Eicosapentaenoic acid membrane incorporation impairs ABCA1-dependent cholesterol efflux via a protein kinase A signaling pathway in primary human macrophages.

A diet rich in n-3/n-6 polyunsaturated fatty acids (PUFAs) is cardioprotective. Dietary PUFAs affect the cellular phospholipids composition, which may influence the function of membrane proteins. We investigated the impact of the membrane incorporation of several PUFAs on ABCA1-mediated cholesterol efflux, a key antiatherogenic pathway. Arachidonic acid (AA) (C20:4 n-6) and docosahexaenoic acid (DHA) (C22:6 n-3) decreased or increased cholesterol efflux from J774 mouse macrophages, respectively, whereas they had no effect on efflux from human monocyte-derived macrophages (HMDM). Importantly, eicosapentaenoic acid (EPA) (C20:5 n-3) induced a dose-dependent reduction of ABCA1 functionality in both cellular models (-28% for 70µM of EPA in HMDM), without any alterations in ABCA1 expression. These results show that PUFA membrane incorporation does not have the same consequences on cholesterol efflux from mouse and human macrophages. The EPA-treated HMDM exhibited strong phospholipid composition changes, with high levels of both EPA and its elongation product docosapentaenoic acid (DPA) (C22:5 n-3), which is associated with a decreased level of AA. In HMDM, EPA reduced the ATPase activity of the membrane transporter. Moreover, the activation of adenylate cyclase by forskolin and the inhibition of cAMP phosphodiesterase by isobutylmethylxanthine restored ABCA1 cholesterol efflux in EPA-treated human macrophages. In conclusion, EPA membrane incorporation reduces ABCA1 functionality in mouse macrophages as well as in primary human macrophages and this effect seems to be PKA-dependent in human macrophages.

Mycophenolate Mofetil and Rapamycin Induce Apoptosis in the Human Monocytic U937 Cell Line Through Two Different Pathways.

Transplant vasculopathy may be considered as an accelerated form of atherosclerosis resulting in chronic rejection of vascularized allografts. After organ transplantation, a diffuse intimal thickening is observed, leading to the development of an atherosclerosis plaque due to a significant monocyte infiltration. This results from a chronic inflammatory process induced by the immune response. In this study, we investigated the impact of two immunosuppressive drugs used in therapy initiated after organ transplantation, mycophenolate mofetil, and rapamycin, on the apoptotic response of monocytes induced or not by oxidized LDL. Here we show the pro-apoptotic effect of these two drugs through two distinct signaling pathways and we highlight a synergistic effect of rapamycin on apoptosis induced by oxidized LDL. In conclusion, since immunosuppressive therapy using mycophenolate mofetil or rapamycin can increase the cell death in a monocyte cell line, this treatment could exert similar effects on human monocytes in transplant patients, and thus, prevent transplant vasculopathy, atherosclerosis development, and chronic allograft rejection. J. Cell. Biochem. 118: 3480-3487, 2017. © 2017 Wiley Periodicals, Inc.

Eicosapentaenoic acid membrane incorporation impairs cholesterol efflux from cholesterol-loaded human macrophages by reducing the cholesteryl ester mobilization from lipid droplets.

A diet containing a high n-3/n-6 polyunsaturated fatty acids (PUFA) ratio has cardioprotective properties. PUFAs incorporation into membranes influences the function of membrane proteins. We investigated the impact of the membrane incorporation of PUFAs, especially eicosapentaenoic acid (EPA) (C20:5 n-3), on the anti-atherogenic cholesterol efflux pathways. We used cholesteryl esters (CE)-loaded human monocyte-derived macrophages (HMDM) to mimic foam cells exposed to the FAs for a long period of time to ensure their incorporation into cellular membranes. Phospholipid fraction of EPA cells exhibited high levels of EPA and its elongation product docosapentaenoic acid (DPA) (C22:5 n-3), which was associated with a decreased level of arachidonic acid (AA) (C20:4 n-6). EPA 70µM reduced ABCA1-mediated cholesterol efflux to apolipoprotein (apo) AI by 30% without any alteration in ABCA1 expression. The other tested PUFAs, DPA, docosahexaenoic acid (DHA) (C22:6 n-3), and AA, were also able to reduce ABCA1 functionality while the monounsaturated oleic FA slightly decreased efflux and the saturated palmitic FA had no impact. Moreover, EPA also reduced cholesterol efflux to HDL mediated by the Cla-1 and ABCG1 pathways. EPA incorporation did not hinder efflux in free cholesterol-loaded HMDM and did not promote esterification of cholesterol. Conversely, EPA reduced the neutral hydrolysis of cytoplasmic CE by 24%. The reduced CE hydrolysis was likely attributed to the increase in cellular TG contents and/or the decrease in apo E secretion after EPA treatment. In conclusion, EPA membrane incorporation reduces cholesterol efflux in human foam cells by reducing the cholesteryl ester mobilization from lipid droplets.

Impact of peri-urban landscape on the organic and mineral contamination of pond waters and related risk assessment.

Ponds are important for their ecological value and for the ecosystem services they provide to human societies, but they are strongly affected by human activities. Peri-urban development, currently one of the most pervasive processes of land use change in Europe, exposes ponds to both urban and agricultural contaminants, causing a potential combination of adverse effects. This study, focused on 12 ponds located in a peri-urban area, has two main objectives: (1) to link the physico-chemical characteristics of the waters and the nature of their contaminants, either organic or mineral, with the human activities around ponds, and (2) to estimate the environmental risk caused by these contaminants. The ponds were sampled during two consecutive years in both spring and in autumn. Although the ponds were distributed over a limited geographical area, their contamination profiles were different and more correlated with the agricultural than the urban land use. In terms of aptitude for biology, half of the ponds were classified in degraded states due to their physico-chemical parameters, but without correlation with the endocrine disrupting activities and the levels of organic pollutants as indicators. The main quantified organic pollutants, however, were pesticides with sufficiently high levels in certain cases to induce an environmental risk exceeding the classical thresholds of risk quotient.

Contrasting effects of membrane enrichment with polyunsaturated fatty acids on phospholipid composition and cholesterol efflux from cholesterol-loaded J774 mouse or primary human macrophages.

A high consumption of polyunsaturated fatty acids (PUFAs), particularly n-3 PUFAs, is atheroprotective. PUFAs incorporation into membrane phospholipids alters the functionality of membrane proteins. We studied the consequences of the in vitro supplementation of several PUFAs on the FA profiles and on ABCA1-dependent cholesterol efflux capacities from cholesterol-loaded macrophages. Arachidonic acid (AA, C20:4 n-6) and, to a lesser extent, eicosapentaenoic acid (EPA, C20:5 n-3), dose-dependently impaired cholesterol efflux from cholesterol-loaded J774 mouse macrophages without alterations in ABCA1 expression, whereas docosahexaenoic acid (DHA, C22:6 n-3) had no impact. AA cells exhibited higher proportions of arachidonic acid and adrenic acid (C22:4 n-6), its elongation product. EPA cells exhibited slightly higher proportions of EPA associated with much higher proportions of docosapentaenoic acid (C22:5 n-3), its elongation product and with lower proportions of AA. Conversely, both EPA and DHA and, to a lesser extent, AA decreased cholesterol efflux from cholesterol-loaded primary human macrophages (HMDM). The differences observed in FA profiles after PUFA supplementations were different from those observed for the J774 cells. In conclusion, we are the first to report that AA and EPA, but not DHA, have deleterious effects on the cardioprotective ABCA1 cholesterol efflux pathway from J774 foam cells. Moreover, the membrane incorporation of PUFAs does not have the same impact on cholesterol efflux from murine (J774) or human (HMDM) cholesterol-loaded macrophages. This finding emphasizes the key role of the cellular model in cholesterol efflux studies and may partly explain the heterogeneous literature data on the impact of PUFAs on cholesterol efflux.

Investigation of lipid modifications in J774 macrophages by vibrational spectroscopies after eicosapentaenoic acid membrane incorporation in unloaded and cholesterol-loaded cells.

Atherosclerosis is an inflammatory disease of the arterial wall caused by the formation of an atheroma plaque in the vessel wall. The uptake of modified LDL lipoproteins by sub-endothelial macrophages induces the latter's transformation into foam cells, which is the key step of atheroma plaque formation. The modifications of neutral lipids caused by foam cells formation are marked by the appearance of lipid droplets. Polyunsaturated fatty acids (PUFAs) incorporation into membrane phospholipids (PL) modifies their composition, which may influence membrane protein functions. The incorporation of eicosapentaenoic acid (EPA) reduces the anti-atherogenic ABCA1 (ATP Binding Cassette transporter A1) pathway and induces PLs modifications. In order to study lipids directly in the cell environment, a comparative study is conducted by vibrational spectroscopies on murine macrophages J774, loaded or not with cholesterol, which were enriched or not with eicosapentaenoic acid (EPA). The study enabled to identify changes in the spectral signature after cells enrichment with fatty acid (FA) relying only on chemometric analysis without deuterium labelling. Results highlighted spectral changes in the regions attributed to lipids associated to triglycerides, phospholipids and cholesterol in both Raman and IR.