Mechanisms of U87 Astrocytoma Cell Uptake and Trafficking of Monomeric versus Protofibril Alzheimer’s Disease Amyloid-β Proteins

Abstract

A significant hallmark of Alzheimer’s disease is the formation of senile plaques in the brain due to the unbalanced levels of amyloid-beta (Aβ). However, although how Aβ is produced from amyloid precursor proteins is well understood, little is known regarding the clearance and metabolism of various Aβ aggregates from the brain. Similarly, little is known regarding how astrocytes internalize and degrade Aβ, although astrocytes are known to play an important role in plaque maintenance and Aβ clearance. The objective of this study is to investigate the cellular mechanisms that mediate the internalization of soluble monomeric versus oligomeric Aβ by astrocytes. We used a combination of laser confocal microscopy and genetic and pharmacological experiments to dissect the internalization of sAβ42 and oAβ42 and their postendocytic transport by U87 human brain astrocytoma cell line. Both Aβ42 species were internalized by U87 cells through fluid phase macropinocytosis, which required dynamin 2. Depleting LDL receptor-related protein 1 (LRP1) decreased sAβ42 uptake more significantly than that of oAβ42. We finally show that both Aβ42 species were rapidly transported to lysosomes through an endolytic pathway and subjected to proteolysis after internalization, which had no significant toxic effects to the U87 cells under relatively low concentrations. We propose that macropinocytic sAβ42 and oAβ42 uptake and their subsequent proteolytic degradation in astroglial cells is a significant mechanism underlying Aβ clearance from the extracellular milieu. Understanding the molecular events involved in astrocytic Aβ internalization may identify potential therapeutic targets for Alzheimer’s disease.

Role of LRP1 in the pathogenesis of Alzheimer's disease: evidence from clinical and preclinical studies

“…LRP1 might facilitate A uptake into cells through cooperating with other A-binding proteins, such as heparan sulfate proteoglycan (17,18). Oligomeric or aggregated A is also a poor ligand for LRP1 (15,19). Of note, the binding of apoE to LRP1 appears to be affected by the conformational and lipidation status of apoE (1).…”

“…The latter point might be more important in a subset of AD patients whose BBB integrity is disturbed (104) and, thus, circulating sLRP1 might enter the brain. Cumulatively, LRP1 expressed in neurons (17,73,75), astrocytes (19,76,105), microglia (106), endothelial cells (16,78,84), vascular smooth muscle cells or pericytes (88)(89)(90)107), choroid plexus (87,108), plasma (32), and liver (97-99) was reported to be involved in A cellular uptake and/or clearance (Fig. 2).…”

Anticancer properties of lipid and poly(ε-caprolactone) nanocapsules loaded with ferrocenyl-tamoxifen derivatives

“…Some of these mechanisms have been reported for U87 (e.g. phagocytosis) and for MDA‐MB‐231 (e.g. clathrin‐ and caveolae‐mediated pathways).…”

Immunoreactivities for hepcidin, ferroportin, and hephaestin in astrocytes and choroid plexus epithelium of human brains

“…In addition, it was suggested that immunoreactivity for ferroportin was located in association with endosomal structures, possibly macropinosomes, of reactive astrocytes. Recent experimental findings on the clearance of intracerebral substances suggest that proteolytic degradation of soluble amyloid‐beta (Aβ) in macropinosomes of astrocytic cells is a significant mechanism underlying Aβ clearance . Accordingly, it is likely that these proteins may also regulate intracellular iron levels in astrocytes.…”