Endothelial NOTCH1 is suppressed by circulating lipids and antagonizes inflammation during atherosclerosis
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE72633 | GEO | 2015/10/26
SECONDARY ACCESSION(S): PRJNA294525
REPOSITORIES: GEO
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Project description:While much progress has been made in identifying the mechanisms that trigger endothelial activation and inflammatory cell recruitment during atherosclerosis, less is known about the intrinsic pathways that counteract these events. Here we identified NOTCH1 as an antagonist of endothelial cell activation. NOTCH1 was constitutively expressed by adult arterial endothelium, but levels were significantly reduced by high fat diet. Furthermore, treatment of human aortic endothelial cells (HAEC) with inflammatory lipids (Ox-PAPC) and pro-inflammatory cytokines (TNFalpha and IL1beta) decreased Notch1 expression and signaling in vitro through a mechanism that requires STAT3 activation. Reduction of NOTCH1 in HAEC by siRNA, in the absence of inflammatory lipids or cytokines, increased inflammatory molecules and binding of monocytes. Conversely, some of the effects mediated by Ox-PAPC were reversed by increased NOTCH1 signaling; suggesting a link between lipid-mediated inflammation and Notch1. Interestingly, reduction of NOTCH1 by Ox-PAPC in HAEC was associated with a genetic variant previously correlated to HDL in a human GWAS. Finally endothelial Notch1 heterozygous mice showed higher diet-induced atherosclerosis. Based on these findings, we propose that reduction of endothelial NOTCH1 is a predisposing factor in the onset of vascular inflammation and initiation of atherosclerosis. Transcript profile from Human Aortic Endothelial Cells (HAEC) transfected with siRNA targeting NOTCH1 (n=3) or treated with Ox-PAPC (Oxidized 1-Palmitoyl-2-Arachidonoyl-sn-glycero-3-PhosphoCholine) for 6 hours (n=3) were compared to control HAEC (transfected with control siRNA and control media; n=3).
2015-10-26 | E-GEOD-72633 | biostudies-arrayexpress
Project description:Analysis of the effect of a bacterial quorum sensing molecule on in-vitro culture of human endothelial cells at gene expression levels. Objective: Chronic infection has long been postulated as a stimulus for atherogenesis. Pseudomonas aeruginosa infection has been associated with increased atherosclerosis in rats, and the bacteria produce a quorum sensing molecule 3-oxo-dodecynoyl-homoserine lactone (3OC12-HSL) that is critical for colonization and virulence. Paraoxonase 2 (PON2) hydrolyzes 3OC12-HSL and also protects against the effects of oxidized phospholipids thought to contribute to atherosclerosis. We now report the response of human aortic endothelial cells (HAEC) to 3OC12-HSL and oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (Ox-PAPC) in relation to PON2 expression. Methods and Results: Using expression profiling and network modeling, we identified the unfolded protein response (UPR), cell cycle genes, and the MAPK signaling pathway to be heavily involved in the HAEC response to 3OC12-HSL. The network also showed striking similarities to a network created based on HAEC response to Ox-PAPC, a major component of minimally-modified LDL. HAEC in which PON2 was silenced by siRNA showed increased pro-inflammatory and UPR responses when treated with 3OC12-HSL or Ox-PAPC. Conclusion: 3OC12-HSL and Ox-PAPC influence similar inflammatory pathways. Quorum sensing molecules such as 3OC12-HSL contribute to the pro-atherogenic effects of chronic infection and the anti-atherogenic effects of PON2 include destruction of quorum sensing molecules. 4 HAEC lines from different donors were treated with 3-O-C12 HSL or control (medium).
2010-12-30 | E-GEOD-26295 | biostudies-arrayexpress
Project description:Analysis of the effect of a bacterial quorum sensing molecule on in-vitro culture of human endothelial cells at gene expression levels. Objective: Chronic infection has long been postulated as a stimulus for atherogenesis. Pseudomonas aeruginosa infection has been associated with increased atherosclerosis in rats, and the bacteria produce a quorum sensing molecule 3-oxo-dodecynoyl-homoserine lactone (3OC12-HSL) that is critical for colonization and virulence. Paraoxonase 2 (PON2) hydrolyzes 3OC12-HSL and also protects against the effects of oxidized phospholipids thought to contribute to atherosclerosis. We now report the response of human aortic endothelial cells (HAEC) to 3OC12-HSL and oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (Ox-PAPC) in relation to PON2 expression. Methods and Results: Using expression profiling and network modeling, we identified the unfolded protein response (UPR), cell cycle genes, and the MAPK signaling pathway to be heavily involved in the HAEC response to 3OC12-HSL. The network also showed striking similarities to a network created based on HAEC response to Ox-PAPC, a major component of minimally-modified LDL. HAEC in which PON2 was silenced by siRNA showed increased pro-inflammatory and UPR responses when treated with 3OC12-HSL or Ox-PAPC. Conclusion: 3OC12-HSL and Ox-PAPC influence similar inflammatory pathways. Quorum sensing molecules such as 3OC12-HSL contribute to the pro-atherogenic effects of chronic infection and the anti-atherogenic effects of PON2 include destruction of quorum sensing molecules.
2010-12-30 | GSE26295 | GEO
Project description:Exposure to air particulate matter is associated with increased cardiovascular morbidity and mortality. One of the possible mechanisms is by promotion of atherosclerosis mediated by synergy of particulate matter chemicals and oxidized phospholipids on vascular endothelial cells. We treated human microvascular endothelial cells (HMEC) in triplicate wells with diesel exhaust particles (DEP) extract at a concentration of 5 and 25 mcg/ml, oxidized 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphorylcholine (ox-PAPC) at concentrations of 10, 20 and 40 mcg/ml or combination of DEP at 5 mcg/ml plus ox-PAPC at 10, 20 and 40 mcg/ml. The 3600 most variably expressed genes were employed to construct a weighted gene co-expression network to assess the degree of connectivity. Module enrichment analysis showed that three modules (Brown, Green and Yellow) were highly significant (p< 0.0001) in enriched genes regulated by the treatments. These three modules exhibited patterns of synergistic/additive interaction and were enriched in pathway genes relevant to vascular inflammation. The in-vivo relevance of this gene clustering analysis was established by demonstrating that liver gene expression of hypercholesterolemic mice exposed to ultrafine particles exhibited significant upregulation of similar gene pathways, as predicted by the in-vitro synergy. Keywords: Expression profiling comparison, effect of DEP vs. ox-PAPC vs. DEP plus ox-PAPC
2007-07-30 | GSE6584 | GEO
Project description:The transition of the endothelium to a pro-inflammatory state is key to progression of chronic inflammatory diseases including rheumatoid arthritis, chronic bowel disease and atherosclerosis. In atherosclerosis it is hypothesized that low density lipoproteins (LDL) that become trapped in the intima of the blood vessels are oxidized to minimally modified LDL (mmLDL) and that these serve as an important contributing factors to endothelial dysfunction. Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphatidylcholine (OX-PAPC), a model of the active phospholipid components of mmLDL affects the expression of hundreds of genes involved in inflammatory and other biological processes in human aortic endothelial cells (HAECs). We hypothesized that microRNAs (miRNAs) partially regulate this response. Using next generation sequencing, we identified miR-21-3p and miR-27a-5p to be induced 4-fold and 3-fold, respectively in response to OX-PAPC treatment compared to control treatment in HAECs. To identify the targets, we performed whole genome transcript profiling following transient over-expression of these two miRNAs followed by. In total, 1254 genes were down-regulated with 925 of them overlapping between the two miRNAs. Functional enrichment analysis using Gene Ontology predicted that the two miRNAs were involved in the regulation of NF-κB signaling. We characterized the Toll/interleukin-1 receptor (TIR) domain-containing adaptor protein TICAM2 as a direct target of miR-21-3p and miR-27a-5p. Furthermore, we showed that over-expression of miR-21-3p and miR-27a-5p lead to decreased p65 translocation to the nucleus and decreased the expression of known NF-κB downstream target genes confirming both miRNAs’ role in negatively regulating NF-κB signaling in endothelial cells. mRNA expression profiling of human aortic endothelial cells from two separate donors that were transfected with 1 nM microRNA mimics and negative control. The miRIDIAN mimics used were miR-21-3p (Catalog Number:C-301023-01-0005), miR-27a-5p (Catalog No: C-301028-01-0005), negative control (Catalog No: CN-001000-01-05)
2014-06-01 | E-GEOD-48006 | biostudies-arrayexpress
Project description:The transition of the endothelium to a pro-inflammatory state is key to progression of chronic inflammatory diseases including rheumatoid arthritis, chronic bowel disease and atherosclerosis. In atherosclerosis it is hypothesized that low density lipoproteins (LDL) that become trapped in the intima of the blood vessels are oxidized to minimally modified LDL (mmLDL) and that these serve as an important contributing factors to endothelial dysfunction. Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphatidylcholine (OX-PAPC), a model of the active phospholipid components of mmLDL affects the expression of hundreds of genes involved in inflammatory and other biological processes in human aortic endothelial cells (HAECs). We hypothesized that microRNAs (miRNAs) partially regulate this response. Using next generation sequencing, we identified miR-21-3p and miR-27a-5p to be induced 4-fold and 3-fold, respectively in response to OX-PAPC treatment compared to control treatment in HAECs. To identify the targets, we performed whole genome transcript profiling following transient over-expression of these two miRNAs followed by. In total, 1254 genes were down-regulated with 925 of them overlapping between the two miRNAs. Functional enrichment analysis using Gene Ontology predicted that the two miRNAs were involved in the regulation of NF-κB signaling. We characterized the Toll/interleukin-1 receptor (TIR) domain-containing adaptor protein TICAM2 as a direct target of miR-21-3p and miR-27a-5p. Furthermore, we showed that over-expression of miR-21-3p and miR-27a-5p lead to decreased p65 translocation to the nucleus and decreased the expression of known NF-κB downstream target genes confirming both miRNAs’ role in negatively regulating NF-κB signaling in endothelial cells.
2014-06-01 | GSE48006 | GEO
Project description:Oxidation products of 1-palmitoyl-2-arachidonoyl-sn-glycerol-2-phosphatidylcholine (PAPC), referred to as OxPAPC, accumulate in atherosclerotic lesions and regulate over 1000 genes in human aortic endothelial cells (HAEC). We previously demonstrated that OxPAPC covalently binds to a number of proteins in HAEC. The goal of these studies was to gain insight into the binding mechanism and determine if binding regulates activity. In whole cells N-acetylcysteine inhibited gene regulation by OxPAPC, and blocking cell cysteines with N-ethylmaleimide strongly inhibited OxPAPC binding. Using MS, we demonstrate that most of the binding of OxPAPC to cysteine is mediated by PEIPC. We also show that OxPAPC binds to a model protein, H-Ras, at cysteines previously shown to regulate activity in response to 15dPGJ2. This binding was observed with recombinant protein and in cells overexpressing H-Ras. 15dPGJ2 and OxPAPC increased H-Ras activity at comparable concentrations. Using microarray analysis, we demonstrate a considerable overlap of gene regulation by OxPAPC and 15dPGJ2 in HAEC, suggesting that some effects attributed to 15dPGJ2 may also be regulated by OxPAPC since OxPAPC lipids and 15dPGJ2 both accumulate in inflammatory sites. Overall, we provide evidence for the importance of OxPAPC-cysteine interactions in regulating HAEC function. Control, OxPAPC, and 15dPGJ2-treated HAECs Samples. 'Control' is media (1%FBS, 99%M199)-only treated HAECs. 15dPGJ2 and OxPAPC are treated with those respective compounds in media.
2012-02-14 | E-GEOD-35709 | biostudies-arrayexpress
Project description:Oxidation products of 1-palmitoyl-2-arachidonoyl-sn-glycerol-2-phosphatidylcholine (PAPC), referred to as OxPAPC, accumulate in atherosclerotic lesions and regulate over 1000 genes in human aortic endothelial cells (HAEC). We previously demonstrated that OxPAPC covalently binds to a number of proteins in HAEC. The goal of these studies was to gain insight into the binding mechanism and determine if binding regulates activity. In whole cells N-acetylcysteine inhibited gene regulation by OxPAPC, and blocking cell cysteines with N-ethylmaleimide strongly inhibited OxPAPC binding. Using MS, we demonstrate that most of the binding of OxPAPC to cysteine is mediated by PEIPC. We also show that OxPAPC binds to a model protein, H-Ras, at cysteines previously shown to regulate activity in response to 15dPGJ2. This binding was observed with recombinant protein and in cells overexpressing H-Ras. 15dPGJ2 and OxPAPC increased H-Ras activity at comparable concentrations. Using microarray analysis, we demonstrate a considerable overlap of gene regulation by OxPAPC and 15dPGJ2 in HAEC, suggesting that some effects attributed to 15dPGJ2 may also be regulated by OxPAPC since OxPAPC lipids and 15dPGJ2 both accumulate in inflammatory sites. Overall, we provide evidence for the importance of OxPAPC-cysteine interactions in regulating HAEC function.
2012-02-14 | GSE35709 | GEO
Project description:Oxidized phospholipids are thought to promote atherogenesis by stimulating endothelial cells (ECs) to produce inflammatory cytokines, such as IL-8. In studies with mouse models, we previously demonstrated that genetic variation in inflammatory responses of endothelial cells to oxidized lipids contributes importantly to atherosclerosis susceptibility. We now show that similar variations occur in cultured aortic ECs derived from multiple heart transplant donors. These variations were stably maintained between passages and, thus, reflect either genetic or epigenetic regulatory differences. Expression array analysis of aortic EC cultures derived from 12 individuals revealed that >1,000 genes were regulated by oxidized phospholipids. We have used the observed variations in the sampled population to construct a gene coexpression network comprised of 15 modules of highly connected genes. We show that several identified modules are significantly enriched in genes for known pathways and confirm a module enriched for unfolded protein response (UPR) genes using siRNA and the UPR inducer tunicamycin. On the basis of the constructed network, we predicted that a gene of unknown function (MGC4504) present in the UPR module is a target for UPR transcriptional activator ATF4. Our data also indicate that IL-8 is present in the UPR module and is regulated, in part, by the UPR. We validate these by using siRNA. In conclusion, we show that interindividual variability can be used to group genes into pathways and predict gene-gene regulatory relationships, thus identifying targets potentially involved in susceptibility to common diseases such as atherosclerosis. Twelve individual HAEC lines were treated in duplicate (in 35-mm dishes) for 4 h in media containing PAPC (40 μg/ml) or oxidized PAPC (OxPAPC) at 20 ug/ml or 40 μg/ml. Cytoplasmic RNA was isolated by using an RNAeasy kit (Qiagen) and analyzed on an Agilent 2100 Bioanalyzer (Agilent, Palo Alto, CA) to assess RNA integrity. Double-stranded cDNAs were synthesized from total RNA by using the cDNA Synthesis System (Invitrogen, Carlsbad, CA). Biotin-labeled cRNA was generated and used to probe Affymetrix (Santa Clara, CA) Human Genome U133 Plus 2.0 arrays according to the manufacturer’s recommendations (Affymetrix). The Microarray Suite 5.0 software (Affymetrix) was used to analyze image data and make the absolute call for each measurement. The array data were normalized with the median invariant method (31).
2011-06-13 | E-GEOD-29903 | biostudies-arrayexpress
Project description:Endothelial NOTCH1 is suppressed by circulating lipids and antagonizes inflammation during atherosclerosis
| PRJNA294525 | ENA