed three-dimensionally at the time of peak migration. After leukocytes were labeled with AO, the mice were perfused with rhodamine-labeled concanavalin A lectin to stain the vascular endothelium. Leukocyte migration into the retina was then monitored three-dimensionally with confocal microscopy, and the velocity of the migration was measured.Results. Both leukocyte rolling and migration peaked at 48 hours after LPS injection. Leukocytes were seen to extravasate from the deeper capillary layers and to migrate toward the outer layer of the retina. The traveling velocity of extravasated leukocytes in retinal tissue was 2.0+/-0.1 mum/h.Conclusions. New methods have been demonstrated for the three-dimensional and quantitative evaluation of leukocyte behavior in mouse retina.