Abe Masaaki, Sano Yasuteru, Maeda Toshihiko, Shimizu Fumitaka, Kashiwamura Yoko, Haruki Hiroyo, Saito Kazuyuki, Tasaki Ayako, Kawai Motoharu, Terasaki Tetsuya, Kanda Takashi
Department of Neurology and Clinical Neuroscience, Yamaguchi University Graduate School of Medicine, 1-1-1 Minamikogushi, Ube, Yamaguchi 755-8505, Japan.
Cell Struct Funct. 2012;37(2):89-100. doi: 10.1247/csf.11042. Epub 2012 May 19.
The blood-nerve barrier (BNB) is a highly specialized unit that maintains the microenvironments of the peripheral nervous system. Since the breakdown of the BNB has been considered a key step in autoimmune neuropathies such as Guillain-Barré syndrome and chronic inflammatory demyelinating polyraduculoneuropathy, it is important to understand the cellular properties of the peripheral nerve microvascular endothelial cells (PnMECs) which constitute the BNB. For this purpose, we established an immortalized cell line derived from human PnMECs. The human PnMECs were transduced with retroviral vectors encoding the temperature-sensitive SV40 large T antigen and human telomerase. This cell line, termed FH-BNB, showed a spindle fiber-shaped morphology, expression of von Willebrand factor and uptake of acetylated low density lipoprotein. These cells expressed tight junction proteins including occludin, claudin-5, ZO-1 and ZO-2 at the cell-cell boundaries. P-glycoprotein and GLUT-1 were also detected by a Western blot analysis and the cells exhibited the functional expression of p-glycoprotein. In addition, transendothelial electrical resistance experiments and paracellular permeabilities of sodium fluorescein and fluorescein isothiocyanate-labeled dextran of molecular weight 4 kDa across these cells demonstrated that FH-BNBs had functional tight junctions. These results indicated that FH-BNBs had highly specialized barrier properties and they might therefore be a useful tool to analyze the pathophysiology of various neuropathies.
血神经屏障（BNB）是一个高度专业化的单元，可维持周围神经系统的微环境。由于血神经屏障的破坏被认为是格林-巴利综合征和慢性炎症性脱髓鞘性多发性神经根神经病等自身免疫性神经病的关键步骤，因此了解构成血神经屏障的周围神经微血管内皮细胞（PnMECs）的细胞特性非常重要。为此，我们建立了一种源自人PnMECs的永生化细胞系。用人PnMECs转导编码温度敏感型SV40大T抗原和人端粒酶的逆转录病毒载体。这个细胞系称为FH-BNB，呈现纺锤形纤维形态，表达血管性血友病因子并摄取乙酰化低密度脂蛋白。这些细胞在细胞-细胞边界处表达紧密连接蛋白，包括闭合蛋白、Claudin-5、ZO-1和ZO-2。通过蛋白质印迹分析还检测到P-糖蛋白和GLUT-1，并且这些细胞表现出P-糖蛋白的功能性表达。此外，跨内皮电阻实验以及荧光素钠和分子量为4 kDa的异硫氰酸荧光素标记的葡聚糖在这些细胞上的细胞旁通透性表明，FH-BNB具有功能性紧密连接。这些结果表明，FH-BNB具有高度专业化的屏障特性，因此它们可能是分析各种神经病病理生理学的有用工具。
