Bis(monoacylglycero)phosphate regulates oxysterol binding protein-related protein 11 dependent sterol trafficking

Bis(Monoacylglycero) Phosphate (BMP) is a unique phospholipid localized in late endosomes, a critical cellular compartment in low density lipoprotein (LDL)-cholesterol metabolism. In previous work, we demonstrated the important role of BMP in the regulation of macrophage cholesterol homeostasis. BMP exerts a protective role against the pro-apoptotic effect of oxidized LDL (oxLDL) by reducing the production of deleterious oxysterols. As the intracellular sterol traffic in macrophages is in part regulated by oxysterol binding protein (OSBP) and OSBP-related proteins (ORPs), we investigated the role of ORP11, localized at the Golgi-late endosomes interface, in the BMP-mediated protection from oxLDL/oxysterol cytotoxicity. Stably silencing of ORP11 in mouse RAW264.7 macrophages via a shRNA lentiviruses system had no effect on BMP production. However, ORP11 knockdown abrogated the protective action of BMP against oxLDL induced apoptosis. In oxLDL treated control cells, BMP enrichment was associated with reduced generation of 7-oxysterols, while these oxysterol species were abundant in the ORP11 knock-down cells. Of note, BMP enrichment in ORP11 knock-down cells was associated with a drastic increase in free cholesterol and linked to a decrease of cholesterol efflux. The expression of ATP-binding cassette-transporter G1 (ABCG1) was also reduced in the ORP11 knock-down cells. These observations demonstrate a cooperative function of OPR11 and BMP, in intracellular cholesterol trafficking in cultured macrophages. We suggest that BMP favors the egress of cholesterol from late endosomes via an ORP11-dependent mechanism, resulting in a reduced production of cytotoxic 7-oxysterols.Peer reviewe

Bis(monoacylglycero)phosphate, an important actor in the host endocytic machinery hijacked by SARS-CoV-2 and related viruses

International audienceViruses, including the novel coronavirus SARS-CoV-2, redirect infected cell metabolism to their own purposes. After binding to its receptor angiotensin-converting enzyme 2 (ACE2) on the cell surface, the SARS-CoV-2 is taken up by receptor-mediated endocytosis ending in the acidic endolysosomal compartment. The virus hijacks the endosomal machinery leading to fusion of viral and endosomal membranes and release of the viral RNA into the cytosol. This mini-review specifically highlights the membrane lipid organization of the endosomal system focusing on the unconventional and late endosome/lysosome-specific phospholipid, bis(monoacylglycero)phosphate (BMP). BMP is enriched in alveolar macrophages of lung, one of the target tissue of SARS-CoV-2. This review details the BMP structure, its unsaturated fatty acid composition and fusogenic properties that are essential for the highly dynamic formation of the intraluminal vesicles inside the endosomes. Interestingly, BMP is necessary for infection and replication of enveloped RNA virus such as SARS-CoV-1 and Dengue virus. We also emphasize the role of BMP in lipid sorting and degradation, especially cholesterol transport in cooperation with Niemann Pick type C proteins (NPC 1 and 2) and with some oxysterol-binding protein (OSBP)-related proteins (ORPs) as well as in sphingolipid degradation. Interestingly, numerous virus infection required NPC1 as well as ORPs along the endocytic pathway. Furthermore, BMP content is increased during pathological endosomal lipid accumulation in various lysosomal storage disorders. This is particularly important knowing the high percentage of patients with metabolic disorders among the SARS-CoV-2 infected patients presenting severe forms of COVID-19

Bis(monoacylglycero)phosphate reduces oxysterol formation and apoptosis in macrophages exposed to oxidized LDL.

International audienceAtherosclerosis is a major cardiovascular complication of diseases associated with increased oxidative stress that favors oxidation of circulating low density lipoproteins (LDLs). Oxidized LDL (oxLDL) is considered as highly atherogenic as it induces a strong accumulation of cholesterol in subendothelial macrophages leading to the formation of foam cells and emergence of atherosclerotic plaque. OxLDL is enriched in oxidation products of cholesterol called oxysterols, some of which have been involved in the ability of oxLDL to induce cellular oxidative stress and cytotoxicity, mainly by apoptosis. Little is known about the possible contribution of cell-generated oxysterols toward LDL-associated oxysterols in cellular accumulation of oxysterols and related apoptosis. Using both radiochemical and mass analyzes, we showed that oxLDL greatly enhanced oxysterol production by RAW macrophages in comparison with unloaded cells or cells loaded with native LDL. Most oxysterols were produced by non-enzymatic routes (7-ketocholesterol and 7α/β-hydroyxycholesterol) but enzymatically formed 7α-, 25- and 27-hydroxycholesterol were also quantified. Bis(monoacylglycero)phosphate (BMP) is a unique phospholipid preferentially found in late endosomes. We and others have highlighted the role of BMP in the regulation of intracellular cholesterol metabolism/traffic in macrophages. We here report that cellular BMP accumulation was associated with a significantly lower production of oxysterols upon oxLDL exposure. Of note, potent pro-apoptotic 7-ketocholesterol was the most markedly decreased. OxLDL-induced cell cytotoxicity and apoptosis were consistently attenuated in BMP-enriched cells. Taken together, our data suggest that BMP exerts a protective action against the pro-apoptotic effect of oxLDL via a reduced production of intracellular pro-apoptotic oxysterols

A critical appraisal of the measurement of serum 'cholesterol efflux capacity' and its use as surrogate marker of risk of cardiovascular disease

The 'cholesterol efflux capacity (CEC)' assay is a simple in vitro measure of the capacities of individual sera to promote the first step of the reverse cholesterol transport pathway, the delivery of cellular cholesterol to plasma HDL. This review describes the cell biology of this model and critically assesses its application as a marker of cardiovascular risk. We describe the pathways for cell cholesterol export, current cell models used in the CEC assay with their limitations and consider the contribution that measurement of serum CEC provides to our understanding of HDL function in vivo

Fatty Acid Composition and Metabolism in Leishmania Parasite Species: Potential Biomarkers or Drug Targets for Leishmaniasis?

International audienceFatty acids have received growing interest in Leishmania biology with the characterization of the enzymes allowing the complete fatty acid synthesis of this trypanosomatid parasite. This review presents a comparative analysis of the fatty acid profiles of the major classes of lipids and phospholipids in different species of Leishmania with cutaneous or visceral tropism. Specificities relating to the parasite forms, resistance to antileishmanial drugs, and host/parasite interactions are described as well as comparisons with other trypanosomatids. Emphasis is placed on polyunsaturated fatty acids and their metabolic and functional specificities, in particular, their conversion into oxygenated metabolites that are inflammatory mediators able to modulate metacyclogenesis and parasite infectivity. The impact of lipid status on the development of leishmaniasis and the potential of fatty acids as therapeutic targets or candidates for nutritional interventions are discussed

Increased ABCA1 (ATP-Binding Cassette Transporter A1)-Specific Cholesterol Efflux Capacity in Schizophrenia

International audienceObjective: Patients with schizophrenia have increased long-term mortality attributable to cardiovascular disease and commonly demonstrate features of mixed dyslipidemia with low HDL-C (high-density lipoprotein cholesterol). The removal of cholesterol from cells by HDL via specific ATP-binding cholesterol transporters is a major functional property of HDL, and its measurement as cholesterol efflux capacity (CEC) can predict cardiovascular risk. Whether HDL function is impaired in patients with schizophrenia is unknown. Approach and Results: We measured basal and ABCA1 (ATP-binding cassette transporter A1)- and ABCG1 (ATP-binding cassette transporter G1)-dependent CEC, comparing patients with schizophrenia with age- and sex-matched healthy controls, and related our findings to NMR analysis of lipoprotein subclasses. Total plasma cholesterol and LDL-C (low-density lipoprotein cholesterol) were comparable between healthy controls (n=51) and patients (n=120), but patients with schizophrenia had increased total plasma triglyceride, low HDL-C and apolipoprotein A-I concentrations. NMR analysis indicated a marked (15-fold) increase in large triglyceride-rich lipoprotein particle concentration, increased small dense LDL particles, and fewer large HDL particles. Despite lower HDL-C concentration, basal CEC was 13.7±1.6% higher, ABCA1-specific efflux was 35.9±1.6% higher, and ABCG1 efflux not different, in patients versus controls. In patients with schizophrenia, ABCA1-specific efflux correlated with the abundance of small 7.8 nm HDL particles but not with serum plasminogen or triglyceride levels.Conclusions: Patients with schizophrenia have increased concentrations of atherogenic apoB-containing lipoproteins, decreased concentrations of large HDL particles, but enhanced ABCA1-mediated CEC. In this population, preventative strategies should focus on reducing atherogenic lipoproteins rather than increasing CEC

THP1 macrophages oxidized cholesterol, generating 7-derivative oxysterols specifically released by HDL

International audienceMacrophages are well recognized as key pathophysiologic agents in many chronic inflammatory diseases, especially atherosclerosis. During atherogenesis process, low density lipoproteins (LDL) undergo oxidation (oxLDL) and become highly atherogenic as they induce a strong accumulation of cholesterol in subendothelial macrophages leading to the formation of foam cells, the major cellular component of fatty streaks. OxLDL are enriched in oxidation products of cholesterol called oxysterols involved in the regulation of cholesterol homeostasis, by their ability to induce cellular oxidative stress and cytotoxicity. Little is known about intracellular oxysterol production in macrophages. Using both radiochemical and mass analyzes, we showed that THP1 macrophages promote the intracellular oxidation of LDL derived-cholesterol as well as intracellular cholesterol, this later mechanism being enhanced by exposure with native or oxLDL. We demonstrated that in both THP1 and Raw 267.4 cells cholesterol oxidation occurs in the late endosomal compartment. Most oxysterols were produced by non-enzymatic routes (7-ketocholesterol and 7alpha/beta-hydroxycholesterol) but enzymatically formed 7alpha-, 27-hydroxycholesterol were also quantified. Incubation of THP1 macrophages with nLDL or oxLDL, induced a 2- and 100-fold increase in oxysterol production, respectively. Both oxysterols derived from LDL cholesterol and cellular cholesterol were readily exported to HDL whereas apoA1 was inefficient, showing that HDL plays a major role in the removal of excess oxysterols in THP1 macrophages

In vitro oxidized HDL and HDL from type 2 diabetes patients have reduced ability to efflux oxysterols from THP-1 macrophages

International audienceOxidized LDL (OxLDL) that are enriched in products of lipid peroxidation including oxysterols have been shown to induce cellular oxidative stress and cytotoxicity therefore accelerating atheroma plaque formation. Upon oxLDL exposure of THP-1 macrophages, intracellular oxidation of LDL derived-cholesterol as well as endogenous cholesterol was increased. The oxysterols intracellularly produced were efficiently exported to HDL whereas apolipoprotein A1 was inefficient. These findings prompted us to investigate the consequences of modification of HDL by oxidation and glycation as observed in type 2 diabetes with respect to oxysterol and cholesterol efflux. We show that efflux of oxysterols was significantly impaired after in vitro oxidation and glycoxidation of HDL whereas glycation alone had no impact. Cholesterol efflux was only slightly decreased by oxHDL or glycoxidized HDL and not changed with glycated HDL. The defect of HDL towards oxysterol efflux was also observed with HDL isolated from diabetic subjects as compared to healthy controls. These findings support a deleterious cellular retention of oxysterols due to dysfunctional HDL in type 2 diabetes

Increased Lipid Peroxidation in LDL from Type-2 Diabetic Patients.

International audienceIncreased oxidative stress is associated with type-2 diabetes and related cardiovascular diseases, but oxidative modification of LDL has been partially characterized. Our aim was to compare the lipid and fatty acid composition as well as the redox status of LDL from diabetic patients and healthy subjects. First, to ensure that isolation of LDL by sequential ultracentrifugation did not result in lipid modifications, lipid composition and peroxide content were determined in LDL isolated either by ultracentrifugation or fast-protein liquid chromatography. Both methods resulted in similar concentrations of lipids, fatty acids, hydroxy-octadecadienoic acid (HODE) and malondialdehyde (MDA). Then, LDLs were isolated by ultracentrifugation from eight type-2 diabetic patients and eight control subjects. Compared to control LDL, diabetic LDL contained decreased cholesteryl esters and increased triglyceride concentrations. Ethanolamine plasmalogens decreased by 49%. Proportions of linoleic acid decreased in all lipid classes, while proportions of arachidonic acid increased in cholesteryl esters. Total HODE concentrations increased by 56%, 12- and 15-hydroxy-eicosatetraenoic acid by 161 and 86%, respectively, and MDA levels increased by twofold. alpha-Tocopherol concentrations, expressed relative to triglycerides, were lower in LDL from patients compared to controls, while gamma-tocopherol did not differ. Overall, LDL from type-2 diabetic patients displayed increased oxidative stress. Determination of hydroxylated fatty acids and ethanolamine plasmalogen depletion could be especially relevant in diabetes

Enriched PUFA environment of Leishmania infantum promastigotes promotes the accumulation of lipid mediators and favors parasite infectivity towards J774 murine macrophages

Leishmania parasites are the causative agents of visceral or cutaneous leishmaniasis in humans and of canine leishmaniosis. The macrophage is the predilected host cell of Leishmania in which the promastigote stage is transformed into amastigote. We previously showed changes in the fatty acid composition (FA) of lipids in two strains of Leishmania donovani upon differentiation of promastigote to amastigote, including increased proportions of arachidonic acid (AA) and to a less extent of docosahexaenoic acid (DHA). Here, we carried out supplementation with AA or DHA on two Leishmania infantum strains, a visceral (MON-1) and a cutaneous (MON-24), to evaluate the role of these FA in parasite/macrophage interactions. The proportions of AA or DHA in total lipids were significantly increased in promastigotes cultured in AA- or DHA-supplemented media compared to controls. The content of FA-derived oxygenated metabolites was enhanced in supplemented strains, generating especially epoxyeicosatrienoic acids (11,12- and 14,15-EET) and hydroxyeicosatetraenoic acids (5- and 8- HETE) from AA, and hydroxydocosahexaenoic acids (14- and 17-HDoHE) from DHA. For both MON-1 and MON-24, AA-supplemented promastigotes showed higher infectivity towards J774 macrophages as evidenced by higher intracellular amastigote numbers. Higher infectivity was observed after DHA supplementation for MON-24 but not MON-1 strain. ROS production by macrophages increased upon parasite infection, but only minor change was observed between control and supplemented parasites. We propose that under high AA or DHA environment that is associated with AA or DHA enrichment of promastigote lipids, FA derivatives can accumulate in the parasite, thereby modulating parasite infectivity towards host macrophages