Aortic Plaque Research Articles

Single-cell profiling of CD11c+ B cells in atherosclerosis.

Circulating CD11c+ B cells, a novel subset of activated B cells, have been linked to autoimmunity and shown to expand with age. Atherosclerosis is an age-associated disease that involves innate and adaptive immune responses to modified self-antigens. Yet, the expression of CD11c on specific B-cell subtypes and its link to atherosclerosis are poorly understood. In this study, we characterized the frequency of CD11c+ B cells in tissues in mice with aging. We observed an age-associated increase in CD11c+ B cells in the spleen and bone marrow of ApoE-/- mice, and this was associated with an increase in aortic plaque. In addition, we also utilized single-cell multi-omics profiling of 60 human subjects undergoing advanced imaging for coronary artery disease (CAD) to subtype CD11c+ B cells and determine their frequency in subjects with high and low severity of CAD. Using unsupervised clustering, we identified four distinct clusters of CD11c+ B cells, which include CD27 and IgD double negative 2 (DN2), age-associated (ABC), CD11c+ unswitched memory (USWM), and activated Naïve (aNav) B cells. We observed an increase in the frequency of both ABC B cells and DN2 B cells in patients with high CAD severity. Pathway analysis further demonstrated augmentation of autophagy, IFNg signaling, and TLR signaling in DN2 cells in high-severity CAD patients. On the other hand, an increase in the negative regulator of BCR signaling through CD72 was found in ABC cells in low-severity CAD patients. Through investigating scRNAseq of atheroma, these DN2 cells were also found to infiltrate human coronary atheroma.

A very-low carbohydrate content in a high-fat diet modifies the plasma metabolome and impacts systemic inflammation and experimental atherosclerosis

Ketogenic diets (KDs) are very high-fat low-carbohydrate diets that promote nutritional ketosis and are widely used for weight loss, although concerns about potential adverse cardiovascular effects remain. We investigated a very high-fat KD's vascular impact and plasma metabolic signature compared to a non-ketogenic high-fat diet (HFD). Apolipoprotein E deficient (ApoE −/−) mice were fed a KD (%kcal:81:1:18, fat/carbohydrate/protein), a non-ketogenic high-fat diet with half of the fat content (HFD) (%kcal:40:42:18, fat/carbohydrate/protein) for 12 weeks. Plasma samples were used to quantify the major ketone body beta−hydroxybutyrate (BHB) and several pro-inflammatory cytokines (IL-6, MCP-1, MIP-1alpha, and TNF alpha), and to targeted metabolomic profiling by mass spectrometry. In addition, aortic atherosclerotic lesions were quantified ex-vivo by magnetic resonance imaging (MRI) on a 14-tesla system. KD was atherogenic when compared to the control diet, but KD mice, when compared to the HFD group (1) had markedly higher levels of BHB and lower levels of cytokines, confirming the presence of ketosis that alleviated the well-established fat-induced systemic inflammation; (2) displayed significant changes in the plasma metabolome that included a decrease in lipophilic metabolites and an increase in hydrophilic metabolites; (3) had significantly lower levels of several atherogenic lipid metabolites, including phosphatidylcholines, cholesterol esters, sphingomyelins, and ceramides; and (4) presented significantly lower aortic plaque burden. KD was atherogenic and was associated with specific metabolic changes but alleviated the fat-induced inflammation and lessened the progression of atherosclerosis when compared to the HFD.

Zexieyin formula alleviates atherosclerosis with cognitive impairment: A novel role in the treatment of comorbidities and its underlying mechanisms

Ethnopharmacological relevancyZexieyin formula (ZXYF) has been identified to have therapeutic actions of atherosclerosis (AS). It's unknown that whether ZXYF has therapeutic potential of atherosclerosis (AS) with cognitive impairment (CI) and its underlying mechanisms. Aim of the studyTo elucidate therapeutic effect of ZXYF for AS with CI as well as its underlying mechanisms in AS with CI mice model. Methods and materialsTo establish AS with CI model, we fed ApoE−/− mice with high-fat diet (HFD) for 8 weeks. Oil red O staining (ORO) and Hematoxylin-eosin staining (HE) were used to detect aortic plaque area. Morris water maze (MWM) and Y-maze were used to measure cognitive function and cognitive improvement after administration of ZXYF and atorvastatin (ATO). Network pharmacology was used to screen for potential mechanisms for improving cognitive function. Western blot was used to detect expressions of MAPK, Aβ and synaptic proteins in hippocampus. ResultsHFD caused and accelerated the AS in ApoE−/− mice, while it was easier able to produce CI than normal mice. Administration of ZXYF or ATO for 8 weeks significantly reduced aortic plaque area in ORO and HE tests, and improved cognitive abilities in MWM and Y-maze tests. Network pharmacology results showed that MAPK or synaptic proteins were highly associated with CI. HFD contributed to abnormal expressions of MAPK (pERK, pP38, pJNK), NF-kB, synaptic proteins (PSD95, synapsin1) and β-amyloid (Aβ) in hippocampus, which were all reversed by ZXYF. However, ERK and PSD95 expressions were not reversed by ATO in hippocampus. ConclusionsZXYF mitigated AS, further alleviating CI by modulating MAPK signaling, relating to synaptic proteins enhancing and Aβ protein decreasing in the hippocampus. This study firstly lit up the new clinical application of ZXYF, which might promote the use of ZXYF in AS and CI patients.

Methyl-CpG-binding 2 K271 lactylation-mediated M2 macrophage polarization inhibits atherosclerosis.

Rationale: Posttranslational modifications of proteins have not been addressed in studies aimed at elucidating the cardioprotective effect of exercise in atherosclerotic cardiovascular disease (ASCVD). In this study, we reveal a novel mechanism by which exercise ameliorates atherosclerosis via lactylation. Methods: Using ApoE-/- mice in an exercise model, proteomics analysis was used to identify exercise-induced specific lactylation of MeCP2 at lysine 271 (K271). Mutation of the MeCP2 K271 lactylation site in aortic plaque macrophages was achieved by recombinant adenoviral transfection. Explore the molecular mechanisms by which motility drives MeCP2 K271 lactylation to improve plaque stability using ATAC-Seq, CUT &Tag and molecular biology. Validation of the potential target RUNX1 for exercise therapy using Ro5-3335 pharmacological inhibition. Results: we showed that in ApoE-/- mice, methyl-CpG-binding protein 2 (MeCP2) K271 lactylation was observed in aortic root plaque macrophages, promoting pro-repair M2 macrophage polarization, reducing the plaque area, shrinking necrotic cores, reducing plaque lipid deposition, and increasing collagen content. Adenoviral transfection, by introducing a mutant at lysine 271, overexpressed MeCP2 K271 lactylation, which enhanced exercise-induced M2 macrophage polarization and increased plaque stability. Mechanistically, the exercise-induced atheroprotective effect requires an interaction between MeCP2 K271 lactylation and H3K36me3, leading to increased chromatin accessibility and transcriptional repression of RUNX1. In addition, the pharmacological inhibition of the transcription factor RUNX1 exerts atheroprotective effects by promoting the polarization of plaque macrophages towards the pro-repair M2 phenotype. Conclusions: These findings reveal a novel mechanism by which exercise ameliorates atherosclerosis via MeCP2 K271 lactylation-H3K36me3/RUNX1. Interventions that enhance MeCP2 K271 lactylation have been shown to increase pro-repair M2 macrophage infiltration, thereby promoting plaque stabilization and reducing the risk of atherosclerotic cardiovascular disease. We also established RUNX1 as a potential drug target for exercise therapy, thereby providing guidance for the discovery of new targets.

Protocatechuic acid alleviates TMAO-aggravated atherosclerosis via mitigating inflammation, regulating lipid metabolism, and reshaping gut microbiota.

Trimethylamine-N-oxide (TMAO) is a risk factor for atherosclerosis. As a natural phenolic acid, protocatechuic acid (PCA) is abundant in various plant foods. The present study investigated the effect of PCA on TMAO-aggravated atherosclerosis in ApoE-/- mice. The mice were randomly divided into five groups and fed one of the following five diets for 12 weeks: namely a low-fat diet (LFD), a western diet (WD), a WD + 0.2% TMAO diet (WDT), a WDT + 0.5% PCA diet (WDT + LPCA), and a WDT + 1.0% PCA diet (WDT + HPCA). Results demonstrated that dietary TMAO exacerbated the development of atherosclerosis by eliciting inflammation and disturbing lipid metabolism. The diet with PCA at 1% reduced TMAO-induced aortic plaque by 30% and decreased the levels of plasma pro-inflammatory cytokines. PCA also improved lipid metabolism by up-regulating the hepatic gene expression of peroxisome proliferator-activated receptor alpha (PPARα). In addition, PCA supplementation enhanced fecal excretion of fatty acids and decreased hepatic fat accumulation. PCA supplementation favorably modulated gut microbiota by increasing the α-diversity with an increase in the abundance of beneficial genera (Rikenella, Turicibacter, Clostridium_sensu_stricto and Bifidobacterium) and a decrease in the abundance of the harmful Helicobacter genus. In summary, PCA could alleviate the TMAO-exacerbated atherosclerosis and inflammation, improve the lipid metabolism, and modulate gut microbiota.

Multimodal Cardiac Imaging in the Assessment of Patients Who Have Suffered a Cardioembolic Stroke: A Review.

Cardioembolic strokes account for 20-25% of all ischaemic strokes, with their incidence increasing with age. Cardiac imaging plays a crucial role in identifying cardioembolic causes of stroke, with early and accurate identification affecting treatment, preventing recurrence, and reducing stroke incidence. Echocardiography serves as the mainstay of cardiac evaluation. Transthoracic echocardiography (TTE) is the first line in the basic evaluation of structural heart disorders, valvular disease, vegetations, and intraventricular thrombus. It can be used to measure chamber size and systolic/diastolic function. Trans-oesophageal echocardiography (TOE) yields better results in identifying potential cardioembolic sources of stroke and should be strongly considered, especially if TTE does not yield adequate results. Cardiac computed tomography and cardiac magnetic resonance imaging provide better soft tissue characterisation, high-grade anatomical information, spatial and temporal visualisation, and image reconstruction in multiple planes, especially with contrast. These techniques are useful in cases of inconclusive echocardiograms and can be used to detect and characterise valvular lesions, thrombi, fibrosis, cardiomyopathies, and aortic plaques. Nuclear imaging is not routinely used, but it can be used to assess left-ventricular perfusion, function, and dimensions and may be useful in cases of infective endocarditis. Its use should be considered on a case-by-case basis. The accuracy of each imaging modality depends on the likely source of cardioembolism, and the choice of imaging approach should be tailored to individual patients.

Novel role of peptidoglycan recognition protein 2 in activating NOD2-NFκB inflammatory axis in coronary artery disease

Backgrounds and aimsThe role of inflammation in driving atherosclerosis is well-established. It exerts systemic effects beyond the local site of plaque formation. In the context of coronary artery disease (CAD), the proteins that show altered levels in the plasma, are potentially important for understanding the key regulatory mechanism in the pathogenesis of atherosclerosis. A case-control study revealed that plasma soluble Peptidoglycan Recognition Protein 2 (PGLYRP2) primarily produced by the liver, is increased in subjects with CAD. Furthermore, the concentration of PGLYRP2 in the blood correlates with the severity of coronary artery disease. Thus, it raises interest in understanding the exact role of the protein in aortic inflammation and plaque progression. MethodsWe evaluated the plasma concentration of PGLYRP2 in three distinct groups: patients with CAD (N = 68), asymptomatic individuals (N = 34), and healthy volunteers (N = 20). Furthermore, we investigated the correlation between disease severity and PGLYRP2 levels in CAD patients. To identify potential binding partners of PGLYRP2, we employed computational analysis. We verified the PGLYRP2-NOD2 interaction in macrophage cells and elucidated the inflammatory pathways activated by PGLYRP2 within these cells. To assess the impact of PGLYRP2, we examined its effects in the atherosclerotic mice model (ApoE−/−). ResultsIn this study, we report for the first time that Nucleotide-binding Oligomerization domain 2 (NOD2) which is expressed on the surface of macrophages, is a receptor of PGLYRP2. The N-terminal domain of PGLYRP2 directly binds to NOD2 and activates the NOD2-RIP2-NFκB cascade that promotes the secretion of proinflammatory cytokines like TNFα, IL1β, and IL-8. In the atherosclerotic mice model (ApoE−/−) we demonstrate that elevated PGLYRP2 level is parallel with increased proinflammatory cytokines in the plasma when fed a High Cholesterol Diet (HCD). Immunohistochemical analysis reveals that PGLYRP2 is co-localized with NOD2 on the macrophages at the site of the lesion. ConclusionsTaken together, our data demonstrate that NOD2 acts as a receptor of PGLYRP2 on macrophages, which mediates the activation of the NOD2-RIP2-NFκB pathway and promotes inflammation, thus significantly contributing to the development and progression of atherosclerosis.

The Duality of Adiponectin: The Role of Sex in Atherosclerosis.

The hormone adiponectin has many beneficial effects in atherosclerosis, as gene deficiency in adiponectin or its receptor has shown detrimental effects on plaque burden in mice. Our objective was to understand the potential roles adiponectin deficiency has on aortic plaque content, inflammation, and markers of cardiovascular disease according to sex and age. To study the influence of adiponectin status on sex and atherosclerosis, we used young male and female adipoq-/-apoe-/-, adipoq+/-apoe-/-, and apoe-/- mice, which were given a high-fat diet (HFD). Even a 50% reduction in the expression of adiponectin led to a plaque reduction in males and an increase in females compared with apoe-/- controls. Changes in plaque were not attributed to changes in cholesterol or cardiovascular disease markers but correlated with inflammatory markers. Plaque reduction in males was associated with reduced monocyte chemoattractant protein 1 (MCP1) and increased colony stimulating factor 3 (CSF3), while the increase in plaque in females correlated with the opposite effect in these markers. In old mice, both adiponectin-deficient genotypes and sexes accumulated more plaque than their respective apoe-/- controls. The increase in plaque with adiponectin deficiency according to age was not explained by a worsening lipid profile but correlated with increased levels of C-C motif chemokine ligand 5 (CCL5). Overall, our study uncovered genotype-specific effects that differed by sex and age of adiponectin deficiency in atherosclerosis.

Macrophage-targeted ultrasound nanobubbles for highly efficient sonodynamic therapy of atherosclerotic plaques by modulating M1-to-M2 polarization

Background and aimsSonodynamic therapy (SDT) is a new approach for the treatment of atherosclerosis (AS), yet the poor targeting ability of sonosensitizers limits its therapeutic efficacy. Herein, we reported a plaque-targeted nanoplatform modified with macrophage type A scavenger receptor (SR-A)-targeted peptide (designated as SR-A-Ce6NB) to augment the efficacy of low-intensity pulsed ultrasound (LIPUS)-mediated SDT of atherosclerotic plaque. MethodsSR-A-Ce6NB was fabricated by thin hydration method and biotin-avidin system, and its physicochemical properties, biocompatibility and plaque-targeting ability were investigated. RAW 264.7 cells were used for in vitro experimental studies. Male 6-week-old apolipoprotein E-deficient mice were fed a high-fat diet for 16 weeks to induce aortic atherosclerotic plaques. Plaque-bearing mice were randomly allocated into five groups (n = 6): control group, Ce6 + LIPUS group, Ce6NB + LIPUS group, SR-A-Ce6NB + LIPUS group and atorvastatin group. After treatment in each group, the aortic artery was harvested for Oil red O, H&E, Masson's trichrome staining, immunohistochemical and immunofluorescent staining. ResultsSR-A-Ce6NB with high stability and excellent biocompatibility was successfully fabricated. SR-A-Ce6NB could actively target activated macrophages and selectively accumulate in the plaque. SR-A-Ce6NB could be triggered by LIPUS and had a more potent sonodynamic effect than free Ce6 to potentiate SDT. SR-A-Ce6NB-mediated SDT enhanced the anti-atherogenic effect via modulating M1-to-M2 macrophage polarization and had an earlier onset of action on plaque than the statin-mediated effect. No apparent side effect was observed after intravenous SR-A-Ce6NB injection and LIPUS exposure. ConclusionsMacrophage-targeted nanoplatform SR-A-Ce6NB-mediated SDT provides a safe, effective and preferable anti-atherogenic therapy by mediating M1-to-M2 macrophage polarization.

Aortic Arch Plaques and the Long-Term Risk of Stroke and Cardiovascular Events in the Statin Era.

Aortic arch plaques are associated with an increased risk of ischemic stroke in patients with cryptogenic stroke or prior embolic events. However, this relationship is unclear in the community. We investigated (1) the long-term risk of stroke and cardiovascular events associated with arch plaques and (2) whether statin therapy prescribed for any indication modified the association. A total of 934 stroke-free participants (72±9 years; 37% men) from the CABL study (Cardiovascular Abnormalities and Brain Lesion) were evaluated. Arch plaques were assessed by suprasternal transthoracic echocardiography; plaques ≥4 mm in thickness were classified as large plaques. The primary outcome was ischemic stroke; the secondary outcome was combined cardiovascular events (ischemic stroke, myocardial infarction, and cardiovascular death). The plaque-related risk of outcomes was also analyzed according to the presence of statin treatment. No plaque was used as a reference. Aortic arch plaques were present in 645 participants (69.1%), with large plaques in 114 (12.2%). During a mean follow-up of 11.3±3.6 years, 236 (25.3%) cardiovascular events occurred (76 ischemic strokes, 27 myocardial infarctions, and 133 cardiovascular deaths). Large arch plaques were independently associated with combined events (adjusted hazard ratio, 2.19 [95% CI, 1.40-3.43]) but not stroke alone (adjusted hazard ratio, 1.09 [95% CI, 0.50-2.38]). The association between large plaques and cardiovascular events was significant in participants receiving statins (adjusted hazard ratio, 2.57 [95% CI, 1.52-4.37]) but not in others; however, participants on statin treatment also had a worse risk profile (higher body mass index, greater frequencies of hypertension, diabetes, and coronary artery disease). Aortic arch plaques may be a marker of cardiovascular risk rather than a direct embolic stroke source in older adults without prior stroke. The efficacy of broader cardiovascular risk factors control, beyond cholesterol levels alone, for primary prevention of cardiovascular events in individuals with aortic arch plaques may require further investigation.

Embolic stroke of undetermined source - ESUS: a new challenge for the cardiologist

Embolic stroke of undetermined source (ESUS) is a new clinical entity which requires an accurate diagnostic work-up. The involvement of the cardiologist in this challenge is of paramount importance as in many cases an occult cardioembolic source is detectable. In the absence of subclinical atrial fibrillation detectable by prolonged ECG monitoring, potential causes of ESUS include atrial cardiomyopathy, left ventricular dysfunction or heart failure, aortic plaques and patent foramen ovale. The assessment of a causal relationship between these cardiac abnormalities and ESUS is often a difficult challenge and the randomized clinical trials performed so far failed to demonstrate a superiority of direct oral anticoagulants (DOAC) over aspirin. In selected individual patients with heart failure in sinus rhythm with high CHA2DS2-VASc score, or aortic plaques with thrombotic component, oral anticoagulation with DOAC might be considered even in the absence of evidence from randomized clinical trials.

Amelioration of Atherosclerosis by lycopene is linked to the modulation of gut microbiota dysbiosis and related gut-heart axis activation in high-fat diet-fed ApoE−/− mice

BackgroundInterplay between gut microbiota and heart, termed “gut-heart” axis, has a crucial role in the pathogenesis of atherosclerosis. Our previous study showed that lycopene possesses anti-inflammatory and anti-atherosclerotic effects, but its link to the gut microbiota is poorly understood. Herein, we surmised that lycopene could regulate the gut microbiota, exert anti-atherosclerotic effect by regulating the “gut-heart” axis.MethodsMale ApoE−/− mice were fed a high-fat diet (HFD) with or without lycopene (0.1% w/w) for 19 weeks. Gut microbiota was analyzed by 16 S rRNA sequencing, the protein levels of zonula occludens-1 (ZO-1), occludin, toll-like receptor 4 (TLR4) and phospho-nuclear factor-κB (NF-κB) p65 were measured by Western blotting, the levels of serum inflammatory factors including monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-6 were assayed using ELISA kits. Also, the concentrations of serum lipopolysaccharide (LPS), D-lactic acid (D-LA) and diamine peroxidase (DAO) were measured through ELISA method.ResultsThe aortic sinus sections revealed that lycopene supplementation significantly reduced the extent of atherosclerotic lesions and inhibited atherosclerosis development caused by HFD. The analysis of gut microbiota showed that lycopene reduced the ratio of Firmicutes/Bacteroides and increased the relative abundance of Verrucomicrobia, Akkermansia and Alloprevotella, which were related to elevated intestinal barrier function and reduced inflammation. Moreover, lycopene up-regulated the expression of intestinal ZO-1 and occludin and decreased serum LPS, D-LA and DAO levels. In addition, lycopene inhibited the expression of TLR4 and phospho-NF-κB p65 in aortic sinus plaque, serum MCP-1, TNF-α, IL-1β, and IL-6 levels were also lowered by lycopene treatment.ConclusionsOur results indicated the protective effect of lycopene against atherosclerosis induced by HFD and further revealed that its mechanism might be its prebiotic effect on maintaining gut microbiota homeostasis and improving intestinal barrier function, consequently reducing serum LPS-triggered inflammatory response in the heart.

Association between aortic thrombi detected using non-obstructive general angioscopy and atrial fibrillation.

Atrial fibrillation (AF) is an independent risk factor for stroke and systemic embolism. Cardiogenic and aortogenic emboli are causes of stroke or systemic embolism. Non-obstructive general angioscopy (NOGA) can be used to diagnose aortic intimal findings, including thrombi and atherosclerotic plaques, but little is known about NOGA-derived aortic intimal findings in patients with AF. This study focused on aortic intimal findings in patients with AF and evaluated the association between AF and aortic thrombi detected using NOGA. We enrolled 283 consecutive patients with coronary artery disease who underwent NOGA of the aorta between January 2017 and August 2022. Aortic intimal findings were screened using NOGA after coronary arteriography. The patients were divided into two groups according to their AF history (AF, n = 50 and non-AF, n = 233). Patients in the AF group were older than those in the non-AF group. Sex, body mass index, and coronary risk factors were not significantly different between the two groups. In the NOGA findings, the presence of intense yellow plaques and ruptured plaques was not significantly different between the two groups. Aortic thrombi were more frequent in the AF group than in the non-AF group (92.0 vs. 71.6%, p < 0.001). Multivariate logistic regression found that AF was independently associated with aortic thrombi (odds ratio 3.87 [95% CI 1.28-11.6], p = 0.016). The presence of aortic thrombi observed using NOGA was associated with AF in patients with coronary artery disease. The roles of aortic thrombi as well as cardiogenic embolism may require clarification.

Mechanisms and Therapeutic Strategies to Reverse TET2 Mutant Clonal Hematopoiesis and the Risk of MDS, AML, and Atherosclerotic Cardiovascular Disease

Inactivating mutations of the epigenetic modifier TET2 are frequent in myeloid malignancies and clonal hematopoiesis of indeterminate potential (CHIP). These mutations in hematopoietic stem and progenitor cells (HSPCs) impart a clonal advantage with increased self-renewal. CHIP is associated with an increased risk of hematological malignancies and, surprisingly, with atherosclerotic cardiovascular disease, supported by studies in mice and humans. The heightened cardiovascular risk in CHIP patients may result from a pro-inflammatory slant of the mutant HSPC progeny, particularly monocytes and macrophages, independent from the risk imparted by elevated LDL cholesterol levels. Targeting molecules like IL-1β may provide a strategy to lower the risk of cardiovascular morbidity in patients with CHIP. However, as an alternative strategy, administration of drugs that selectively suppress the growth of mutant CHIP clones in the bone marrow (BM) might reduce the risk of both i) progression to MDS or hematologic malignancy and of ii) atherosclerotic cardiovascular disease. This study focused on identifying drugs that reverse the inflammatory and atherosclerotic properties of TET2-mutant macrophages. Our previous investigations demonstrated that the nuclear export inhibitor eltanexor selectively kills Tet2-mutant HSPCs in zebrafish embryos and reduces the number of mutant colonies in murine colony formation assays. A murine competitive repopulation model of CHIP tested the in vivo efficacy of eltanexor. We found that eltanexor selectively reduces Tet2-mutant circulating monocytes while having no effect on overall total white blood cell counts. Moreover, a second trial using Ldlr-/- atherosclerosis-prone mice consuming a high-cholesterol diet, showed that treatment with eltanexor selectively reduces the amount of aortic atherosclerotic plaque formation as well as spleen weight in mice injected with Tet2+/- (mutant) BM cells, whereas eltanexor had no significant effect in mice injected with Tet2+/+ (wild-type) BM cells. Single-cell CITE-seq analysis of Ldlr-/- mice injected with a mix of Tet2+/- and Tet2+/+ BM cells showed that eltanexor selectively reduces the percentage of Tet2+/- HSPCs. Additionally, CITE-seq analysis revealed a significant reduction in pro-inflammatory macrophages in the arterial wall after eltanexor treatment, along with a decrease in IL-1β expression by these cells. As TET2 functions in mechanistically regulating gene regulation through epigenetic DNA methylation, we employed ChIP-seq and CUT&amp;RUN-Seq technologies to investigate the mechanisms underlying enhancer dysregulation caused by loss of Tet2. Enhancer profiling of bone marrow-derived macrophages showed that the negative regulator of the macrophage inflammatory response, ATF3, is associated with a super-enhancer. Notably, the ATF3 super-enhancer emerged as the largest super-enhancer in wild-type macrophages but was reduced in Tet2-mutant macrophages. Our investigation also revealed that ATF3 binds with IL-1β at H3K27ac modified regions, and this binding was significantly diminished in Tet2-mutant macrophages. Remarkably, treatment with eltanexor restored the binding of ATF3 to IL1β, providing a mechanism to explain the anti-inflammatory effect of eltanexor concordant with our in vivo results. In summary, this study proposes eltanexor treatment as a novel therapeutic approach to specifically target TET2-mutant cells in individuals with CHIP and sheds light on the underlying mechanisms of the pro-inflammatory slant of TET2-mutant macrophages. These findings offer potential new avenues for precise therapies in humans with CHIP and associated cardiovascular risks from atherosclerosis.

Bacterial butyrate mediates the anti-atherosclerotic effect of silybin

Silybin (SIL) is a versatile bioactive compound used for improving liver damage and lipid disorders and is also thought to be beneficial for atherosclerosis (AS). The goal of this study was to investigate the efficacy of SIL in the treatment of AS in ApoE−/−mice fed a high-fat diet and explore the mechanism underlying treatment outcomes. We found that SIL significantly alleviated AS-related parameters, including the extent of aortic plaque formation, hyperlipidemia, and adhesion molecule secretion in the vascular endothelium. 16 S rRNA gene sequencing analysis, together with the application of antibiotics, showed that intestinal butyrate-producing bacteria mediated the ameliorative effect of SIL on AS. Further analysis revealed that SIL facilitated butyrate production by increasing the level of butyryl-CoA: acetate CoA-transferase (BUT). The increased expression of monocarboxylic acid transporter-1 (MCT1) induced by butyrate and MCT4 induced by SIL in the apical and basolateral membranes of colonocytes, respectively, resulted in enhanced absorption of intestinal butyrate into the circulation, leading to the alleviation of arterial endothelium dysfunction. Moreover, the SIL-mediated increase in intestinal butyrate levels restored gut integrity by upregulating the expression of tight junction proteins and promoting gut immunity, thus inhibiting the AS-induced inflammatory response. This is the first study to show that SIL can alleviate AS by modulating the production of bacterial butyrate and its subsequent absorption.

Circular RNA ZBTB46 depletion alleviates the progression of Atherosclerosis by regulating the ubiquitination and degradation of hnRNPA2B1 via the AKT/mTOR pathway

BackgroundCircZBTB46 has been identified as being associated with the risk of coronary artery disease (CAD) and has the potential to be a diagnostic biomarker for CAD. However, the specific function and detailed mechanism of circZBTB46 in CAD are still unknown.MethodsThe expression levels and properties of circRNAs were examined using qRT‒PCR, RNA FISH, and subcellular localization analysis. ApoE−/− mice fed a high-fat diet were used to establish an atherosclerosis model. HE, Masson, and Oil Red O staining were used to analyze the morphological features of the plaque. CCK-8, Transwell, and wound healing assays, and flow cytometric analysis were used to evaluate cell proliferation, migration, and apoptosis. RNA pull-down, silver staining, mass spectrometry analysis, and RNA-binding protein immunoprecipitation (RIP) were performed to identify the interacting proteins of circZBTB46.ResultsCircZBTB46 is highly conserved and is significantly upregulated in atherosclerotic lesions. Functional studies revealed that knockdown of circZBTB46 significantly decreased the atherosclerotic plaque area, attenuating the progression of atherosclerosis. In addition, silencing circZBTB46 inhibited cell proliferation and migration and induced apoptosis. Mechanistically, circZBTB46 physically interacted with hnRNPA2B1 and suppressed its degradation, thereby regulating cell functions and the formation of aortic atherosclerotic plaques. Additionally, circZBTB46 was identified as a functional mediator of PTEN-dependent regulation of the AKT/mTOR signaling pathway and thus affected cell proliferation and migration and induced apoptosis.ConclusionOur study provides the first direct evidence that circZBTB46 functions as an important regulatory molecule for CAD progression by interacting with hnRNPA2B1 and regulating the PTEN/AKT/mTOR pathway.

Essential oil from Fructus Alpinia zerumbet ameliorates atherosclerosis by activating PPARγ-LXRα-ABCA1/G1 signaling pathway

BackgroundAtherosclerosis (AS) is a progressive chronic disease. Currently, cardiovascular diseases (CVDs) caused by AS is responsible for the global increased mortality. Yanshanjiang as miao herb in Guizhou of China is the dried and ripe fruit of Fructus Alpinia zerumbet. Accumulated evidences have confirmed that Yanshanjiang could ameliorate CVDs, including AS. Nevertheless, its effect and mechanism on AS are still largely unknown. PurposeTo investigate the role of essential oil from Fructus Alpinia zerumbet (EOFAZ) on AS, and the potential mechanism. MethodsA high-fat diet (HFD) ApoE−/− mice model of AS and a oxLDL-induced model of macrophage-derived foam cells (MFCs) were reproduced to investigate the pharmacological properties of EOFAZ on AS in vivo and foam cell formation in vitro, respectively. The underlying mechanisms of EOFAZ were investigated using Network pharmacology and molecular docking. EOFAZ effect on PPARγ protein stability was measured using a cellular thermal shift assay (CETSA). Pharmacological agonists and inhibitors and gene interventions were employed for clarifying EOFAZ's potential mechanism. ResultsEOFAZ attenuated AS progression in HFD ApoE−/− mice. This attenuation was manifested by the reduced aortic intima plaque development, increased collagen content in aortic plaques, notable improvement in lipid profiles, and decreased levels of inflammatory factors. Moreover, EOFAZ inhibited the formation of MFCs by enhancing cholesterol efflux through activiting the PPARγ-LXRα-ABCA1/G1 pathway. Interestingly, the pharmacological knockdown of PPARγ impaired the beneficial effects of EOFAZ on MFCs. Additionally, our results indicated that EOFAZ reduced the ubiquitination degradation of PPARγ, and the chemical composition of EOFAZ directly bound to the PPARγ protein, thereby increasing its stability. Finally, PPARγ knockdown mitigated the protective effects of EOFAZ on AS in HFD ApoE−/− mice. ConclusionThese findings represent the first confirmation of EOFAZ's in vivo anti-atherosclerotic effects in ApoE−/− mice. Mechanistically, its chemical constituents can directly bind to PPARγ protein, enhancing its stability, while reducing PPARγ ubiquitination degradation, thereby inhibiting foam cell formation via activation of the PPARγ-LXRα-ABCA1/G1 pathway. Simultaneously, EOFAZ could ameliorates blood lipid metabolism and inflammatory microenvironment, thus synergistically exerting its anti-atherosclerotic effects.

Deletion of Elovl5 leads to dyslipidemia and atherosclerosis in LDLR-deficient mice

Atherosclerosis is a chronic inflammatory disease for which hepatic steatosis and atherogenic dyslipidemia are significant risk factors. We investigated the effects of endogenously generated very-long-chain polyunsaturated fatty acids (VL-PUFAs) on dyslipidemia and atherosclerosis development using mice that lack ELOVL5, a PUFA elongase that is required for the synthesis of arachidonic acid, EPA, and DHA from the essential fatty acids linoleic and linolenic acids, and the LDL receptor (LDLR). Elovl5−/−;Ldlr−/− mice manifest increased liver triglyceride and cholesterol concentrations due to the activation of sterol regulatory element binding protein-1, a transcription factor that activates enzymes required for de novo lipogenesis. Plasma levels of triglycerides and cholesterol in VLDL, IDL, and LDL were markedly elevated in Elovl5−/−;Ldlr−/− mice fed a chow and the mice exhibited marked aortic atherosclerotic plaques. Bone marrow-derived monocytes from wild-type (WT) and Elovl5−/− mice were polarized to M1 and M2 macrophages, and the effects of ELOVL5 on inflammatory activity were determined. There were no differences in most of the markers tested for M1 and M2 polarized cells between WT and Elovl5−/− cells, except for a slight increase in PGE2 secretion in Elovl5−/− cells, likely due to elevated Cox-2 expression. These results suggest that the deletion of Elovl5 leads to hepatic steatosis and dyslipidemia, which are the major factors in severe atherosclerosis in Elovl5−/−;Ldlr−/− mice.

2-(2-Phenylethyl)chromone-enriched extract of Chinese agarwood (Aquilaria sinensis) inhibits atherosclerosis progression through endoplasmic reticulum stress-mediated CD36 expression in macrophages

Ethnopharmacological relevanceChinese agarwood, derived from the Aquilaria sinensis (Lour.) Gilg (Thymelaeaceae), has a long history of use in Traditional Chinese Medicine for the management of cardiovascular disease. However, the specific active ingredients responsible for its impact on atherosclerosis are yet to be fully understood.Aim of the study: The aim of this study is to investigate the anti-atherosclerotic effectiveness of the 2-(2-phenylethyl)chromone-enriched extract derived from Chinese agarwood (CPE) through the ER stress-mediated CD36 pathway. Materials and methodsTo assess the effectiveness of CPE, an atherosclerotic mouse model was established using ApoE−/− mice with a high-fat diet. Then we assessed the impact of CPE on lipid accumulation in THP-1 macrophages that were exposed to oxLDL. Subsequently, the effect of CPE on the expression of CD36 and markers related to ER stress was characterized. ResultsOur in vivo research confirmed that CPE effectively reduces the formation of aortic plaques in atherosclerotic ApoE−/− mice. Additionally, our in vitro study observed that CPE inhibits the uptake of oxLDL and hinders the generation of foam cells. This effect is achieved by downregulating the level of CD36 in macrophages. Furthermore, our study revealed that the increase in CD36 expression, resulting from oxLDL exposure, is governed by the activation of JNK1/2/3 signaling pathways and the initiation of ER stress. ConclusionCPE demonstrated significant efficacy to inhibit the atherosclerosis. The ER stress/P-JNK/PPARγ/CD36 signaling pathway plays critical involvement in modulating the foam cell formation in vitro and in vivo. These findings underscore the efficacy of CPE as a viable therapeutic intervention for the treatment of atherosclerosis.

Long-term pulmonary iron oxide nanoparticles exposure disrupts hepatic iron-lipid homeostasis and increases plaque vulnerability in ApoE−/− mice

Iron oxide nanoparticles (Fe3O4 NPs) have attracted great attention due to their extensive applications, which warranted their environmental concerns. Although recent advances have proposed the relevance of Fe3O4 NPs to cardiovascular disease, the intrinsic mechanisms underlying the effects of NPs remain indistinct. ApoE−/− mice were chosen as a long-term exposure model to explore the immanent association between respiratory exposure to Fe3O4 NPs and the development of cardiovascular diseases. Pulmonary exposure to 20 nm and 200 nm Fe3O4 NPS resulted in significant lung injury, and pulmonary histopathological examination displayed inflammatory cell infiltration, septal thickening and alveolar congestion. Intriguingly, liver iron deposition and variations in the hepatic lipid homeostasis were found in Fe3O4 NPs-exposed mice, eventually leading to dyslipidemia, hinting the potential cardiovascular toxicity of Fe3O4 NPs. In addition, we not only found that Fe3O4 NPs exposure increased aortic plaque area, but also increased M1 macrophages in the plaque, which yielding plaque vulnerability in ApoE−/− mice Of note, 20 nm Fe3O4 NPs showed enhanced capability on the progression of atherosclerosis than 200 nm Fe3O4 NPs. This study may propose the potential mechanism for adverse cardiovascular disease induced by Fe3O4 NPs and provide convincing evidence for the safety evaluation of Fe3O4 NPs.