Immunogold localization of tight junctional proteins in normal and osmotically-affected rat blood-brain barrier
Immunogold localization of tight junctional proteins in normal and osmotically-affected rat blood-brain barrier
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DOI：
10.1007/s10735-004-1318-3
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发表时间：
2004-06-01
影响因子：
3.2
通讯作者：
Vorbrodt, AW
中科院分区：
文献类型：
--
作者：
Dobrogowska, DH;Vorbrodt, AW
应用免疫金细胞化学技术，在电镜下观察了大鼠血脑屏障（BBB）微血管内皮细胞间紧密连接（TJ）分子组分的分布。对正常（未受影响）和受累（颈动脉内输注1.8 M L（+）阿拉伯糖后1、5和30 min）的大鼠脑样品进行处理，用于TJ特异性整合膜（闭合蛋白、JAM-1、claudin-5）和外周（ZO-1）蛋白分子的免疫细胞化学定位。在未受影响的内皮间连接的对照大鼠的免疫信号（由金颗粒表示）的occludin和ZO-1是最高的，而claudin-5和JAM-1的密度较低。输注后1分钟，未观察到连接相关分子分布的明显变化。然而，在5分钟时，变化是最明显的，它们包括节段性衰减的内皮衬里和扩张（开放）的一些交界处裂缝伴随着减少的密度的免疫信号TJ特异性跨膜和外周蛋白。这是由于这些分子与连接复合物的空间关系的无序化。30分钟后，许多内皮间连接似乎仍然开放，而其他连接部分或完全关闭。在开放的内皮间连接的TJ相关分子的表达弱于封闭的连接。我们的观察结果表明，TJ特异性蛋白质，特别是occludin的定位和表达，并在较低程度的claudin-5和JAM-1，连同周边ZO-1分子，渗透压休克的影响。据推测，这些蛋白质中的一些（例如，封闭蛋白、封闭蛋白-5和ZO-1）可以被认为是正常的以及也是BBB功能状态紊乱的敏感指标。
The distribution of molecular components of interendothelial tight junctions (TJs) was studied in rat blood-brain barrier (BBB) microvessels, using immunogold cytochemistry applied to electron microscopy. Samples of rat brains, both normal (unaffected) and osmotically-affected (1, 5, and 30 min after intracarotid infusion of 1.8 M L(+) arabinose), were processed for immunocytochemical localization of TJ-specific integral membrane (occludin, JAM-1, claudin-5) and peripheral (ZO-1) protein molecules. In unaffected interendothelial junctions of control rats the immunosignals (represented by gold particles) for occludin and ZO-1 were of highest, whereas for claudin-5 and JAM-1 were of lower density. At 1 min after infusion, no discernible changes in distribution of junction-associated molecules were noted. At 5 min, however, changes were most conspicuous, and they consisted of segmental attenuation of the endothelial lining and dilatation (opening) of some junctional clefts accompanied by the diminution of the density of immunosignals for TJ-specific transmembrane and peripheral proteins. It was paralleled by disorganization of the spatial relation of these molecules to the junctional complexes. After 30 min, many interendothelial junctions appeared to be still open, whereas other junctions were partially or totally closed. In the opened interendothelial junctions the expression of TJ-associated molecules was weaker than in closed junctions. Our observations indicate that the localization and expression of TJ-specific proteins, especially occludin, and in lower degree claudin-5 and JAM-1, together with the peripheral ZO-1 molecules, are affected by osmotic shock. Presumably, some of these proteins (e.g., occludin, claudin-5 and ZO-1) could be considered sensitive indicators of normal and also of disturbed functional state of the BBB.