背景:患有阿尔茨海默病(AD)的人通常需要许多药物;然而,这些药物使用推荐用于无AD患者的方案给药.尽管AD中血脑屏障(BBB)中P-糖蛋白(P-gp)的丰度和功能降低,这会影响大脑暴露的药物。导致散发性AD中P-gp丰度降低的基本机制仍然未知;然而,已知载脂蛋白E(apoE)基因与散发性AD的发生具有最强的遗传联系,和apoE亚型可以不同地改变BBB功能。这项研究的目的是使用人脑微血管内皮细胞(hCMEC/D3)模型评估apoE是否以同工型依赖性方式影响P-gp丰度和功能。
方法:本研究评估了apoE亚型对hCMEC/D3细胞中P-gp丰度(通过蛋白质印迹)和功能(通过罗丹明123(R123)摄取)的影响。将细胞以2-10μg/mL暴露于重组apoE3和apoE424-72小时。hCMEC/D3细胞也暴露于来自表达人源化apoE同种型的星形胶质细胞的星形胶质细胞条件培养基(ACM)72小时。
结果:相对于重组apoE3,重组apoE4没有改变hCMEC/D3细胞中的P-gp丰度,含有人apoE亚型的ACM也没有改变P-gp丰度。重组apoE同种型处理在hCMEC/D3细胞中的R123积累也没有改变,表明P-gp功能没有变化,尽管丰度和功能都被阳性对照SR12813(5µM)和PSC833(5µM)改变,分别。
结论:在该模型中,不同的apoE同工型对P-gp丰度或功能没有直接影响,需要进一步的体内研究来解决散发性AD中P-gp丰度或功能是否以apoE同工型特异性方式降低。
BACKGROUND: Individuals with Alzheimer\'s disease (AD) often require many medications; however, these medications are dosed using regimens recommended for individuals without AD. This is despite reduced abundance and function of P-glycoprotein (P-gp) at the blood-brain barrier (BBB) in AD, which can impact brain exposure of drugs. The fundamental mechanisms leading to reduced P-gp abundance in sporadic AD remain unknown; however, it is known that the apolipoprotein E (apoE) gene has the strongest genetic link to sporadic AD development, and apoE isoforms can differentially alter BBB function. The aim of this study was to assess if apoE affects P-gp abundance and function in an isoform-dependent manner using a human cerebral microvascular endothelial cell (hCMEC/D3) model.
METHODS: This study assessed the impact of apoE isoforms on P-gp abundance (by western blot) and function (by rhodamine 123 (R123) uptake) in hCMEC/D3 cells. Cells were exposed to recombinant apoE3 and apoE4 at 2 - 10 µg/mL over 24 - 72 hours. hCMEC/D3 cells were also exposed for 72 hours to astrocyte-conditioned media (ACM) from astrocytes expressing humanised apoE isoforms.
RESULTS: P-gp abundance in hCMEC/D3 cells was not altered by recombinant apoE4 relative to recombinant apoE3, nor did ACM containing human apoE isoforms alter P-gp abundance. R123 accumulation in hCMEC/D3 cells was also unchanged with recombinant apoE isoform treatments, suggesting no change to P-gp function, despite both abundance and function being altered by positive controls SR12813 (5 µM) and PSC 833 (5 µM), respectively.
CONCLUSIONS: Different apoE isoforms have no direct influence on P-gp abundance or function within this model, and further in vivo studies would be required to address whether P-gp abundance or function are reduced in sporadic AD in an apoE isoform-specific manner.