The Identification and Investigation of In-vitro Blood-Brain Barrier Integrity Enhancers

摘要

Alzheimer's disease (AD) is known for its hallmark features such as increased levels of tau hyperphosphorylation and amyloid-beta (Aβ) deposition. Cerebral vascular dysfunction constitutes an important feature of AD as well, which includes amyloid angiopathy and impaired clearance of Aβ across the blood-brain barrier (BBB). In the search for potential drugs that may enhance or maintain the BBB integrity and function, we screened the Sigma Lopac??1280 compound library using a high-throughput screening (HTS) assay developed in our laboratory. In the process of identifying a hit compound, an in-vitro BBB model monolayer utilizing bEnd3 cells was constructed. Compounds were first identified as hits if they were able to enhance the monolayer's integrity by decreasing Lucifer Yellow (LY) permeation. Next, hit compounds were secondary screened for their effect on BBB model function. Utilizing the same cell line, western blot analysis for major transport proteins of Aβ and the tight junction proteins ZO1, Occludin, and Claudin-5 was performed. Finally Aβ transport study was conducted to measure the effect of hit compounds on Aβ transport across the membrane. The HTS assay identified 3 unique compounds that decreased LY permeation, indicating enhancement of the in-vitro BBB model integrity. Western blot analysis showed these hit compounds to deferentially alter transport and tight junction protein expression indicating increased phenotypic function. In addition, hit compounds enhanced the Aβ transport across the monolayer in this model. In conclusion, these hit compounds are compelling candidates for further in vivo investigation in AD animal models.