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Analysis of L-leucine amino acid transporter species activity and gene expression by human blood brain barrier hCMEC/D3 model reveal potential LAT1, LAT4, B0AT2 and y+LAT1 functional cooperation
Journal of Cerebral Blood Flow & Metabolism ( IF 6.3 ) Pub Date : 2021-08-24 , DOI: 10.1177/0271678x211039593
Mehdi Taslimifar 1, 2 , Martin Faltys 2, 3 , Vartan Kurtcuoglu 1, 4 , François Verrey 2, 4 , Victoria Makrides 1, 2, 5
Affiliation  

In the CNS, amino acid (AA) neurotransmitters and neurotransmitter precursors are subject to tight homeostatic control mediated by blood-brain barrier (BBB) solute carrier amino acid transporters (AATs). Since the BBB is composed of multiple closely apposed cell types and opportunities for human in vivo studies are limited, we used in vitro and computational approaches to investigate human BBB AAT activity and regulation. Quantitative real-time PCR (qPCR) of the human BBB endothelial cell model hCMEC/D3 (D3) was used to determine expression of selected AAT, tight junction (TJ), and signal transduction (ST) genes under various culture conditions. L-leucine uptake data were interrogated with a computational model developed by our group for calculating AAT activity in complex cell cultures. This approach is potentially applicable to in vitro cell culture drug studies where multiple “receptors” may mediate observed responses. Of 7 Leu AAT genes expressed by D3 only the activity of SLC7A5-SLC3A2/LAT1-4F2HC (LAT1), SLC43A2/LAT4 (LAT4) and sodium-dependent AATs, SLC6A15/B0AT2 (B0AT2), and SLC7A7/y+LAT1 (y+LAT1) were calculated to be required for Leu uptake. Therefore, D3 Leu transport may be mediated by a potentially physiologically relevant functional cooperation between the known BBB AAT, LAT1 and obligatory exchange (y+LAT1), facilitative diffusion (LAT4), and sodium symporter (B0AT2) transporters.



中文翻译:

人血脑屏障 hCMEC/D3 模型分析 L-亮氨酸氨基酸转运蛋白活性和基因表达揭示潜在的 LAT1、LAT4、B0AT2 和 y+LAT1 功能合作

在 CNS 中,氨基酸 (AA) 神经递质和神经递质前体受到血脑屏障 (BBB) 溶质载体氨基酸转运蛋白 (AAT) 介导的严格稳态控制。由于 BBB 由多种紧密并列的细胞类型组成,人体体内研究的机会有限,我们在体外使用和计算方法来研究人类 BBB AAT 活动和调节。人 BBB 内皮细胞模型 hCMEC/D3 (D3) 的定量实时 PCR (qPCR) 用于确定各种培养条件下选定的 AAT、紧密连接 (TJ) 和信号转导 (ST) 基因的表达。L-亮氨酸摄取数据用我们小组开发的计算模型进行询问,用于计算复杂细胞培养物中的 AAT 活性。这种方法可能适用于体外细胞培养药物研究,其中多个“受体”可能介导观察到的反应。在 D3 表达的 7 个 Leu AAT 基因中,只有 SLC7A5-SLC3A2/LAT1-4F2HC (LAT1)、SLC43A2/LAT4 (LAT4) 和钠依赖性 AAT、SLC6A15/B 0 AT2 (B 0 AT2) 和 SLC7A7/y 的活性+ LAT1 (y + LAT1) 被计算为 Leu 摄取所必需的。因此,D3 Leu 转运可能由已知的 BBB AAT、LAT1 和强制性交换 (y + LAT1)、促进扩散 (LAT4) 和钠同向转运蛋白 (B 0 AT2) 转运蛋白之间潜在的生理相关功能合作介导。

更新日期:2021-08-24
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