Abstract 143: Macropinocytosis Mediates Enzymatically Modified LDL (ELDL)-Induced Murine Smooth Muscle Cell Formation: A Role for RAGE In ELDL Endocytosis and Upregulation of Scavenger Receptor LOX1

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Bijoy Chellan ◽  
Catherine A Reardon ◽  
Marion Hofmann Bowman
Keyword(s):  
Smooth Muscle ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Foam Cell Formation ◽  
Wild Type ◽  
Pi3k Inhibitors ◽  
Atherosclerotic Lesions ◽  
Ldl Particles

Background: ELDL is present in human atherosclerotic lesions and promotes foam cell formation in cultured macrophages and vascular smooth muscle cells (SMC). Here we study mechanism of ELDL uptake and its effects on SMCs. Methods and Results: Incubation of wild type murine aortic SMCs with 10 μg/ml ELDL (trypsin, cholesterol esterase modified) results in enhanced foam cell formation (analyzed by Oil Red O, lipid measurement) compared to SMCs incubated with acetylated LDL (500 μg/ml; -50%, p<0.01) and oxidized LDL (200 μg/ml; -75%, p<0.01). Inhibitors of macropinocytosis (50 μM LY294006, 2 μM wortmannin, and 3 mM amiloride) attenuated ELDL uptake (-50%, -50%, -100% respectively). In contrast, inhibitors of receptor mediated endocytosis (100 μM dynasore, 0.1 M Sucrose), and inhibitors of caveolae /lipid raft mediated endocytosis (5mM MBCD, 5 μM filipin) had no effect on ELDL uptake in SMCs. Moreover, ELDL incubation led to increased expression of scavenger receptor LOX1 (+ 3 fold, p<0.01) in wild type SMC’s, but not in SMC deficient in Receptor for AGE (RAGE-/-), while CD36 and SRA1 remained unchanged in both the SMCs. Importantly, RAGE-/- SMCs upon pretreatment with PI3K inhibitors that only partially inhibited macropinocytosis of ELDL in wild type SMCs, completely prevented ELDL uptake in RAGE-/- SMCs. Mechanistically, ELDL upregulates ROS (detected using H2DCFDA) and down regulates PIP3 (detected by pAkt immunoblotting) in wild type, but not in RAGE-/- SMCs. Since ROS is known to regulate macropinocytosis via increased Ca2+ levels, we tested Ca2+ channel inhibitor lacidipine (30 μM), and found complete inhibition of ELDL uptake in both, wild type and RAGE-/- SMCs. Lastly, we speculate that the fused structure of LDL in the ELDL preparation is preferentially activating RAGE, since oligomerization of ligands are known to increase RAGE signaling, and FPLC analysis demonstrated that ELDL consists mostly of fused LDL particles. Conclusions: ELDL is highly potent in inducing foam cells in aortic SMCs. ELDL endocytosis is mediated by RAGE-regulated, Ca2+ dependent macropinocytosis.

Abstract 633: Macrophage SR-BI Suppresses Atherosclerosis by Modulation of Autophagy via VPS34/Belclin-1 Pathway

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Huan Tao ◽  
Patricia G Yancey ◽  
John L Blakemore ◽  
Youmin Zhang ◽  
Lei Ding ◽  
...  
Keyword(s):  
Foam Cell ◽  
Oxidized Ldl ◽  
Cell Formation ◽  
Atherosclerotic Lesion ◽  
Beclin 1 ◽  
Foam Cell Formation ◽  
Aortic Tissue ◽  
Type I ◽  
Macrophage Foam Cell ◽  
Atherosclerotic Lesions

Background: Autophagy modulates vascular cell lipid metabolism, lipid droplet turnover, foam cell formation, cell survival and death, and inflammation. Scavenger receptor class B type I (SR-BI) deficiency causes impaired lysosome function in macrophages and erythrocytes. Methods and Results: Bone marrow transplantation studies were performed in ApoE and LDLR deficient mice to examine the effects of hematopoietic SR-BI deletion on atherosclerotic lesion autophagy. In addition, in vitro studies compared WT versus SR-BI -/- macrophages. Under conditions of cholesterol induced stress, the mRNA and protein levels of critical autophagy players including ATG5, ATG6/Belcin-1, ATG7 and LC3II were decreased by 37.8% to 84.6% (P<0.05 to 0.01) in SR-B1 -/- macrophages and atherosclerotic aortic tissue compared to controls. Electron microscopic analysis showed that SR-BI -/- versus WT macrophages had 80% fewer (P<0.05) autophagsomes in response to cholesterol enrichment. Macrophage SR-BI deficiency led to 1.8-fold (P<0.05) more lipid deposition and 2.5-fold more (P<0.01) apoptosis in response to oxidized LDL. Furthermore, hematopoietic SR-BI deletion caused 2.3 fold (P<0.05) more cell death in aortic atherosclerotic lesions compared to the WT control. Pharmacologic activation of autophagy did not reduce the levels of lipid droplets or cell apoptosis in SR-BI null macrophages vs WT control. WT peritoneal macrophages were used to examine SR-BI subcellular distribution and its interaction with VPS34/Beclin-1. In response to induction of autophagy, macrophage SR-BI was expressed in lysosomes and co-localized with LC3-II. Furthermore, we found that SR-BI directly interacted with the VPS34/Beclin-1 complex. Conclusions: SR-BI deficiency leads to defective autophagy and accelerates macrophage foam cell formation and apoptosis in experimental mouse atherosclerotic lesions. Macrophage SR-BI regulates expression of critical autophagy players and directly modulates autophagy via the VPS34/Beclin-1 pathway, identifying novel targets for the treatment of atherosclerosis.

Abstract 333: CD36 Mediates Cholesterol Uptake via Na/K-ATPase/lyn Signaling in Macrophages

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Yiliang Chen ◽  
David Kennedy ◽  
Zhichuan Li ◽  
Zijian Xie ◽  
Roy L Silverstein
Keyword(s):  
Kinase Activity ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Developed Countries ◽  
Foam Cell Formation ◽  
Macrophage Foam Cell ◽  
Cholesterol Uptake ◽  
Lyn Kinase

Atherosclerosis, the leading cause of death in the developed countries, is characterized by macrophage foam cell formation. We previously showed that CD36, a scavenger receptor highly expressed in macrophages, mediates oxidized-LDL uptake, contributes to intracellular cholesterol accumulation and foam cell formation, and regulates macrophage migration and pro-inflammatory signaling. Consistently, cd36 deletion in mice protects from diet-induced atherosclerosis. Mechanistically, we discovered a novel signaling pathway, in which oxidized LDL (oxLDL) binding to CD36 activates Lyn kinase and initiates a cascade that is necessary for the pro-atherogenic cellular phenotype. How CD36 regulates Lyn kinase remains undefined. Since we previously showed that the Na/K-ATPase (NKA) regulates Src family kinases, including Lyn, we hypothesized that CD36 regulates Lyn kinase via an interaction with NKA. We used co-immunoprecipitation, FRET, and a novel cross linking assay to demonstrate that CD36 physically associates with NKA on the macrophage surface. Using a Lyn kinase activity assay, we showed that the interaction regulates Lyn kinase activity in response to oxLDL in macrophages. Moreover, a newly developed peptide inhibitor specifically blocked Lyn activation in response to oxLDL and attenuated oxLDL-stimulated cholesterol uptake (135.6±3.4 μM cholesterol/mg protein after 24 hours vs 173.8±7.7 μM cholesterol/mg protein in vehicle treated cells; p=0.0005; n=6). Taken together, we conclude that CD36 signals through NKA to regulate Lyn kinase activity in macrophages, which may be a molecular mechanism underlying cholesterol overloading and foam cell formation.

Role of Ox-LDL and LOX-1 in Atherogenesis

2019 ◽  
Vol 26 (9) ◽  
pp. 1693-1700 ◽  
Author(s):  
Ajoe John Kattoor ◽  
Sri Harsha Kanuri ◽  
Jawahar L. Mehta
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Foam Cell ◽  
Leukocyte Adhesion ◽  
Oxidized Ldl ◽  
Cell Formation ◽  
Muscle Cells ◽  
Foam Cell Formation ◽  
Plaque Instability

Oxidized LDL (ox-LDL) plays a central role in atherosclerosis by acting on multiple cells such as endothelial cells, macrophages, platelets, fibroblasts and smooth muscle cells through LOX-1. LOX-1 is a 50 kDa transmembrane glycoprotein that serves as receptor for ox-LDL, modified lipoproteins, activated platelets and advance glycation end-products. Ox- LDL through LOX-1, in endothelial cells, causes increase in leukocyte adhesion molecules, activates pathways of apoptosis, increases reactive oxygen species and cause endothelial dysfunction. In vascular smooth muscle cells and fibroblasts, they stimulate proliferation, migration and collagen synthesis. LOX-1 expressed on macrophages inhibit macrophage migration and stimulate foam cell formation. They also stimulate generation of metalloproteinases and contribute to plaque instability and thrombosis. Drugs that modulate LOX-1 are desirable targets against atherosclerosis. Many naturally occurring compounds have been shown to modulate LOX-1 expression and atherosclerosis. Currently, novel drug design techniques are used to identify molecules that can bind to LOX-1 and inhibit its activation by ox-LDL. In addition, techniques using RNA interference and monoclonal antibody against LOX-1 are currently being investigated for clinical use.

OxLDL-mediated survival of macrophages does not require LDL internalization or signalling by major pattern recognition receptors

2011 ◽  
Vol 89 (4) ◽  
pp. 387-395 ◽  
Author(s):  
Maziar Riazy ◽  
Johnny H. Chen ◽  
Yasuhiko Yamamato ◽  
Hiroshi Yamamato ◽  
Vincent Duronio ◽  
...  
Keyword(s):  
Pattern Recognition ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Pi3k Pathway ◽  
Pattern Recognition Receptors ◽  
Foam Cell Formation ◽  
Macrophage Populations ◽  
Phosphoinositide 3 Kinase

Macrophages play a key role in the pathogenesis of atherosclerosis, in part by destabilizing plaques. We and others have shown that low concentrations of oxidized LDL (oxLDL) inhibit macrophage apoptosis. As oxLDL is present in lesions, this may be a mechanism by which macrophage populations in the intima are expanded. We have previously shown that oxLDL activates prosurvival signalling pathways such as the phosphoinositide 3-kinase (PI3K) pathway in bone marrow derived macrophages (BMDMs). However, little is known about more upstream signalling events especially at the receptor level. The endocytic pattern recognition receptors (PRRs), scavenger receptor A (SR-A) and CD36, are the main receptors on macrophages for uptake of oxLDL and are therefore important in foam cell formation. The signalling PRRs such as toll-like receptor (TLR) 2 and 4 also bind some types of oxLDL. This study was done to determine if any of the known PRRs are required for the anti-apoptotic effects of oxLDL in BMDMs. To do this, we tested the effect of oxLDL on viability of BMDMs lacking both SR-A and CD36 or lacking TLR2, TLR4, CD14, FcγRIIb, or RAGE. Our results indicate that none of these receptors are essential for activating the oxLDL prosurvival pathway. Furthermore, we show that the anti-apoptotic effect is not dependent on the uptake of oxLDL.

scholarly journals Chondroitin Sulfate N -acetylgalactosaminyltransferase-2 Impacts Foam Cell Formation and Atherosclerosis by Altering Macrophage Glycosaminoglycan Chain

2021 ◽  
Vol 41 (3) ◽  
pp. 1076-1091
Author(s):  
Imam Manggalya Adhikara ◽  
Keiko Yagi ◽  
Dyah Samti Mayasari ◽  
Yoko Suzuki ◽  
Koji Ikeda ◽  
...  
Keyword(s):  
Chondroitin Sulfate ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Density Lipoprotein ◽  
Peritoneal Macrophages ◽  
Metabolic Phenotype ◽  
Foam Cell Formation ◽  
Macrophage Foam Cell

Objective: Chondroitin sulfate proteoglycans are the primary constituents of the macrophage glycosaminoglycan and extracellular microenvironment. To examine their potential role in atherogenesis, we investigated the biological importance of one of the chondroitin sulfate glycosaminoglycan biosynthesis gene, ChGn-2 (chondroitin sulfate N -acetylgalactosaminyltransferase-2), in macrophage foam cell formation. Approach and Results: ChGn-2-deficient mice showed decreased and shortened glycosaminoglycans. ChGn-2 −/− /LDLr −/− (low-density lipoprotein receptor) mice generated less atherosclerotic plaque after being fed with Western diet despite exhibiting a metabolic phenotype similar to that of the ChGn-2 +/+ /LDLr −/− littermates. We demonstrated that in macrophages, ChGn-2 expression was upregulated in the presence of oxLDL (oxidized LDL), and glycosaminoglycan was substantially increased. Foam cell formation was significantly altered by ChGn-2 in both mouse peritoneal macrophages and the RAW264.7 macrophage cell line. Mechanistically, ChGn-2 enhanced oxLDL binding on the cell surface, and as a consequence, CD36—an important macrophage membrane scavenger receptor—was differentially regulated. Conclusions: ChGn-2 alteration on macrophages conceivably influences LDL accumulation and subsequently accelerates plaque formation. These results collectively suggest that ChGn-2 is a novel therapeutic target amenable to clinical translation in the future. Graphic Abstract: A graphic abstract is available for this article.

scholarly journals Blockade of Macrophage CD147 Protects Against Foam Cell Formation in Atherosclerosis

2021 ◽  
Vol 8 ◽  
Author(s):  
Jian-Jun Lv ◽  
Hao Wang ◽  
Hong-Yong Cui ◽  
Ze-Kun Liu ◽  
Ren-Yu Zhang ◽  
...  
Keyword(s):  
Foam Cell ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Foam Cell Formation ◽  
Macrophage Foam Cell ◽  
Atherosclerotic Lesions ◽  
Cd147 Expression ◽  
Compound Screening ◽  
Genetic Deletion ◽  
Atherosclerosis Development

The persistence of macrophage-derived foam cells in the artery wall fuels atherosclerosis development. However, the mechanism of foam cell formation regulation remains elusive. We are committed to determining the role that CD147 might play in macrophage foam cell formation during atherosclerosis. In this study, we found that CD147 expression was primarily increased in mouse and human atherosclerotic lesions that were rich in macrophages and could be upregulated by ox-LDL. High-throughput compound screening indicated that ox-LDL-induced CD147 upregulation in macrophages was achieved through PI3K/Akt/mTOR signaling. Genetic deletion of macrophage CD147 protected against foam cell formation by impeding cholesterol uptake, probably through the scavenger receptor CD36. The opposite effect was observed in primary macrophages isolated from macrophage-specific CD147-overexpressing mice. Moreover, bioinformatics results indicated that CD147 suppression might exert an atheroprotective effect via various processes, such as cholesterol biosynthetic and metabolic processes, LDL and plasma lipoprotein clearance, and decreased platelet aggregation and collagen degradation. Our findings identify CD147 as a potential target for prevention and treatment of atherosclerosis in the future.

Function of scavenger receptor class A type I/II is not important for smooth muscle foam cell formation

2008 ◽  
Vol 87 (2) ◽  
pp. 91-99 ◽  
Author(s):  
Birgit Luechtenborg ◽  
Oliver Hofnagel ◽  
Gabriele Weissen-Plenz ◽  
Nick J. Severs ◽  
Horst Robenek
Keyword(s):  
Smooth Muscle ◽  
Foam Cell ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Foam Cell Formation ◽  
Type I ◽  
Class A ◽  
Scavenger Receptor Class

scholarly journals 10: CHOLESTEROL FLUX PATHWAY ABNORMALITIES INDUCED BY PLASMA FROM PATIENTS WITH CHRONIC KIDNEY DISEASE

2016 ◽  
Vol 64 (3) ◽  
pp. 803.1-803
Author(s):  
N Miyawaki ◽  
F Daccueil ◽  
NM Siegart ◽  
J Mattana ◽  
I Voloshyna ◽  
...  
Keyword(s):  
Chronic Kidney Disease ◽  
Kidney Disease ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Pcr Analysis ◽  
Foam Cell Formation ◽  
Cvd Risk ◽  
Healthy Control

Purpose of StudyChronic kidney disease (CKD) is a known risk factor for cardiovascular disease (CVD). Patients with CKD have a high prevalence of atherosclerosis. However, CVD risk associated with CKD is not entirely explained by standard lipid profile or liver handling of cholesterol, as evidenced by the resistance to statin benefits seen in later stages of CKD. This study aims to detect changes in expression of cholesterol transport proteins in the setting of CKD and to determine if such changes adversely affect lipid handling by macrophages leading to cholesterol overload and atheromatous foam cell formation.Methods UsedTHP-1 human macrophages (106/ml) were incubated for 18 h–24 h with plasma obtained from 10 CKD patients (7 male, 3 female) or 10 healthy control subjects (4 male, 6 female). CKD patients were not on dialysis and had not received renal transplant. Following incubation, mRNA was isolated and reverse transcribed. The resulting cDNA was subjected to quantitative real-time PCR using specific primers for ATP binding cassette transporter (ABC)A1 (cholesterol efflux protein) and CD36, (a scavenger receptor with the capacity to endocytose oxidized LDL).Summary of ResultsPCR analysis showed that ABCA1 mRNA was reduced by 23±5% (p<0.0001) while CD36 mRNA was decreased by 36±7% (p<0.0001) in macrophages exposed to CKD plasma as compared to healthy control.ConclusionsThese findings suggest a different mechanism of lipid dysregulation associated with CKD that may explain the pathogenesis of elevated CVD risk in CKD and lack of response to statins. This mechanism, through pro-atherogenic suppression of ABCA1, differs from our finding in autoimmune rheumatic diseases where, in addition to lowering of ABCA1, augmentation of CD36 was also observed. In CKD, a paradoxical decrease in CD36 could compromise macrophage clearance of lipids, increasing vulnerability to lipoprotein thrombi in kidney. Further lowering of monocyte CD36 with statins would be of little benefit if CD36 is already low in CKD. Defining changes in lipid handling in CKD could lead to novel, targeted CVD treatment approaches in the CKD population.

Abstract 639: CD36 Recruits Na/K-ATPase/Lyn Complex to Mediate Proatherogenic Phenotypes in Macrophages

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Yiliang Chen ◽  
David Kennedy ◽  
Devi Prasadh Ramakrishnan ◽  
Huang Wenxin ◽  
Zhichuan Li ◽  
...  
Keyword(s):  
Foam Cell ◽  
Oxidized Ldl ◽  
Scavenger Receptor ◽  
Cell Formation ◽  
Foam Cell Formation ◽  
Null Mouse ◽  
Macrophage Migration ◽  
Lesion Area ◽  
Downstream Signaling ◽  
Lyn Kinase

Atherosclerosis is characterized by accumulation of macrophage foam cells in the arterial wall. We previously showed that CD36, a scavenger receptor highly expressed in macrophages, mediates oxidized-LDL (oxLDL) uptake, contributes to intracellular cholesterol accumulation and foam cell formation, and regulates macrophage migration and pro-inflammatory signaling. Genetic deletion of cd36 in mice is protective against diet-induced atherosclerosis. Mechanistically, we discovered that binding of oxLDL to CD36 activates Lyn kinase and initiates a cascade that is necessary for the pro-atherogenic cellular phenotype. Nevertheless, how CD36 regulates Lyn kinase remains undefined. We previously showed that Na/K-ATPase (NKA) regulates Src family kinases, including Lyn and we now hypothesized that CD36 regulates Lyn kinase via an interaction with NKA. We used co-immunoprecipitation and a novel immunofluorescence-based cell surface cross linking assay to demonstrate that CD36 physically associates with NKA on the macrophage surface. In a NKA α1 subunit heterozygous null mouse model in which ~60% of macrophage NKA expression is downregulated, we demonstrated that recruitment and activation of Lyn and its downstream signaling events in response to oxLDL were abolished. Functionally, we showed that NKA haploinsufficiency significantly blunted oxLDL uptake, foam cell formation and macrophage migration under atherogenic conditions. Importantly NKA α1 heterozygous null mice when bred into an apoe null background developed less atherosclerosis (26.7% lesion area in NKA control mice v.s. 13.4% lesion area in NKA heterozygous null mice) assessed by en face oil red O staining of aortae after 12 weeks on high fat diet. We conclude that by controlling Lyn kinase activity NKA critically regulates oxLDL/CD36 induced pro-atherogenic signaling.

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