title: Consumption of a high-fat, high-calorie meal is associated with an increase in intracellular co-localization of PPAR-γ mRNA and protein in monocytes.
abstract: Acute and habitual dietary habits contribute to the onset and progression of many forms of cardiovascular disease. Circulating peripheral blood monocytes have been a target of pre-clinical research related to the risk of atherosclerosis. Specifically, when monocytes migrate into the subendothelial space and endocytosize modified LDL (i.e. acLDL or oxLDL) they phenotypically transform into foam cells. The endocytosis of modified LDL is mediated by the scavenger receptor CD36, whose expression is in tern regulated by the transcription factor PPAR-γ. In this report, we describe a novel technique for the simultaneous measurement of intracellular PPAR-γ mRNA and protein in peripheral blood monocytes collected from human subjects in fasted state or 3 and 5-h after consuming a high-calorie (65% of daily calorie needs), high-fat meal. Intracellular detection and co-localization of PPAR-γ was made possible using a combination of image-based flow cytometry (MilliporeSigma FlowSight) and an amplified mRNA FISH staining technique (Affymetrix/eBioscience PrimeFlow). Consumption of a high-calorie, high-fat meal increased the percentage of co-localization at both 3 and 5-h post prandial compared to pre-meal. No obvious difference in co-localization was observed when cells were treated by acLDL in vitro. More research is needed to determine how to best use this method to study pre-clinical risk of atherosclerosis.
authors.affiliation: University of North Texas, Applied Physiology Laboratory, United States; University of North Texas, Department of Biological Sciences, United States.
authors.forename: Andrea L
authors.initials: AL
authors.lastname: Henning
authors.affiliation: University of North Texas, Applied Physiology Laboratory, United States; University of North Texas, Department of Biological Sciences, United States. Electronic address: brian.mcfarlin@unt.edu.
authors.forename: Brian K
authors.initials: BK
authors.lastname: McFarlin
mesh_terms: D000906:Antibodies / Q000737:chemistry; D018955:CD36 Antigens / Q000235:genetics / Q000378:metabolism; D059305:Diet, High-Fat / Q000009:adverse effects; D004041:Dietary Fats / Q000008:administration & dosage*; D005247:Feeding Behavior; D005434:Flow Cytometry / Q000379:methods*; D015870:Gene Expression; D006801:Humans; D019044:Image Cytometry / Q000379:methods*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D047495:PPAR gamma / Q000819:agonists / Q000235:genetics* / Q000378:metabolism; D061251:Primary Cell Culture; D012333:RNA, Messenger / Q000819:agonists / Q000235:genetics / Q000378:metabolism; D013194:Staining and Labeling / Q000379:methods; D015854:Up-Regulation
chemical_list: D000906:Antibodies; D018955:CD36 Antigens; D004041:Dietary Fats; D008077:Lipoproteins, LDL; D047495:PPAR gamma; D012333:RNA, Messenger; C099252:oxidized low density lipoprotein
keywords: Atherosclerosis; Flow cytometry; Foam cells; Lipoproteins; Monocytes