(338f) Amyloid-Beta Induced Endothelial-Monocyte Interactions Involved in Cerebral Amyloid Angiopathy and Alzheimer's Disease

Authors: 
Moss, M. A., University of South Carolina
Gonzalez, F. J., University of South Carolina
Barcelo, A. R., University of South Carolina


Cerebral amyloid angiopathy (CAA) associated with Alzheimer's disease (AD) and hereditary cerebral hemorrhage with amyloidosis (HCHWA) is characterized by deposition of the fibrillar form of the amyloid-beta protein in the cerebral vasculature. Monomeric amyloid-beta self-associates to form the fibrillar amyloid-beta that is deposited. Vascular accumulation of fibrillar amyloid-beta occurs primarily in the basement membrane and initiates a cascade of events culminating in a weakening of the vessel wall that may lead to stroke as a result of cerebral hemorrhage. Among these events is an increase in vessel-associated monocyte/macrophage lineage cells,1,2 which could promote vascular damage and may contribute to the elevated immune response observed in AD brain. Cell culture experiments have shown that the presence of amyloid-beta aggregation mixtures enhances monocyte adhesion to endothelial monolayers and subsequent transendothelial migration.3 Yet, the mechanism for these interactions remains unknown.

We have investigated the ability of direct interactions between amyloid-beta(1-40) and endothelial monolayers to enhance adhesion and subsequent transendothelial migration of monocyte cells. Confluent human umbilical vein endothelial cell (HUVEC) monolayers were grown on gelatin-coated 96-well plates for adhesion assays, or gelatin-coated polyethylene terephthalate (PET) membrane inserts for transendothelial migration assays. Confluent monolayers were incubated with varying concentrations of amyloid-beta(1-40) reaction mixtures, mature amyloid-beta(1-40) fibrils, or unaggregated amyloid-beta(1-40) monomer. Subsequently, monolayers were washed and exposed to Calcein-labeled THP-1 monocytes. Cells remaining adherent following a series of washes or cells that had transmigrated through membrane inserts were quantified using Calcein fluorescence.

Experimental data demonstrates that treatment of endothelial monolayers with aggregation mixtures of amyloid-beta(1-40), containing mature fibrils, protofibril intermediates, and unassembled monomer, enhances the adhesion of THP-1 monocytes and also promotes subsequent transendothelial migration. In contrast, treatment of endothelial monolayers with either amyloid-beta(1-40) fibril alone or amyloid-beta(1-40) monomer alone have no effect. Parallel toxicity experiments verify that amyloid-beta-induced changes in endothelial adhesion and transmigration cannot be attributed to toxicity and subsequent disruption of endothelial monolayers. Together, these results demonstrate that the presence of aggregated amyloid-beta alone is insufficient to stimulate endothelium for monocyte adhesion. Instead, the presence of smaller aggregates, monomer, or the combination present in the undefined reaction mixture is required. These observations are in agreement with other studies which have suggested that amyloid-beta-induced vascular responses correlate with amyloid-beta assembly,4-6 require the presence of both amyloid-beta monomer and aggregate,7 and are observed only during active amyloid-beta polymerization.6 Future work will focus upon identifying the assembly events that promote amyloid-beta(1-40) stimulation of endothelial monolayers.

As the contribution of CAA in the progression of AD continues to gain support, therapeutic strategies should begin to encompass this vital facet of the disease. Resolution of the interactions between vascular amyloid-beta deposits and the endothelium will distinguish potential therapeutic targets for the reduction of vascular damage associated with CAA and AD.

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