Polarized hemichannel opening of pannexin 1/connexin 43 contributes to dysregulation of transport function in blood-brain barrier endothelial cells
Polarized hemichannel opening of pannexin 1/connexin 43 contributes to dysregulation of transport function in blood-brain barrier endothelial cells
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DOI：
10.1016/j.neuint.2019.104600
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发表时间：
2020-01-01
影响因子：
4.2
通讯作者：
Hosoya, Ken-ichi
中科院分区：
文献类型：
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作者：
Tachikawa, Masanori;Murakami, Koji;Hosoya, Ken-ichi
血脑屏障（BBB）运输失调会加剧急性缺血性中风的脑损伤。在这里，我们的目的是通过研究大鼠（TR-BBB13细胞）和人类（hCMEC/D3细胞）血脑屏障内皮细胞中pannexin（Px）和connexin（Cx）半通道在急性缺血性中风模拟缺氧/葡萄糖剥夺（OGD）和无细胞外Ca2+（[Ca2+](e)）条件下的定位和功能来研究这种BBB转运失调的机制条件。 TR-BBB13 细胞显示半通道可渗透的磺基罗丹明 101 的摄取增加，并且这种增加被半通道抑制剂甘酸索酮显着抑制。在TR-BBB13细胞和新鲜分离的脑微血管内皮细胞中检测到Px1和Cx43的转录本。在基底 Transwell 室中无 [Ca2+](e) 的条件下，hCMEC/D3 细胞中半通道可渗透的碘化丙啶的基底室到细胞的摄取选择性增强。免疫组织化学分析揭示了 Cx43 在 hCMEC/D3 细胞侧膜上的主要定位。在缺乏[Ca2+](e)的情况下，hCMEC/D3细胞对[H-3]牛磺酸的摄取显着减少。在无 [Ca2+](e) 的条件下，用模拟肽功能性敲低 Px1 和 Cx43 显着抑制 hCMEC/D3 细胞 ATP 释放的增加。这些结果表明，在模拟急性缺血性中风的条件下，脑毛细血管内皮细胞中的极化 Px1/Cx43 半通道开放导致 BBB 转运功能失调，导致细胞内牛磺酸和 ATP 的释放。
Dysregulation of blood-brain barrier (BBB) transport exacerbates brain damage in acute ischemic stroke. Here, we aimed to investigate the mechanism of this BBB transport dysregulation by studying the localization and function of pannexin (Px) and connexin (Cx) hemichannels in blood-brain barrier endothelial cells of rat (TR-BBB13 cells) and human (hCMEC/D3 cells) under acute ischemic stroke-mimicking oxygen/glucose deprivation (OGD) and extracellular Ca2+ ([Ca2+](e))-free conditions. TR-BBB13 cells showed increased uptake of hemichannel-permeable sulforhodamine 101, and this increase was markedly inhibited by carbenoxolone, a hemichannel inhibitor. Transcripts of Px1 and Cx43 were detected in TR-BBB13 cells and freshly isolated brain microvascular endothelial cells. The basal compartment-to-cell uptake of hemichannel-permeable propidium iodide was selectively enhanced in hCMEC/D3 cells under [Ca2+](e)-free conditions in the basal Transwell chamber. Immunohistochemical analysis revealed the predominant localization of Cx43 on the lateral membranes of hCMEC/D3 cells. [H-3]Taurine uptake by hCMEC/D3 cells was significantly reduced in the absence of [Ca2+](e). Functional knock-down of Px1 and Cx43 with mimetic peptides significantly inhibited the increase of ATP release from hCMEC/D3 cells under [Ca2+](e)-free conditions. These results suggest that polarized Px1/Cx43 hemichannel opening in brain capillary endothelial cells under acute ischemic stroke-mimicking conditions contributes to dysregulation of BBB transport function, resulting in release of intracellular taurine and ATP.