1. Nile Red Quantifier: a novel and quantitative tool to study lipid accumulation in patient-derived circulating monocytes using confocal microscopy
- Author
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Erik S.G. Stroes, Feiko Tiessens, Guido J. Bakker, Jeffrey Kroon, Sophie J. Bernelot Moens, Johan G. Schnitzler, Max Nieuwdorp, Geesje M. Dallinga-Thie, Albert K. Groen, Center for Liver, Digestive and Metabolic Diseases (CLDM), Lifestyle Medicine (LM), Internal medicine, ACS - Diabetes & metabolism, Graduate School, Amsterdam Cardiovascular Sciences, Vascular Medicine, Amsterdam Gastroenterology Endocrinology Metabolism, Experimental Vascular Medicine, ACS - Atherosclerosis & ischemic syndromes, and ACS - Microcirculation
- Subjects
0301 basic medicine ,Pathology ,medicine.medical_specialty ,BLOOD ,lipid droplets ,Inflammation ,QD415-436 ,Biology ,Biochemistry ,DISEASE ,Monocytes ,law.invention ,03 medical and health sciences ,chemistry.chemical_compound ,Endocrinology ,INFLAMMATION ,Confocal microscopy ,law ,Lipid droplet ,Oxazines ,fluorescence and confocal imaging ,Methods ,medicine ,Humans ,FAMILIAL HYPERCHOLESTEROLEMIA ,cholesterol and trafficking ,Microscopy, Confocal ,Cholesterol ,CHOLESTEROL ,Nile red ,DROPLETS ,ASSOCIATION ,Cell Biology ,ANGINA-PECTORIS ,Lipids ,Molecular biology ,3. Good health ,Staining ,030104 developmental biology ,ATHEROSCLEROSIS ,chemistry ,NEUTRAL LIPIDS ,medicine.symptom ,Blood Chemical Analysis ,Ex vivo ,Intracellular - Abstract
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, however, 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. Circulating monocytes were isolated from blood and used for the NR staining procedure. In monocytes stained with NR, we clearly distinguished, based on 3D imaging, phospholipids and exclusively intracellular neutral lipids. Next, we developed and validated NRQ, a semi-automated quantification program that detects alterations in lipid accumulation. NRQ was able to detect LD alterations after ex vivo exposure of isolated monocytes to freshly isolated LDL in a time-and dose-dependent fashion. Finally, we validated NRQ in patients with familial hypercholesterolemia and obese subjects in pre- and postprandial state. In conclusion, NRQ is a suitable tool to detect even small differences in neutral LD content in circulating monocytes using NR staining.
- Published
- 2017
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