(191dn) Maximizing P-Glycoprotein Expression and Transport in the Presence of Therapeutic Compounds

In 2004, a study of brain tissue obtained from elderly patients showed a local inverse correlation for the expression of P‑glycoprotein (P-gp), a blood-brain barrier (BBB) transport protein, and the deposition of aggregated amyloid-β protein (Aβ), a hallmark of Alzheimer’s disease (AD). The most common form of neurodegenerative disorder, AD affects over 5 million people and is the 6th leading cause of death in the United States. AD is characterized by an accumulation of Aβ in the brain. Currently under study as a therapeutic approach is the transport of excess Aβ out of the brain through the single endothelial cell thickness of the BBB. Our lab has shown that P-gp can transport oligomeric aggregates of Aβ. While monomeric Aβ is inert, oligomeric Aβ exhibits neurotoxicity and leads to the formation of Aβ fibrils that deposit as amyloid plaques, which are characteristic of AD brain. However, formation of oligomeric Aβ may be important to its clearance from the brain.

It has been recently discovered that the presence of certain forms of Aβ in the cells of the BBB can reduce P-gp levels through the ubiquitin-proteasome degradation pathway. These findings identify P-gp as a novel therapeutic target for AD. This study seeks to identify therapeutic compounds that will increase the expression levels of P-gp at the blood-brain barrier in order to mitigate the accumulation of Aβ by allowing for increased clearance of Aβ from the brain.

Madin-Darby Canine Kidney (MDCK) epithelial cells continuously express the multidrug resistance 1 (MDR1) gene responsible for the presence of P-gp. Cells were treated with three prospective therapeutic compounds: verapamil, caffeine, and rifampicin. Verapamil is a known P-gp substrate recently suggested to up-regulate P-gp expression and has exhibited a neuroprotective effect on AD model rats. Rifampicin, an antibiotic used to treat tuberculosis and leprosy, and caffeine have been shown in epidemiological studies to reduce dementia symptoms. Following treatment, alteration of P-gp expression was determined using fluorescent microscopy to visualize cell membrane location of P-gp and SDS-PAGE with Western blotting to quantify P-gp expression. A cell transport assay was used to quantify the transport of Aβ the treated MDCK cells. Results supporting increased P-gp expression and Aβ transport highlight the prospective therapeutic potential of these compounds.