Visualizing Cholesterol Uptake by Self-Assembling Rhodamine B-Labeled Polymer Inside Living Cells via FLIM-FRET Microscopy

Abstract

Atherosclerosis is a widespread and hazardous disease characterized by the formation of arterial plaques mostly composed of fat, cholesterol, and calcium ions. The direct solubilization of cholesterol represents a promising, atheroprotective strategy to subside lipid blood levels and reverse atherosclerosis. This study deals with the in-depth analysis of polymer-mediated cholesterol dissolution inside living human cells. To this end, a recently described multifunctional block-polymer is labeled with Rhodamine B (RhoB) to investigate its interaction with cells via fluorescence microscopy. This gives insight into the cellular internalization process of the polymer, which appears to be clathrin- and caveolae/raft-dependent endocytosis. In cell single particle tracking reveals an active transport of RhoB polymer including structures. Förster resonance energy transfer (FRET) measurements of cells treated with a fluorophore-tagged cholesterol derivative and the RhoB polymer indicates the uptake of cholesterol by the polymeric particles. Hence, these results present a first step toward possible applications of cholesterol-absorbing polymers for treating atherosclerosis.