Atherosclerosis Formation In Mice Research Articles

Exploring the role of myeloperoxidase in the atherosclerotic process in hypoxic mice based on the MAPK signaling pathway

Atherosclerosis (AS) is the common pathophysiological basis of various cardiovascular diseases and the leading cause of death from cardiovascular disease worldwide. When the body is in a hypoxic environment, enhanced oxidative stress and significant accumulation of reactive oxygen species (ROS) in tissue cells exacerbate the inflammatory response, resulting in increased release of myeloperoxidase (MPO), catalyzing the formation of large quantities of hypochlorous acid (HOCl), further oxidative modification of low-density lipoprotein (LDL), and exacerbating the formation and progression of atherosclerotic plaques. The MAPK signaling pathway is important in oxidative stress-mediated promotion of atherogenesis. MPO -/- mice were used in this study to establish a hypoxia model simulating 5000 m altitude and a Western high-fat diet-induced atherosclerosis model for 12 weeks. Exploring the role of MPO in the atherosclerotic process in hypoxic mice by observing the MAPK signaling pathway to provide a therapeutic target for the prevention and treatment of hypoxic atherosclerotic disease in the plateau. We found that hypoxia promotes the formation of atherosclerosis in mice, and the mechanism may be that increased MPO in vivo promotes an inflammatory response, which plays a crucial role in the formation of atherosclerosis. In addition, hypoxia further exacerbates plaque instability by activating the MAPK signaling pathway to upregulate vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP9), which in turn promotes angiogenesis within the plaque. Therefore, a potential target for preventing and treating hypoxic atherosclerotic disease is the inhibition of MPO.

Global deficiency in the inflammatory chemokine receptors 1,2,3 and 5 ameliorates atherosclerosis and selectively modulates aortic myeloid cell phenotype

Abstract Background Atherosclerosis is a chronic inflammatory disease in which leukocytes traffic to the vessel wall, a process in part mediated by the inflammatory chemokine receptors (iCCRs): Ccr1, Ccr2, Ccr3 and Ccr5. ICCRs display significant ligand promiscuity and redundancy, which have hampered the definitive deduction of the isolated and collective roles of these receptors. Purpose To assess iCCR expression in human and mouse atherosclerosis and determine the contribution of global iCCR expression to atherosclerosis formation in mice. Methods We derived data from the human carotid endarterectomy biobanks, Athero-Express and Biobank of Karolinska Endarterectomy (BiKE) to investigate iCCR expression. Next, we employed fluorescent reporter mice (iREP) to track iCCR expression via flow cytometry and confocal microscopy. To ascertain the contribution of iCCRs to atherosclerosis, we utilised mice in which the complete iCcr locus incorporating Ccr1, Ccr2, Ccr3 and Ccr5 has been deleted (TACKO mice). We induced atherosclerosis using tail vein injection of AAV8- proprotein convertase subtilisin/kexin type 9 (PCSK9) in combination with 12- or 20-weeks Western diet (WD). Plaque size and composition was assessed via histology while cytometry of time-of-flight (CyTOF) and single-cell RNA-sequencing (scRNA-seq) were used to assess cellular changes. Results By analysing scRNA-seq data from Athero-Express, CCR1 emerged as the most strongly expressed iCCR and was highly localised to macrophages. Next, by utilizing BiKE data, we discovered that CCR1, 2 and 5 expression increased in plaques compared to healthy vessels, with CCR1 and CCR5 more strongly associated with symptomatic patients compared with asymptomatic patients. In iREP mice, CCR1, 2 and 5+ cell numbers were significantly increased in the atherosclerotic aorta compared with healthy mice. Cellular expression levels were variable with CCR2 being dominant in the plaque and increasing markedly on macrophages. Following 12 weeks WD, there was a reduction in mean plaque area within the aortic sinus of TACKO mice (WT 0.20 mm² vs. TACKO 0.10 mm², n=19-20, P<0.0001). Following 20 weeks WD, mean plaque area was also reduced in TACKO mice (WT 0.41 mm² vs. TACKO 0.25 mm², n=9-12, P<0.01) and this was associated with a significant reduction in necrotic core area and a reduction in calcification, reflecting a shift towards a more stable plaque phenotype, as classified according to the Virmani grading system. CyTOF analysis revealed reductions in inflammatory monocytes and conventional type 2 dendritic cells in TACKO aorta accompanied by a phenotypic switch in macrophages towards a less inflammatory phenotype. Finally, scRNA-seq analysis revealed phenotypic differences in non-immune vascular cells between WT and TACKO mice. Conclusion Our results support an athero-protective, plaque-stabilizing effect of iCcr deletion with a selective reduction in inflammatory myeloid cell aortic homing.

Phthalate promotes atherosclerosis through interacting with long-non coding RNA and induces macrophage foam cell formation and vascular smooth muscle damage

BackgroundPhthalates are commonly used in variety of plastic products. Previously it has been revealed that di (2-ethylhexyl) phthalate (DEHP), as the most common member of the class of phthalates, may disturb cholesterol homeostasis and deregulate the inflammatory response, and leading to accelerate the atherosclerosis process. In this regard, the aim of the current study is to explore the underlying mechanism of DEHP-induced atherosclerosis through the increasing of foam cell formation and Vascular Smooth Muscle Cells (VSMCs) damage via the interaction of long-non coding RNA (GAS5) and miR-145–5p. MethodsApoE−/− mice were used to evaluate the in vivo study. RAW264.7 and VSMCs were used to evaluate the effect of DEHP on formation of foam cell, cell proliferation, and cell damage in vitro. Animals were treated with DEHP (5% w/w of food) orally and cells were treated with medium containing of 100 μM DEHP; qRT-PCR, Western blotting, flowcytometry, IHC, oil red O, BODIPY, and autophagic vacuoles assay were used to evaluate the effect of DEHP on formation of atherosclerosis. ResultsDEHP significantly accelerated the formation of atherosclerosis in mice and alter the lipid profile in mice. In addition, after treating VSMCs with DEHP, GAS5 was significantly up-regulated and miR-145–5p was down-regulated. In VSMCs treated with DEHP, we observed that GAS5 could be used as the competing endogenous RNA (ceRNA) of miR-145–5p to regulate the proliferation and apoptosis of VSMCs; and the expression of GAS5 was correlated with the expression of miR-145–5p. DEHP increased the ox-LDL uptake by macrophage and increasing the formation of foam cells. Besides, GAS5 knocking down reversed the effect of DEHP on foam cell formation and ox-LDL uptake. ConclusionDEHP could accelerate the atherosclerosis process through increasing VSMCs damage and formation of macrophage foam cell by increasing lipid uptake though down regulating lncRNA GAS5 and altering in regulation of miR-145–5p.

Effects of coal-fired PM2.5 on the expression levels of atherosclerosis-related proteins and the phosphorylation level of MAPK in ApoE\u2212/\u2212 mice

BackgroundAir pollution increases the morbidity and mortality of cardiovascular disease (CVD). Atherosclerosis (AS) is the pathological basis of most CVD, and the progression of atherosclerosis and the increase of fragile plaque rupture are the mechanism basis of the relationship between atmospheric particulate pollution and CVD. The aim of the present study was to investigate the effects of coal-fired fine particulate matter (PM2.5) on the expression levels of atherosclerosis-related proteins (von Willebrand factor (vWF), Endothelin-1 (ET-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin, and to explore the role and mechanism of the progression of atherosclerosis induced by coal-fired PM2.5 via the mitogen-activated protein kinase (MAPK) signaling pathways.MethodsDifferent concentrations of PM2.5 were given to apolipoprotein-E knockout (ApoE−/−) mice via intratracheal instillation for 8 weeks. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of vWF, ET-1 in serum of mice. Immunohistochemistry was used to observe the expression and distribution of ICAM-1 and E-selectin in the aorta of mice. Western blot was used to investigate the phosphoylation of proteins relevant to MAPK signaling pathways.ResultsCoal-fired PM2.5 exacerbated atherosclerosis induced by a high-fat diet. Fibrous cap formation, foam cells accumulation, and atherosclerotic lesions were observed in the aortas of PM2.5-treated mice. Coal-fired PM2.5 increased the protein levels of ET-1, ICAM-1, and E-selectin, but there was no significant difference in the vWF levels between the PM2.5-treatment mice and the HFD control mice. Coal-fired PM2.5 promoted the phosphorylation of p38, c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) in aortic tissues of mice.ConclusionCoal-derived PM2.5 exacerbated the formation of atherosclerosis in mice, increased the expression levels of atherosclerosis-related proteins in mice serum, and promoted the phosphorylation of proteins relevant to MAPK signaling pathway. Thus, MAPK signaling pathway may play a role in the atherosclerosis pathogenesis induced by Coal-derived PM2.5.

Allergic asthma aggravated atherosclerosis increases cholesterol biosynthesis and foam cell formation in apolipoprotein E-deficient mice

Several studies have demonstrated that allergic asthma can induce atherosclerosis formation in mice. Moreover, allergic asthma and atherosclerosis have been shown to be strongly associated with dyslipidemia. In this study, we investigated the underlying mechanism of allergic asthma-aggravated atherosclerosis-induced cholesterol metabolism disorder in asthmatic apoE−/- mice. We found that allergic asthma increased the expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) in the liver and CD36 in the aorta during the acute and advanced stages of atherosclerosis, respectively. These results indicate that cholesterol biosynthesis is increased during acute atherosclerosis and cholesterol uptake and foam cell formation is increased during advanced atherosclerosis. Simvastatin administration significantly ameliorated the aortic root lesion size of asthmatic mice and significantly decreased HMGCR and CD36 expression. However, the expression of the low-density lipoprotein receptor and ATP-binding cassette transporter A1 was markedly increased, indicating that the beneficial effect of statins in allergic asthma and coronary artery disease was mediated, at least in part, by decreasing cholesterol biosynthesis and foam cell formation. In conclusion, allergic asthma aggravates atherosclerosis by regulating cholesterol metabolism in apoE−/- mice. Allergic asthma selectively promotes cholesterol biosynthesis in acute atherosclerosis and increases foam cell formation in advanced atherosclerosis.

ApoAI-derived peptide increases glucose tolerance and prevents formation of atherosclerosis in mice

Aims/hypothesisFinding new treatment alternatives for individuals with diabetes with severe insulin resistance is highly desired. To identify novel mechanisms that improve glucose uptake in skeletal muscle, independently from insulin levels and signalling, we have explored the therapeutic potential of a short peptide sequence, RG54, derived from apolipoprotein A-I (ApoA-I).MethodsINS-1E rat clonal beta cells, C2C12 rat muscle myotubes and J774 mouse macrophages were used to study the impact of RG54 peptide on glucose-stimulated insulin secretion, glucose uptake and cholesterol efflux, respectively. GTTs were carried out on diet-induced insulin-resistant and Leprdb diabetic mouse models treated with RG54 peptide, and the impact of RG54 peptide on atherosclerosis was evaluated in Apoe−/− mice. Control mice received ApoA-I protein, liraglutide or NaCl.ResultsThe synthetic RG54 peptide induced glucose uptake in cultured muscle myotubes by a similar amount as insulin, and also primed pancreatic beta cells for improved glucose-stimulated insulin secretion. The findings were verified in diet-induced insulin-resistant and Leprdb diabetic mice, jointly confirming the physiological effect. The RG54 peptide also efficiently catalysed cholesterol efflux from macrophages and prevented the formation of atherosclerotic plaques in Apoe−/− mice.Conclusions/interpretationThe RG54 peptide exhibits good prospects for providing glucose control and reducing the risk of cardiovascular disease in individuals with severe insulin resistance.

A Novel Inhibitory Effect of Naloxone on Macrophage Activation and Atherosclerosis Formation in Mice

We investigated whether naloxone could reduce macrophage activation and influence atherosclerotic lesion formation in mice. Macrophages play an important role in the inflammatory process in atherosclerosis. Naloxone could inhibit activation of microglia, the resident macrophage in the nervous system. The anti-inflammatory effect of naloxone was evaluated by stimulating the macrophage cell culture and FVB mice with lipopolysaccharide or oxidized low-density lipoprotein with and without naloxone pretreatment. Apolipoprotein-E (apoE)-deficient mice received naloxone injection for 10 weeks, and the severity of aortic atherosclerosis was measured. The left common carotid arteries of C57BL/6 mice were ligated near the carotid bifurcation. The mice then received naloxone injection for 4 weeks after ligation, and the severity of neointima formation was evaluated. Naloxone pretreatment significantly suppressed the production of tumor necrosis factor-alpha (TNF-alpha), interleukin-6, monocyte chemoattractant protein-1, and superoxide in macrophages after stimulation. In FVB mice, naloxone reduced the TNF-alpha level in circulation, inflammatory cell infiltration in lungs, and superoxide production in aorta. Naloxone injection significantly decreased the severity of aortic atherosclerosis in the apoE-deficient mice and carotid neointima formation in the C57BL/6 mice after ligation. Naloxone, with its novel anti-inflammatory effect, significantly reduces atherosclerosis and neointima formation in mice.

N-[2-[ N′-pentyl-(6,6-dimethyl-2,4-heptadiynyl) amino]ethyl]-(2-methyl-1-naphthylthio)acetamide (FY-087) : A new acyl coenzyme A:Cholesterol acyltransferase (ACAT) inhibitor of diet-induced atherosclerosis formation in mice

FY-087 ( N-[2-[ N′-pentyl-(6,6-dimethyl-2,4-heptadiynyl)amino]ethyl]-(2-methyl-1-naphthyl-thio)acetamide) was found to be a competitive inhibitor of human microsomal acyl coenzyme A:cholesterol acyltransferase (ACAT) with an ic 50 value of 0.11 μM. Under our assay conditions, other ACAT inhibitors tested, specifically YM-750, E-5324, and melinamide, all of which are now in phase I clinical trials or in clinical use in Japan, inhibited this enzyme with ic 50 values of 0.18, 0.14, and 3.2 μM, respectively. FY-087 also inhibited ACAT in acetyl-low density lipoprotein loaded human macrophages (THP-1 cells) with an ic 50 of 0.17 μM. Following the oral administration of FY-087 (30 mg/ kg) to rats, the plasma concentration of FY-087 reached 0.42 μg/mL after 2 hr. This concentration of FY-087 was enough to inhibit blood vessel ACAT activity. Cholesterol-lowering and anti-atherogenic effects of FY-087 were examined using C57BL/6J mice fed an atherogenic diet. In this mouse model, treatment with FY-087 (28 mg/kg) inhibited the increase in plasma cholesterol levels by about 20% and decreased the hepatic accumulation of free and esterified cholesterol by 61 and 67%, respectively. FY-087 also significantly inhibited the atherogenic diet-induced increase in the fatty-streak lesion area of the proximal aorta by 57% in C57BL/6J mice. These results indicate that FY-087 is not only a therapeutically bioavailable ACAT inhibitor that lowers plasma cholesterol levels, but also an effective anti-atherogenic agent in mice fed an atherogenic diet.