Characterisation of a mouse cerebral microvascular endothelial cell line (bEnd.3) after oxygen glucose deprivation and reoxygenation
Characterisation of a mouse cerebral microvascular endothelial cell line (bEnd.3) after oxygen glucose deprivation and reoxygenation
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DOI：
10.1111/1440-1681.12587
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发表时间：
2016-08-01
影响因子：
2.9
通讯作者：
Miller, Alyson A.
中科院分区：
文献类型：
--
作者：
Ku, Jacqueline M.;Taher, Mohammadali;Miller, Alyson A.
研究利用暴露于氧葡萄糖剥夺（OGD）的永生化小鼠脑内皮细胞（bEnd.3）来研究缺血后血脑屏障（BBB）的破坏。然而，缺乏描述最佳模拟体内BBB破坏所需的OGD（和再氧合[RO]）持续时间的文献。在这项研究中，我们评估了bEnd.3细胞在暴露于一系列OGD期后以及OGD+RO后的BBB破坏。将bEnd.3单层暴露于4、6、16或24小时的OGD导致渗透性显著增加。16或24小时后的高通透性与紧密连接蛋白（occludin和claudin-5）的表达减少有关。此外，细胞活力降低，促凋亡蛋白，裂解的半胱天冬酶-3的表达增加。相对于1小时OGD，将bEnd.3单层暴露于1小时OGD+23小时RO加剧了高渗透性，这与闭合蛋白和ZO-1的表达水平降低有关，但细胞活力或半胱天冬酶-3无变化。4小时OGD+23小时RO加重了细胞通透性增高，降低了细胞紧密连接蛋白的表达水平，降低了细胞活力，并增加了caspase-3的表达。因此，bEnd.3细胞在暴露于长时间的OGD后表现出高渗透性、紧密连接蛋白的损失，并经历细胞死亡。此外，在OGD+RO后，它们表现出加重的高通透性、紧密连接蛋白的损失和半胱天冬酶-3的表达增加。这些发现将促进该细胞系在BBB破坏研究和治疗剂测试中的使用。
Studies have utilised immortalised mouse cerebral endothelial cells (bEnd.3) exposed to oxygen glucose deprivation (OGD) to study blood-brain barrier (BBB) disruption after ischaemia. However, there is a paucity of literature describing the duration of OGD (and reoxygenation [RO]) required to best simulate BBB disruption in vivo. In this study we assessed BBB disruption in bEnd.3 cells after exposure to a range of OGD periods, and also after OGD+RO. Exposure of bEnd.3 monolayers to 4, 6, 16, or 24hours of OGD resulted in a significant increase in permeability. The hyperpermeability after 16 or 24hours was associated with decreased expression of tight junction proteins (occludin and claudin-5). Furthermore, there was a decrease in cell viability and increased expression of the pro-apoptotic protein, cleaved caspase-3. Exposure of bEnd.3 monolayers to 1hour OGD+ 23hours RO exacerbated hyperpermeability relative to 1 hour OGD, which was associated with decreased expression levels of occludin and ZO-1, but no change in cell viability or caspase-3. 4hours OGD+23hours RO exacerbated hyperpermeability, decreased expression levels of tight junction proteins, decreased cell viability, and increased caspase-3 expression. Thus, bEnd.3 cells exhibit hyperpermeability, a loss of tight junction proteins, and undergo cell death, after exposure to prolonged periods of OGD. Moreover, they exhibit exacerbated hyperpermeability, a loss of tight junction proteins, and increased expression of caspase-3 after OGD+RO. These findings will facilitate the use of this cell line in studies of BBB disruption and for the testing of therapeutics.