Membrane-exposed sulfatides are proposed to contribute to P-selectin-dependent platelet aggregation. Here, we demonstrated that P-selectin-mediated platelet aggregation on a collagen-coated surface... Show moreMembrane-exposed sulfatides are proposed to contribute to P-selectin-dependent platelet aggregation. Here, we demonstrated that P-selectin-mediated platelet aggregation on a collagen-coated surface under flow indeed depended on sulfatides and that this interaction differed considerably from the interaction of P-selectin with P-selectin Glycoprotein Ligand-1 (PSGL-1), which underlies leukocyte-endothelium adhesion.\n= 7-12 μM).\nOur data suggest that the sulfatide/P-selectin interaction implicates multiple binding pockets, which only partly overlap with that of PSGL-1. These observations open ways to selectively interfere with sulfatide/P-selectin-dependent platelet aggregation without affecting PSGL-1-dependent cell adhesion. Show less
OBJECTIVE The ABCA2 transporter shares high structural homology to ABCA1, which is crucial for the removal of excess cholesterol from macrophages and, by extension, in atherosclerosis. It has been... Show moreOBJECTIVE The ABCA2 transporter shares high structural homology to ABCA1, which is crucial for the removal of excess cholesterol from macrophages and, by extension, in atherosclerosis. It has been suggested that ABCA2 sequesters cholesterol inside the lysosomes, however, little is known of the macrophage-specific role of ABCA2 in regulating lipid homeostasis in vivo and in modulating susceptibility to atherosclerosis. METHODS Chimeras with dysfunctional macrophage ABCA2 were generated by transplantation of bone marrow from ABCA2 knockout (KO) mice into irradiated LDL receptor (LDLr) KO mice. RESULTS Interestingly, lack of ABCA2 in macrophages resulted in a diminished lesion size in the aortic root (-24.5%) and descending thoracic aorta (-36.6%) associated with a 3-fold increase in apoptotic cells, as measured by both caspase 3 and TUNEL. Upon oxidized LDL exposure, macrophages from wildtype (WT) transplanted animals developed filipin-positive droplets in lysosomal-like compartments, corresponding to free cholesterol (FC) accumulation. In contrast, ABCA2-deficient macrophages displayed an abnormal diffuse distribution of FC over peripheral regions. The accumulation of neutral sterols in lipid droplets was increased in ABCA2-deficient macrophages, but primarily in cytoplasmic clusters and not in lysosomes. Importantly, apoptosis of oxLDL loaded macrophages lacking ABCA2 was increased 2.7-fold, probably as a consequence of the broad cellular distribution of FC. CONCLUSIONS Lack of functional ABCA2 generates abnormalities in intracellular lipid distribution/trafficking in macrophages consistent with its lysosomal sequestering role, leading to an increased susceptibility to apoptosis in response to oxidized lipids and reduced atherosclerotic lesion development. Show less
Calpe-Berdiel, L.; Zhao, Y.; Graauw, M. de; Ye, D.; Santbrink, P.J. van; Mommaas, A.M.; ... ; Berkel, T.J.C. van 2012
ABCA7, a close relative of ABCA1 which facilitates cholesterol efflux to lipid-poor apoproteins, has been implicated in macrophage lipid efflux and clearance of apoptotic cells in in vitro studies... Show moreABCA7, a close relative of ABCA1 which facilitates cholesterol efflux to lipid-poor apoproteins, has been implicated in macrophage lipid efflux and clearance of apoptotic cells in in vitro studies. In the current study, we investigated the in vivo effects of macrophage ABCA7 deficiency on lipid metabolism and atherosclerosis. Chimeras with dysfunctional ABCA7 in macrophages and other blood cells were generated by transplantation of bone marrow from ABCA7 knockout (KO) mice into irradiated low-density lipoprotein receptor (LDLr) KO mice. Unexpectedly, macrophage ABCA7 deficiency did not significantly affect atherosclerosis susceptibility of LDLr KO mice after 10 weeks Western-type diet feeding. However, ABCA7 deficiency was associated with 2-fold (p<0.05) higher macrophage ABCA1 mRNA expression levels. Combined disruption of ABCA1 and ABCA7 in bone-marrow-derived cells increased atherosclerotic lesion development (1.5-fold (p>0.05) as compared to wild type transplanted mice. However, single deletion of ABCA1 had a similar effect (1.8-fold, p<0.05). Macrophage foam cell accumulation in the peritoneal cavity was reduced in ABCA1/ABCA7 dKO transplanted animals as compared to single ABCA1 KO transplanted mice, which was associated with increased ABCG1 expression. Interestingly, spleens of ABCA1/ABCA7 double KO transplanted mice were significantly larger as compared to the other 3 groups and showed massive macrophage lipid accumulation, a reduction in CD3+ T-cells, and increased expression of key regulators of erythropoiesis. In conclusion, deletion of ABCA7 in bone marrow-derived cells does not affect atherogenesis in the arterial wall neither in the absence or presence of ABCA1. Interestingly, combined deletion of bone marrow ABCA1 and ABCA7 causes severe splenomegaly associated with cellular lipid accumulation, a reduction in splenic CD3+ T cells, and induced markers of erythropoeisis. Our data indicate that ABCA7 may play a role in T cell proliferation and erythropoeisis in spleen. Show less
Meurs, I.; Lammers, B.; Zhao, Y.; Out, R.; Hildebrand, R.B.; Hoekstra, M.; ... ; Eck, M. van 2012
Aim ABCA1 protects against atherosclerosis by facilitating cholesterol efflux from macrophage foam cells in the arterial wall to extracellular apolipoprotein (apo) A-I. In contrast to apoA-I,... Show moreAim ABCA1 protects against atherosclerosis by facilitating cholesterol efflux from macrophage foam cells in the arterial wall to extracellular apolipoprotein (apo) A-I. In contrast to apoA-I, apoE is secreted by macrophages and can, like apoA-I, induce ABCA1-mediated cholesterol efflux. Yet, the combined effect of macrophage ABCA1 and apoE on lesion development is unexplored. Methods and ResultsLDL receptor knockout (KO) mice were transplanted with bone marrow from ABCA1/apoE double KO (dKO) mice, their respective single KO's, and wild-type (WT) controls and were challenged with a high-fat/high-cholesterol diet for 9 weeks. In vitro cholesterol efflux experiments showed no differences between ABCA1 KO and dKO macrophages. The serum non-HDL/HDL ratio in dKO transplanted mice was 1.7-fold and 2.4-fold (p<0.01) increased compared to WT and ABCA1 KO transplanted mice, respectively. The atherosclerotic lesion area in dKO transplanted animals (650±94×103 µm2), however, was 1.9-fold (p<0.01) and 1.6-fold (p<0.01) increased compared to single knockouts (ABCA1 KO: 341±20×103 µm2; apoE KO: 402±78×103 µm2, respectively) and 3.1-fold increased (p<0.001) compared to WT (211±20×103 µm2). When normalized for serum cholesterol exposure, macrophage ABCA1 and apoE independently protected against atherosclerotic lesion development (p<0.001). Moreover, hepatic expression levels of TNFα and IL-6 were highly induced in dKO transplanted animals (3.0-fold; p<0.05, and 4.3-fold; p<0.001, respectively). In agreement, serum IL-6 levels were also enhanced in ABCA1 KO transplanted mice (p<0.05) and even further enhanced in dKO transplanted animals (3.1-fold as compared to ABCA1 KO transplanted animals; p<0.05). Conclusions Combined deletion of macrophage ABCA1 and apoE results in a defect in cholesterol efflux and, compared to ABCA1 KO transplanted mice, elevated serum total cholesterol levels. Importantly, these mice also suffer from enhanced systemic and hepatic inflammation, together resulting in the observed augmented atherosclerotic lesion development. Show less
Excessive accumulation of cholesterol by macrophage-derived foam cells is one of the characteristic features of atherosclerotic lesion development. Macrophages not only play an important role in... Show moreExcessive accumulation of cholesterol by macrophage-derived foam cells is one of the characteristic features of atherosclerotic lesion development. Macrophages not only play an important role in the initiation of the early atherosclerotic lesion, during further progression of the lesions macrophages also contribute to the formation of a necrotic core, thereby affecting the stability of the atherosclerotic plaque. Especially in the initiation of atherosclerosis the balance between cholesterol influx and efflux in macrophages is of prime importance. This balance is maintained by scavenger receptors and ATP-binding cassette (ABC) transporters, which are key mediators for macrophage cholesterol homeostasis as they facilitate the influx and efflux of lipids. Macrophages are incapable of limiting the uptake of cholesterol by scavenger receptors, including scavenger receptor class A (SR-A) and CD36. Therefore, prevention of lipid accumulation in macrophages largely depends on cholesterol efflux pathways, mainly mediated by ABC transporters. Gaining more knowledge on macrophage lipid homeostasis is of prime importance for the development of new therapeutic strategies to prevent atherosclerotic lesion development or induce regression of existing lesions. The aim of the studies described in this thesis was investigation of the role of several ABC transporters and SR-BI in (macrophage) lipid metabolism and atherogenesis. Show less
Hildebrand, R.B.; Lammers, B.; Meurs, I.; Korporaal, S.J.A.; Haan, W. de; Zhao, Y.; ... ; Eck, M. van 2010
Objective-Disruption of scavenger receptor class B type I (SR-BI) in mice impairs high-density lipoprotein (HDL)-cholesterol (HDL-C) delivery to the liver and induces susceptibility to... Show moreObjective-Disruption of scavenger receptor class B type I (SR-BI) in mice impairs high-density lipoprotein (HDL)-cholesterol (HDL-C) delivery to the liver and induces susceptibility to atherosclerosis. In this study, it was investigated whether introduction of cholesteryl ester transfer protein (CETP) can normalize HDL-C transport to the liver and reduce atherosclerosis in SR-BI knockout (KO) mice. Methods and Results-Expression of human CETP in SR-BIKO mice resulted in decreased plasma HDL-C levels, both on chow diet (1.8-fold, P < 0.001) and on challenge with Western-type diet (1.6-fold, P < 0.01). Furthermore, the presence of CETP partially normalized the abnormally large HDL particles observed in SR-BIKO mice. Unexpectedly, expression of CETP in SR-BIKO mice did not reduce atherosclerotic lesion development, probably because of consequences of SR-BI deficiency, including the persistence of higher VLDL-cholesterol (VLDL-C) levels, unchanged elevated free cholesterol/total cholesterol ratio, and the increased oxidative status of the animals. In addition, CETP expression did not normalize other characteristics of SR-BI deficiency, including female infertility, reticulocytosis, thrombocytopenia, and impaired platelet aggregation. Conclusion-CETP restores HDL-C levels in SR-BIKO mice, but it does not change the susceptibility to atherosclerosis and other typical characteristics that are associated with SR-BI disruption. This may indicate that the pathophysiology of SR-BI deficiency is not a direct consequence of changes in the HDL pool. (Arterioscler Thromb Vasc Biol. 2010; 30: 1439-1445.) Show less
Hoekstra, M.; Meurs, I.; Koenders, M.; Out, R.; Hildebrand, R.B.; Kruijt, J.K.; ... ; Berkel, T.J.C. van 2008
Receptor-mediated cholesterol uptake has been suggested to play a role in maintaining the adrenal intracellular free cholesterol pool and the ability to produce hormones. Therefore, in the current... Show moreReceptor-mediated cholesterol uptake has been suggested to play a role in maintaining the adrenal intracellular free cholesterol pool and the ability to produce hormones. Therefore, in the current study, we evaluated the importance of scavenger receptor class B type I (SR-BI)-mediated cholesteryl ester uptake from HDL for adrenal glucocorticoid hormone synthesis in vivo. No difference was observed in the plasma level of corticosterone between SR-BI-deficient and wild-type mice under ad libitum feeding conditions. Overnight fasting (∼16 h) stimulated the plasma level of corticosterone by 2-fold in wild-type mice. In contrast, no effect of fasting on plasma corticosterone levels was observed in SR-BI-deficient mice, leading to a 44% lower plasma corticosterone level compared with their wild-type littermate controls. In parallel, an almost complete depletion of lipid stores in the adrenal cortex of fasted SR-BI-deficient mice was observed. Plasma adrenocorticotropic hormone levels were increased by 5-fold in fasted SR-BI-deficient mice. SR-BI deficiency induced marked changes in the hepatic expression of the glucocorticoid-responsive genes cholesterol 7α-hydroxylase, HMG-CoA synthase, apolipoprotein A-IV, corticosteroid binding globulin, interleukin-6, and tumor necrosis factor-α, which coincided with a 42% decreased plasma glucose level under fasting conditions. In conclusion, we show that the absence of adrenal HDL cholesteryl ester uptake in SR-BI-deficient mice impairs the adrenal glucocorticoid-mediated stress response to fasting as a result of adrenal glucocorticoid insufficiency and attenuated liver glucocorticoid receptor signaling, leading to hypoglycemia under fasting conditions. Show less
