The impact of agrin on the formation of orthogonal arrays of particles in cultured astrocytes from wild-type and agrin-null mice
The impact of agrin on the formation of orthogonal arrays of particles in cultured astrocytes from wild-type and agrin-null mice
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DOI：
10.1016/j.brainres.2010.09.092
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发表时间：
2011-01-07
期刊：
影响因子：
2.9
通讯作者：
Noell, Susan
中科院分区：
文献类型：
--
作者：
Fallier-Becker, Petra;Sperveslage, Jan;Noell, Susan
星形细胞终足膜上布满了水通道蛋白-4(AQP4)，其中含有颗粒的正交排列(OAP)，这可以通过冷冻断裂的方法来唯一地观察到。它们主要表达在星形胶质细胞膜与血管浅层和血管周围基底板接触的地方。这种极性似乎对血脑屏障(BBB)的完整性至关重要。基膜含有许多细胞外基质(ECM)成分，如胶原、层粘连蛋白和聚集素等硫酸乙酰肝素蛋白多糖，被认为影响星形胶质细胞的这种与OAP相关的极性。最近，我们发现集聚蛋白，特别是神经元亚型A4B8，能够影响星形胶质细胞在集聚蛋白条件培养液中OAP的形成。在这篇文章中，我们想要研究与其他ECM成分涂层相比，外源性集聚蛋白涂层是否会诱导集聚蛋白缺失小鼠星形胶质细胞中OAP的形成。为此，我们将来自集聚蛋白缺失和野生型小鼠的星形胶质细胞培养在集聚蛋白或细胞外基质涂层的表面。AQP4的免疫荧光细胞化学染色显示，AQP4在聚集素或细胞外基质包被的培养物中的表达水平高于未包被的培养物，而蛋白质印迹分析和聚合酶链式反应显示没有差异。α-营养不良聚糖被认为是一种潜在的集聚蛋白受体，在野生型和集聚蛋白缺失的星形胶质细胞中均有免疫染色。在冷冻断裂的复制品中，我们观察到星形胶质细胞在琼脂和细胞外基质涂层上生长时OAP密度增加。这些结果与其他实验一致，这些实验测量了低渗应激后的体积变化，支持了外源agrin对AQP4插入细胞膜、OAP的形成和水分运输的积极影响。(C)2010爱思唯尔B.V.保留所有权利。
Astrocytic endfeet membranes are studded with aquaporin-4 (AQP4) containing orthogonal arrays of particles (OAP) which can be visualized exclusively by the freeze-fracturing method. They are predominantly expressed where the astroglial membrane is in contact with the superficial and perivascular basal lamina. This polarity seems to be essential for the integrity of the blood-brain barrier (BBB). The basal lamina containing many extracellular matrix (ECM) components such as collagen, laminin and heparansulfate proteoglycans like agrin is thought to influence this OAP-related polarity of astrocytes. Recently, we have shown that agrin, in particular the neuronal isoform A4B8, is capable of influencing the formation of OAPs in astrocytes when cultured in the presence of agrin-conditioned media. In this paper we wanted to investigate whether coating with exogenous agrin compared to coating with other ECM components would induce OAP formation in astrocytes of the agrin-null mouse. For this purpose, we cultured astrocytes from agrin-null and wild-type mice on agrin- or ECM-coated surfaces. Immunofluorescent cytochemical staining of AQP4 indicated a higher AQP4 expression level in cultures with agrin- or ECM-coated than in cultures with uncoated surfaces, whereas western blot analyses and PCR showed no differences. alpha-Dystroglycan is thought to be a potential receptor of agrin and was immunostained in wild-type as well as in agrin-null astrocytes. In freeze-fracture replicas, we observed an increase in OAP density in astrocytes when growing on agrin- and ECM-coatings. These results concurred with other experiments in which changes in volume were measured following hypotonic stress, which supported the positive influence of exogenous agrin on AQP4 insertion into the membrane, on OAP formation and on water transport. (C) 2010 Elsevier B.V. All rights reserved.