Scavenging of Alzheimer's amyloid ?-protein by microglia in culture

Abstract

Deposits of amyloid β-protein (Aβ) form the cores of the pathological plaques which characterize Alzheimer's disease. The mechanism of formation of the deposits is unknown; one possibility is failure of a clearance mechanism that would normally remove the protein from brain parenchyma. This study has investigated the capacity of the central nervous system (CNS) phagocytes, microglia cells, to clear exogenous Aβ_1–42 from their environment. Cultured microglia from adult rat CNS have a high capacity to remove Aβ from serum-free medium, shown by immunoblotting experiments. Aβ from incubation medium was attached to the cell surface and could be identified by immunocytochemistry at the light or electron microscopic (EM) level; by EM, Aβ also appeared in phagosome-like intracellular vesicles. Light microscopic immunocytochemistry combined with computer-assisted image analysis showed that cells accumulated Aβ within 24 hr. from culture medium containing from 1 to 20 μg/ml Aβ. Microglial accumulation of Aβ was substantially reduced in the presence of fetal bovine serum. Addition of the protease inhibitor leupeptin to incubation medium with serum resulted in accumulation of Aβ in a membrane-bound intracellular compartment, but not at the cell surface. The increase in intracellular accumulation in the presence of the protease inhibitor indicates a microglial capacity for intracellular degradation of Aβ in the absence of inhibition. The change from predominantly cell-surface accumulation in serum-free medium to predominantly intracellular accumulation with serum may be explained by the presence in serum of carrier proteins that complex with Aβ and target it to cell surface receptors capable of stimulating endocytosis. Microglia were also cultured on unfixed cryostat sections of human brain tissue containing Alzheimer's plaques. Very little Aβ from the tissue was accumulated by the cells, although cultured microglia were found in direct contact with anti-Aβ immunopositive plaques. Possibly Aβ in tissue sections was complexed with other proteins which either inhibited its uptake by microglia or enhanced its proteolysis, preventing cellular accumulation of immunostainable Aβ. The results indicate that cultured microglia effectively remove Aβ from tissue culture medium and from the surface of the dish and concentrate monomer and aggregates of Aβ either on the cell surface or intracellularly. This process may be modified by proteins present in Alzheimer's brain sections.