Phospholipase A2 Mediates Apolipoprotein-Independent Uptake of Chylomicron Remnant-Like Particles by Human Macrophages

Apolipoprotein E-receptor-mediated pathways are the main routes by which macrophages take up chylomicron remnants, but uptake may also be mediated by receptor-independent routes. To investigate these mechanisms, triacylglycerol (TG) accumulation induced by apolipoprotein-free chylomicron remnant-like particles (CRLPw/o) in human monocyte-derived macrophages was evaluated. Macrophage TG content increased about 5-fold after incubation with CRLPw/o, and this effect was not reduced by the inhibition of phagocytosis, macropinocytosis, apolipoprotein E function, or proteoglycan bridging. The role of lipases, including lipoprotein lipase, cholesteryl ester hydrolase, and secretory (sPLA2) and cytosolic phospholipase A2, was studied using [3H]TG-labelled CRLPw/o. Total cell radioactivity after incubation with [3H]TG CRLPw/o was reduced by 15–30% by inhibitors of lipoprotein lipase and cholesteryl ester hydrolase and by about 45% by inhibitors of sPLA2 and cytosolic PLA2 . These results suggest that macrophage lipolytic enzymes mediate the internalization of postprandial TG-rich lipoproteins and that sPLA2 and cytosolic PLA2, play a more important role than extracellular lipoprotein lipase-mediated TG hydrolysis

Apatite - Cholesterol Agglomerates in Human Atherosclerotic Lesions

The purpose of this study was to examine the ultrastructural relationships of cholesterol crystals and apatite deposits in human atherosclerotic lesions. Segments of human aortic atherosclerotic lesions were obtained at autopsy , fixed in glutaraldehyde and dehydrated without using any organic solvents. The aortic segments were coated with carbon and subjected to various scanning electron microscope analyses. These included secondary electron imaging, back scattering of primary electrons, energy dispersive X-ray analysis of selected spots followed by area mapping of calcium and phosphorus , and cathodoluminescence. The information gathered from scanning of selected areas in the lesions by all the techniques showed that cholesterol crystals and apatite deposits are close to each other, within 10 μm distance or less. Cholesterol crystals are often surrounded by or adjacent to apatite. The results indicate that cholesterol and apatite crystals form closely linked agglomerates in human atherosclerotic lesions. Further studies are needed to determine whether precipitation of calcium and cholesterol are somehow linked during atherosclerotic lesion development

Evaluation of transduction efficiency in macrophage colony-stimulating factor differentiated human macrophages using HIV-1 based lentiviral vectors

<p>Abstract</p> <p>Background</p> <p>Monocyte-derived macrophages contribute to atherosclerotic plaque formation. Therefore, manipulating macrophage function could have significant therapeutic value. The objective of this study was to determine transduction efficiency of two HIV-based lentiviral vector configurations as delivery systems for the transduction of primary human blood monocyte-derived macrophages.</p> <p>Results</p> <p>Human blood monocytes were transduced using two VSV-G pseudotyped HIV-1 based lentiviral vectors containing EGFP expression driven by either native HIV-LTR (VRX494) or EF1α promoters (VRX1090). Lentiviral vectors were added to cultured macrophages at different times and multiplicities of infection (MOI). Transduction efficiency was assessed using fluorescence microscopy and flow cytometry. Macrophages transduced between 2 and 120 hours after culturing showed the highest transduction efficiency at 2-hours transduction time. Subsequently, cells were transduced 2 hours after culturing at various vector concentrations (MOIs of 5, 10, 25 and 50) to determine the amount of lentiviral vector particles required to maximally transduce human monocyte-derived macrophages. On day 7, all transduced cultures showed EGFP-positive cells by microscopy. Flow cytometric analysis showed with all MOIs a peak shift corresponding to the presence of EGFP-positive cells. For VRX494, transduction efficiency was maximal at an MOI of 25 to 50 and ranged between 58 and 67%. For VRX1090, transduction efficiency was maximal at an MOI of 10 and ranged between 80 and 90%. Thus, transductions performed with VRX1090 showed a higher number of EGFP-positive cells than VRX494.</p> <p>Conclusions</p> <p>This report shows that VSV-G pseudotyped HIV-based lentiviral vectors can efficiently transduce human blood monocyte-derived macrophages early during differentiation using low particle numbers that do not interfere with differentiation of monocytes into macrophages.</p

UBIAD1 Mutation Alters a Mitochondrial Prenyltransferase to Cause Schnyder Corneal Dystrophy

Peer reviewe

Jet size dependence of single jet suppression in lead-lead collisions at sqrt(s(NN)) = 2.76 TeV with the ATLAS detector at the LHC

Measurements of inclusive jet suppression in heavy ion collisions at the LHC provide direct sensitivity to the physics of jet quenching. In a sample of lead-lead collisions at sqrt(s) = 2.76 TeV corresponding to an integrated luminosity of approximately 7 inverse microbarns, ATLAS has measured jets with a calorimeter over the pseudorapidity interval |eta| < 2.1 and over the transverse momentum range 38 < pT < 210 GeV. Jets were reconstructed using the anti-kt algorithm with values for the distance parameter that determines the nominal jet radius of R = 0.2, 0.3, 0.4 and 0.5. The centrality dependence of the jet yield is characterized by the jet "central-to-peripheral ratio," Rcp. Jet production is found to be suppressed by approximately a factor of two in the 10% most central collisions relative to peripheral collisions. Rcp varies smoothly with centrality as characterized by the number of participating nucleons. The observed suppression is only weakly dependent on jet radius and transverse momentum. These results provide the first direct measurement of inclusive jet suppression in heavy ion collisions and complement previous measurements of dijet transverse energy imbalance at the LHC.Comment: 15 pages plus author list (30 pages total), 8 figures, 2 tables, submitted to Physics Letters B. All figures including auxiliary figures are available at http://atlas.web.cern.ch/Atlas/GROUPS/PHYSICS/PAPERS/HION-2011-02

UBIAD1 Mutation Alters a Mitochondrial Prenyltransferase to Cause Schnyder Corneal Dystrophy

Abstract Background: Mutations in a novel gene, UBIAD1, were recently found to cause the autosomal dominant eye disease Schnyder corneal dystrophy (SCD). SCD is characterized by an abnormal deposition of cholesterol and phospholipids in the cornea resulting in progressive corneal opacification and visual loss. We characterized lesions in the UBIAD1 gene in new SCD families and examined protein homology, localization, and structure

Fluorescent pegylated nanoparticles demonstrate fluid-phase pinocytosis by macrophages in mouse atherosclerotic lesions

The uptake of lipoproteins by macrophages is a critical step in the development of atherosclerotic lesions. Cultured monocyte-derived macrophages take up large amounts of native LDL by receptor-independent fluid-phase pinocytosis, either constitutively or in response to specific activating stimuli, depending on the macrophage phenotype. We therefore sought to determine whether fluid-phase pinocytosis occurs in vivo in macrophages in atherosclerotic lesions. We demonstrated that fluorescent pegylated nanoparticles similar in size to LDL (specifically nontargeted Qtracker quantum dot and AngioSPARK nanoparticles) can serve as models of LDL uptake by fluid-phase pinocytosis in cultured human monocyte-derived macrophages and mouse bone marrow-derived macrophages. Using fluorescence microscopy, we showed that atherosclerosis-prone Apoe-knockout mice injected with these nanoparticles displayed massive accumulation of the nanoparticles within CD68 + macrophages, including lipid-containing foam cells, in atherosclerotic lesions in the aortic arch. Similar results were obtained when atherosclerotic mouse aortas were cultured with nanoparticles in vitro. These results show that macrophages within atherosclerotic lesions can take up LDL-sized nanoparticles by fluid-phase pinocytosis and indicate that fluid-phase pinocytosis of LDL is a mechanism for macrophage foam cell formation in vivo

Murine bone marrow-derived macrophages differentiated with GM-CSF become foam cells by PI3K?-dependent fluid-phase pinocytosis of native LDL

Accumulation of cholesterol by macrophage uptake of LDL is a key event in the formation of atherosclerotic plaques. Previous research has shown that granulocyte-macrophage colony-stimulating factor (GM-CSF) is present in atherosclerotic plaques and promotes aortic lipid accumulation. However, it has not been determined whether murine GM-CSF-differentiated macrophages take up LDL to become foam cells. GM-CSF-differentiated macrophages from LDL receptor-null mice were incubated with LDL, resulting in massive macrophage cholesterol accumulation. Incubation of LDL receptor-null or wild-type macrophages with increasing concentrations of ¹²?I-LDL showed nonsaturable macrophage LDL uptake that was linearly related to the amount of LDL added, indicating that LDL uptake was mediated by fluid-phase pinocytosis. Previous studies suggest that phosphoinositide 3-kinases (PI3K) mediate macrophage fluid-phase pinocytosis, although the isoform mediating this process has not been determined. Because PI3K? is known to promote aortic lipid accumulation, we investigated its role in mediating macrophage fluid-phase pinocytosis of LDL. Wild-type macrophages incubated with LDL and the PI3K? inhibitor AS605240 or PI3K?-null macrophages incubated with LDL showed an ?50% reduction in LDL uptake and cholesterol accumulation compared with wild-type macrophages incubated with LDL only. These results show that GM-CSF-differentiated murine macrophages become foam cells by fluid-phase pinocytosis of LDL and identify PI3K? as contributing to this process