A high-content microscopy drug screening platform for regulators of the extracellular digestion of lipoprotein aggregates by macrophages

Abstract

The recruitment of macrophages to the intima of arteries is a critical event in atherosclerotic progression. These macrophages accumulate excessive lipid droplets and become "foam cells", a hallmark of atherosclerosis. Most studies focus on lipid accumulation through macrophage interaction with modified monomeric low-density lipoprotein (LDL). However, in the intima, macrophages predominantly encounter aggregated LDL (agLDL), an interaction that has been studied far less. Macrophages digest agLDL and generate free cholesterol in an extracellular, acidic, hydrolytic compartment. They form a tight seal around agLDL through actin polymerization and deliver lysosomal contents into this space in a process termed digestive exophagy. There is some evidence that inhibiting digestive exophagy to slow cholesterol accumulation in macrophages protects them from becoming foam cells and slows the progression of atherosclerotic lesions. Thus, understanding the mechanisms of digestive exophagy is critical. Here, we describe a high-content microscopy screen on a library of repurposed drugs for compounds that inhibit lysosome exocytosis during digestive exophagy. We identified many hit compounds and further characterized the effects that five of these compounds have on various aspects of digestive exophagy. In addition, three of the five compounds do not inhibit oxidized LDL-induced foam cell formation, indicating the two pathways to foam cell formation can be targeted independently. We demonstrate that this high-content screening platform has great potential as a drug discovery tool with the ability to effectively and efficiently screen for modulators of digestive exophagy.