Macrophage-specific Abca1 knock-out (Abca1؊M/؊M ) mice were generated to determine the role of macrophage ABCA1 expression in plasma lipoprotein concentrations and the innate immune response of macrophages. Plasma lipid and lipoprotein concentrations in chow-fed Abca1 ؊M/؊M and wild-type (WT) mice were indistinguishable. Compared with WT macrophages, Abca1 ؊M/؊M macrophages had a >95% reduction in ABCA1 protein, failed to efflux lipid to apoA-I, and had a significant increase in free cholesterol (FC) and membrane lipid rafts without induction of endoplasmic reticulum stress. Lipopolysaccharide (LPS)-treated Abca1 ABCA1 (ATP-binding cassette transporter A1) is a plasma membrane protein that is widely expressed throughout the body (1, 2) and functions as a primary gatekeeper for eliminating excess free cholesterol (FC) 2 from tissues by effluxing cellular FC and phospholipid (PL) to lipid-free apoA-I, resulting in the formation of nascent high density lipoprotein (HDL) particles (3, 4). The nascent discoid-shaped HDL then undergoes a maturation process that involves additional lipid acquisition and conversion of FC to cholesteryl ester (CE) by lecithin:cholesterol acyltransferase to become mature spherical plasma HDL. Mutations that inactivate the human ABCA1 gene result in Tangier disease, which is characterized by extremely low HDL cholesterol concentrations, mildly elevated plasma trigelyceride levels, and accumulation of cholesterol in macrophages (5-10). Targeted deletion of Abca1 in mice and a natural mutation of Abca1 in the Wisconsin hypoalpha mutant chicken recapitulate the Tangier plasma lipid phenotype, supporting the essential role of ABCA1 in HDL formation (11-15). Although ABCA1 is expressed in many cells in the body, recent studies in hepatocyte-and intestinal epithelium-specific Abca1 knock-out mice suggest that the liver contributes 70 -80% of the plasma HDL pool, whereas the intestine contributes 20 -30% (16, 17). Although mobilization of excess FC from macrophages is dependent on ABCA1 and results in the formation of nascent HDL particles, transplantation of bone marrow from Abca1 knock-out (KO) mice into wild-type (WT) mice or transplantation of WT marrow into Abca1 KO recipients has little effect on plasma HDL concentrations, suggesting that macrophage ABCA1 expression has minimal impact on plasma HDL concentrations (18,19).Macrophages are a primary cell type involved in innate immunity. Although macrophage ABCA1 has a minimal impact on plasma lipid levels, there is evidence that its activity modulates the inflammatory response of macrophages to pathogen-associated molecules such as lipopolysaccharide

ATP-binding cassette transporter A1 (ABCA1) is a plasma membrane protein that functions to eliminate excess free cholesterol (FC) from tissues by effl uxing cellular FC and phospholipid (PL) to lipid-free apolipoprotein AI, forming nascent HDL particles ( 1, 2 ). Therefore, ABCA1 plays a critical role in the movement of cholesterol from peripheral tissues to the liver in a process known as reverse cholesterol transport. Mutations that inactivate the human ABCA1 gene result in Tangier disease, which is characterized by extremely low plasma HDL cholesterol concentrations, mildly elevated plasma triglyceride levels, and accumulation of cholesterol in macrophages ( 3-5 ). ABCA1 protein is expressed to a variable extent in most cells in the body, and its expression is regulated by transcriptional activation and protein degradation ( 6, 7 ), making it diffi cult to determine HL-49373, HL-094525, and AT-27820 (J.S.P.) Ϫ / Ϫ , total Abca1 knockout; ABCG1, ATP-binding cassette transporter G1; BMDM, bone marrow-derived macrophage; CT-B, cholera toxin B; FC, free cholesterol; fPEG-chol, fl uorescein ester of polyethylene glycol-derivatized Macrophage ABCA1 reduces MyD88-dependent Toll-like This work was supported by National Institutes of Health Grants

Rationale Plasma HDL levels are inversely correlated with atherosclerosis. Although it is widely assumed that this is due to the ability of HDL to promote cholesterol efflux from macrophage foam cells, direct experimental support for this hypothesis is lacking. Objective To assess the role of macrophage cholesterol efflux pathways in atherogenesis. Methods and Results We developed MAC-ABCDKO mice with efficient deletion of the ATP Binding Cassette Transporters A1 and G1 (ABCA1 and ABCG1) in macrophages but not in hematopoietic stem or progenitor populations. MAC-ABCDKO bone marrow (BM) was transplanted into Ldlr-/- recipients. On the chow diet, these mice had similar plasma cholesterol and blood monocyte levels but increased atherosclerosis compared to controls. On the Western type diet (WTD), MAC-ABCDKO BM transplanted Ldlr-/- mice had disproportionate atherosclerosis, considering they also had lower VLDL/LDL cholesterol levels than controls. ABCA1/G1 deficient macrophages in lesions showed increased inflammatory gene expression. Unexpectedly, WTD-fed MAC-ABCDKO BM transplanted Ldlr-/- mice displayed monocytosis and neutrophilia in the absence of HSPC proliferation. Mechanistic studies revealed increased expression of M-CSF and G-CSF in splenic macrophage foam cells, driving BM monocyte and neutrophil production. Conclusion These studies 1) show that macrophage deficiency of ABCA1/G1 is pro-atherogenic likely by promoting plaque inflammation and 2) uncover a novel positive feedback loop in which cholesterol-laden splenic macrophages signal BM progenitors to produce monocytes, with suppression by macrophage cholesterol efflux pathways.

The liver X receptors (LXRs) are transcriptional regulators of lipid homeostasis that also have potent anti-inflammatory effects. The molecular basis for their anti-inflammatory effects is incompletely understood, but has been proposed to involve the indirect tethering of LXRs to inflammatory gene promoters. Here we demonstrate that the ability of LXRs to repress inflammatory gene expression in cells and mice derives primarily from their ability to regulate lipid metabolism through transcriptional activation and can occur in the absence of SUMOylation. Moreover, we identify the putative lipid transporter Abca1 as a critical mediator of LXR's anti-inflammatory effects. Activation of LXR inhibits signaling from TLRs 2, 4 and 9 to their downstream NF-κB and MAPK effectors through Abca1-dependent changes in membrane lipid organization that disrupt the recruitment of MyD88 and TRAF6. These data suggest that a common mechanism-direct transcriptional activation-underlies the dual biological functions of LXRs in metabolism and inflammation.DOI: http://dx.doi.org/10.7554/eLife.08009.001

Background-Stearoyl-coenzyme A desaturase 1 (SCD1) is a well-known enhancer of the metabolic syndrome. The purpose of the present study was to investigate the role of SCD1 in lipoprotein metabolism and atherosclerosis progression. Methods and Results-Antisense oligonucleotides were used to inhibit SCD1 in a mouse model of hyperlipidemia and atherosclerosis (LDLr Ϫ/Ϫ Apob 100/100 ). In agreement with previous reports, inhibition of SCD1 protected against diet-induced obesity, insulin resistance, and hepatic steatosis. Unexpectedly, however, SCD1 inhibition strongly promoted aortic atherosclerosis, which could not be reversed by dietary oleate. Further analyses revealed that SCD1 inhibition promoted accumulation of saturated fatty acids in plasma and tissues and reduced plasma triglyceride, yet had little impact on low-density lipoprotein cholesterol. Because dietary saturated fatty acids have been shown to promote inflammation through toll-like receptor 4, we examined macrophage toll-like receptor 4 function. Interestingly, SCD1 inhibition resulted in alterations in macrophage membrane lipid composition and marked hypersensitivity to toll-like receptor 4 agonists. Conclusions-This study demonstrates that atherosclerosis can occur independently of obesity and insulin resistance and argues against SCD1 inhibition as a safe therapeutic target for the metabolic syndrome.

In humans, the plasma concentration of HDLs has been repeatedly shown to be inversely correlated with the risk of developing coronary heart disease ( 1, 2 ). The concentrations of plasma HDL has been shown to be largely dependent on hepatic ATP-binding cassette transporter A1 (ABCA1) ( 3-5 ), which transports and promotes the effl ux of glycerophosphocholine (PC), free cholesterol (FC), and sphingomyelin (SM) to wild-type apolipoprotein A-I (apoA-I), resulting in the formation of nascent HDLs (nHDLs). Functional mutations in human ABCA1 cause Tangier disease ( 6, 7 ), which is characterized by very low levels of plasma HDL apoA-I. Tangier disease is believed to alter a process termed "reverse cholesterol transport." Although defects in ABCA1 function have been identifi ed by Abstract This report details the lipid composition of nascent HDL (nHDL) particles formed by the action of the ATP binding cassette transporter A1 (ABCA1) on apolipoprotein A-I (apoA-I). nHDL particles of different size (average diameters of ‫ف‬ 12, 10, 7.5, and <6 nm) and composition were purifi ed by size-exclusion chromatography. Electron microscopy suggested that the nHDL were mostly spheroidal. The proportions of the principal nHDL lipids, free cholesterol, glycerophosphocholine, and sphingomyelin were similar to that of lipid rafts, suggesting that the lipid originated from a raft-like region of the cell. Smaller amounts of glucosylceramides, cholesteryl esters, and other glycerophospholipid classes were also present. The largest particles, ‫ف‬ 12 nm and 10 nm diameter, contained ‫ف‬ 43% free cholesterol, 2-3% cholesteryl ester, and three apoA-I molecules. Using chemical cross-linking chemistry combined with mass spectrometry, we found that three molecules of apoA-I in the ‫ف‬ 9-14 nm nHDL adopted a belt-like conformation. The smaller (7.5 nm diameter) spheroidal nHDL particles carried 30% free cholesterol and two molecules of apoA-I in a twisted, antiparallel, double-belt conformation. Overall, these new data offer fresh insights into the biogenesis and structural constraints involved in forming nascent HDL from ABCA1 . -Sorci-Thomas, M. G., J. S. Owen, B. Fulp, S. Bhat, These studies were supported by grants from the National Institutes of Health grants and Abbreviations: BMDM, bone marrow-derived macrophage; CE, cholesteryl ester; DSP, dithiobis(succinimidylpropionate) ; EM, electron microscopy; FC, free cholesterol; FPLC, fast protein liquid chromatography; GP, glycerophospholipids; HEK, human embryonic kidney; HexCer; hexosylceramides; LPC, lyso-glycerophosphocholine; MSCE, mass spectrometer collision energy; NDGGE, nondenaturing gradient gel electrophoresis; nHDL, nascent HDL; PC, glycerophosphocholine; PE, glycerophospho ethanolamine; PG, glycerophosphoglycerol; PI, glyerophos phoinositol; PM, plasma membrane; POPC, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine; PS, glycerophosphoserine; rHDL, recombinant HDL; RT, room temperature; SL, sphingolipid; TC, total cholesterol.1 To whom correspondence should be addressed. e-mail...

The collapse of thymic stromal cell microenvironment with age and resultant inability of the thymus to produce naïve T cells contributes to lower immune-surveillance in the elderly. Here we show that age-related increase in ‘lipotoxic danger signals’ such as free cholesterol (FC) and ceramides, leads to thymic caspase-1 activation via the Nlrp3 inflammasome. Elimination of Nlrp3 and Asc, a critical adaptor required for inflammasome assembly, reduces age-related thymic atrophy and results in an increase in cortical thymic epithelial cells, T cell progenitors and maintenance of T cell repertoire diversity. Using a mouse model of irradiation and hematopoietic stem cell transplantation (HSCT), we show that deletion of the Nlrp3 inflammasome accelerates T cell reconstitution and immune recovery in middle-aged animals. Collectively, these data demonstrate that lowering inflammasome-dependent caspase-1 activation increases thymic lymphopoiesis and suggest that Nlrp3 inflammasome inhibitors may aid the reestablishment of a diverse T cell repertoire in middle-aged or elderly patients undergoing HSCT.

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