Zhang Xiaobin, Alakhova Daria Y, Batrakova Elena V, Li Shu, Yang Zhihui, Li Yili, Kabanov Alexander V
Department of Pharmaceutical Sciences and Center for Drug Delivery and Nanomedicine, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE 68198, USA.
J Neuroimmune Pharmacol. 2009 Mar;4(1):35-46. doi: 10.1007/s11481-008-9119-1. Epub 2008 Aug 2.
A synthetic amphiphilic block copolymer Pluronic P85 (P85) was shown to be among the most potent inhibitors of Pgp efflux system in the blood-brain barrier (BBB) and capable of enhancing delivery of Pgp substrates to the brain. The purpose of this work is to evaluate the effects of P85 on amino acid transport in BBB. Primary bovine brain microvessel endothelial cells (BBMEC) grown on membrane inserts were used as an in vitro BBB model. Expression of amino acid transporters, like large neutral amino acid transporter 1, cationic amino acid transporter 1, and small neutral amino acid transporter 1, were confirmed by reverse transcriptase polymerase chain reaction. Effects of P85 on amino acid transporters were examined using their substrates: (3)H-phenylalanine, (3)H-lysine, and (3)H-methylaminoisobutyric acid, respectively. BBMEC permeability studies were carried out in apical (AP) to basolateral (BL) and BL to AP directions. P85 added at the AP side had little, if any, effect on AP to BL ("blood to brain") transport for all examined amino acids in BBMEC monolayers. However, 0.1% P85 added at the BL side significantly increased the BL to AP transport of these substrates. Furthermore, the effective concentrations of P85 were also shown to induce plasma membrane depolarization and increase intracellular sodium concentration in BBMEC, which can contribute to the effects of the copolymer on the energy-dependent transport systems. All together, despite profound effects on transport system(s) at the brain side of cell monolayers, P85 had no effect on AP to BL transport of amino acids in brain microvessel endothelial cell model.
合成两亲性嵌段共聚物普朗尼克P85（P85）被证明是血脑屏障（BBB）中P-糖蛋白外排系统最有效的抑制剂之一，并且能够增强P-糖蛋白底物向脑内的递送。本研究的目的是评估P85对BBB中氨基酸转运的影响。将生长在膜插入物上的原代牛脑微血管内皮细胞（BBMEC）用作体外BBB模型。通过逆转录聚合酶链反应确认氨基酸转运体的表达，如大中性氨基酸转运体1、阳离子氨基酸转运体1和小中性氨基酸转运体1。分别使用它们的底物（3）H-苯丙氨酸、（3）H-赖氨酸和（3）H-甲基氨基异丁酸来检测P85对氨基酸转运体的影响。在顶侧（AP）至基底外侧（BL）以及BL至AP方向上进行BBMEC通透性研究。在AP侧添加P85对BBMEC单层中所有检测氨基酸的AP至BL（“血至脑”）转运几乎没有影响（如果有影响的话）。然而，在BL侧添加0.1%的P85显著增加了这些底物的BL至AP转运。此外，还表明P85的有效浓度可诱导BBMEC中的质膜去极化并增加细胞内钠浓度，这可能有助于该共聚物对能量依赖性转运系统的影响。总体而言，尽管P85对细胞单层脑侧的转运系统有深远影响，但在脑微血管内皮细胞模型中，P85对氨基酸的AP至BL转运没有影响。
