Study on the Function of Non-esterified Fatty Acid in the Process of THP-1 Derived Foam Cell Formation | Best Medical Research Papers For Sale

Study on the Function of Non-esterified Fatty Acid in the Process of THP-1 Derived Foam Cell Formation

21. June 2012 · Comments Off · Categories: Basic Medical · Tags: , , ,
Title Study on the Function of Non-esterified Fatty Acid in the Process of THP-1 Derived Foam Cell Formation
Abstract

Atherosclerosis (AS) is the buildup of plaque on the inside walls of arteries. The first stage in the development of atherosclerosis contains the formation of foam cells, which derived from macrophages in the artery wall. Fatty acid, elevated in visceral obesity, plays a vital role in atherogenesis and acute coronary syndromes. Increased circulating non-esterified fatty acids (NEFAs) in healthy subjects and in subjects with type 2 diabetes may contribute to activation of macrophages, and thus the development of AS. The circulating free fatty acids are mainly composed of lipid in diet. According to different chain length of fatty acids, NEFA can be divided into saturated fatty acid (SFA), monounsaturated fatty acid (MUFA), and polyunsaturated fatty acid (PUFA). NEFA can regulate expression changes of related genes during the formation of foam cell.The most representative and common NEFA in diet are palmitic acid (PA), oleic acid (OA), linoleic acid (LA) and eicosapentaenoic acid (EPA). They are prevalent in milk products and meat (PA), vegetable oils (OA, LA) and fish oils (EPA 20:5n-3). Four representative NEFAs different in saturation degree and chain length were employed in this study. PA (16:0), OA (18:1n-9), and LA (18:2n-6) are the most abundant fatty acids in fatty streaks and atheromatous plaques in humans. LA and EPA are two representative PUFAs.Atherosclerosis contains metabolism of TG, phospholipid, cholesterol and cholesterol esters. Expression changes of several related molecules are closely related with lipid accumulation and exertion. These fatty-acid-metabolism-related genes play crucial roles in intracellular lipid accumulation and efflux, and are involved in the mechanism of foam cell formation. PPARγ/LXR/ABCA1 pathway molecules such as PPARγ, ABCA1 and scavenger receptors, have long been evidenced to regulate the homeostasis of lipid in macrophage.Lipid droplet has a core consisting of neutral lipid and several lipid droplet associated proteins surrounding its surface. These lipid droplet associated proteins play important roles in lipid metabolism. Our previous work had identified that the cell death-inducing DFF45-like effecter (CIDE) family members and Perilipin-ADRP-TIP47 (PAT) family members had great effect on lipid formation in vitro. But the exact mechanism of lipid uptake and efflux in foam cells has not been deeply known.[Objectives]1. Human acute monocytic leukemia cell (THP-1) culture and induction of foam cell formation.2. Pretreatment of THP-1 derived macrophages by NEFAs and identification of the optimum culture conditions. 3. Morphology identification of lipid in foam cell and detection of lipid content.4. Quantitative detection and comparison of total cholesterol (TC) and triglyceride (TG) in foam cells.5. Detection and analysis on the expression profiles of lipid droplet associated molecules during THP-1 derived foam cell formation.6. Compare and analyze the different functions between NEFAs in foam cell formation of AS.[Methods]1. Induce THP-1 into macrophage by phorbol myristate acetate (PMA) and add oxidized low density lipoprotein (ox-LDL) to further induce foam cell formation in another 48h.2. Detect the effects of NEFAs under different duration and concentration on macrophage viability by MTT method. Draw survival cures of macrophages.3. Oil red O stained lipids in macrophages and foam cells and collected images by microscope. The lipid content in all kinds of NEFA groups and control group are measured by Image-Pro Plus 6.0 software.4. Detect TC and TG content in foam cells using Wako LabAssay kits.5. Detect and analyze the expression profiles of lipid droplet associated molecules during THP-1 derived foam cell formation by real-time PCR.[Results]1. We identified an optimum condition by using MTT assay. 100Μm and 72h can be reasonable for the incubation with NEFAs. Oil Red O staining and cytosolic TG and TC test showed that macrophage pretreated with NEFA could increase lipid content. PUFAs, including LA and EPA were more prone to result in lower lipid content than OA and PA did according to less increase of cytosolic total cholesterol (TC). PUFA could elevate PPARγlevel and enhance the expression of scavenger receptors, CD36 and SR-AI, but decrease ABCA1 level.2. The expression profiles of lipid droplet associated proteins, including CIDE and PAT family members were measured. Members of these proteins only coated on mature lipids, such as perilipin and Cidec, descended, but proteins mainly emerging on nascent lipid droplets, such as ADRP and TIP47, increased significantly. Our data indicated that PUFA can not only reduce the relative levels of TC, but also inhibit the maturation of lipid droplets and thus retard the formation of foam cells.[Conclusion]PPARγ/LXR/ABCA1 pathway molecules can regulate lipid metabolism in macrophages, and lipid droplet associated proteins have close correlation with cytosolic lipid content. A specific NEFA could enhance lipid accumulation in THP-1 derived macrophages. PUFA can relatively reduce TC content and retard foam cell formation.
Category Basic Medical
Keywords Atherosclerosis, Foam Cell, non-esterified fatty acid, THP-1,
FileType PDF
Pages 142
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