Aβ40 Reduces P-Glycoprotein at the Blood–Brain Barrier through the Ubiquitin–Proteasome Pathway

Abstract

Failure to clear amyloid-␤ (A␤) from the brain is in part responsible for A␤ brain accumulation in Alzheimer's disease (AD).A critical protein for clearing A␤ across the blood-brain barrier is the efflux transporter P-glycoprotein (P-gp) in the luminal plasma membrane of the brain capillary endothelium. P-gp is reduced at the blood-brain barrier in AD, which has been shown to be associated with A␤ brain accumulation. However, the mechanism responsible for P-gp reduction in AD is not well understood. Here we focused on identifying critical mechanistic steps involved in reducing P-gp in AD. We exposed isolated rat brain capillaries to 100 nM A␤40, A␤40, aggregated A␤40, and A␤42. We observed that only A␤40 triggered reduction of P-gp protein expression and transport activity levels; this occurred in a dose-and time-dependent manner. To identify the steps involved in A␤-mediated P-gp reduction, we inhibited protein ubiquitination, protein trafficking, and the ubiquitin-proteasome system, and monitored P-gp protein expression, transport activity, and P-gpubiquitin levels. Thus, exposing brain capillaries to A␤40 triggers ubiquitination, internalization, and proteasomal degradation of P-gp. These findings may provide potential therapeutic targets within the blood-brain barrier to limit P-gp degradation in AD and improve A␤ brain clearance.

Caloric restriction preserves memory and reduces anxiety of aging mice with early enhancement of neurovascular functions

“…P-gp is an ATP-driven transporter highly expressed at the BBB that facilitates clearance of Aβ, a hallmark of AD. We previously established a confocal imaging-based assay to assess P-gp transport activity in freshly isolated brain capillaries from mice [26, 27]. This assay measures within capillary lumens accumulation of [N-ε(4-nitro-benzofurazan-7-yl)-D-Lys(8)]-cyclosporin A (NBD-CSA), a fluorescent P-glycoprotein substrate.…”

Lymphatics in Neurological Disorders: A Neuro-Lympho-Vascular Component of Multiple Sclerosis and Alzheimer’s Disease?

“…Efflux of Aβ from the ISF into the blood is largely influenced by the expression of p-glycoprotein (Pgp) and low-density lipoprotein receptor-related protein 1 (LRP1) by the vascular endothelial cells (Erickson et al, 2012; Hartz et al, 2016; Shibata et al, 2000). Pgp is less efficient than LRP1 in promoting Aβ excretion; it is possible (though unconfirmed) that Pgp modulates the abluminal efflux of Aβ by boosting the interaction between Apo-E and Aβ in the vicinity of the blood vessels, thereby indirectly promoting the excretion of Apo-E/Aβ complexes through binding to LRP1 (Akanuma et al, 2008; Fitz et al, 2012; Lam et al, 2001).…”

Selective induction of P-glycoprotein at the CNS barriers during symptomatic stage of an ALS animal model

“…NBD-CSA was custom synthesized [3, 8, 12]. Bodipy-Prazosin was purchased from Life Technologies (Grand Island, NY).…”

“…Brain capillary isolation protocol was adopted previously with the following minor modification [12]. Cerebral cortex grey matter and spinal cord tissues were homogenized 40 up-and-down strokes using a tissue grinder (clearance: 150–230 µm) rotating at 50 rpm in ice-cold incubation buffer [2.7 mM KCl, 1.46 mM KH 2 PO 4 , 136.9 mM NaCl, 8.1 mM Na 2 HPO 4 , 0.9 mM CaCl 2 , 0.5 mM MgCl 2 , 5 mM D-Glucose and 1 mM Na + -pyruvate at pH 7.4].…”