Abstract
Aim: The inflammatory profile of circulating monocytes is an important biomarker for atherosclerotic plaque vulnerability. Recent research revealed that peripheral lipid uptake by monocytes alters their phenotype towards an inflammatory state and this coincides with an increased lipid droplet (LD) content. Determination of lipid content of circulating monocytes is, however, not very well established. Based on Nile Red (NR) LD-imaging using confocal microscopy and computational analysis, we developed Nile Red Quantifier (NRQ); a novel quantification method to assess lipid droplet content in monocytes.
Highlights
The inflammatory profile of circulating monocytes is an important biomarker for atherosclerotic plaque vulnerability
To test whether Nile Red (NR) staining was equivalent to Oil Red O (ORO), we examined lipid accumulation in isolated monocytes and stained lipid droplet (LD) using both ORO and NR
This revealed a dose-dependent increase in cholesterol ester (CE)/ TG ratio (CE/TG 3.46, 8.28 vs. 1.00-fold for 1, 2 vs. 0 mg/ml LDL, respectively; supplemental Fig. S1B)
Summary
The inflammatory profile of circulating monocytes is an important biomarker for atherosclerotic plaque vulnerability. Recent research revealed that peripheral lipid uptake by monocytes alters their phenotype toward an inflammatory state and this coincides with an increased lipid droplet (LD) content. Determination of lipid content of circulating monocytes is, not very well established. Based on Nile Red (NR) neutral LD imaging, using confocal microscopy and computational analysis, we developed NR Quantifier (NRQ), a novel quantification method to assess LD content in monocytes. In monocytes stained with NR, we clearly distinguished, based on 3D imaging, phospholipids and exclusively intracellular neutral lipids. We developed and validated NRQ, a semi-automated quantification program that detects alterations in lipid accumulation. NRQ is a suitable tool to detect even small differences in neutral LD content in circulating monocytes using NR staining.—Schnitzler, J. Nile Red Quantifier: a novel and quantitative tool to study lipid accumulation in patient-derived circulating monocytes using confocal microscopy J.
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