Atherosclerotic Plaque Stability Research Articles

Association between circulating CTRP9 levels and coronary artery disease: a systematic review and meta-analysis.

C1q tumor necrosis factor (TNF) related proteins 9 (CTRP9) is a novel adipocytokine that has been shown to have a cardioprotective effect in coronary artery disease (CAD). However, there are conflicting results on circulating levels of CTRP9 in patients with and without CAD. This meta-analysis was conducted to investigate the association between circulating CTRP9 levels and CAD. The aim of this meta-analysis was to re-examine the relationship between circulating CTRP9 levels and CAD. We searched PubMed, Web of Science, Embase, Cochrane Library, CNKI, VIP, Wanfang Data, and CBM for relevant studies up to October 2023, and 193 articles were identified. After reading the title, abstract and full text, a total of 25 articles were included in this meta-analysis. A prespecified protocol registered at INPLASY was followed (INPLASY202450066). Due to the high heterogeneity, we performed subgroup analyses and meta-regression based on patient characteristics, complications, clinical biochemical indicators, coronary artery lesion, and CAD classification. Publication bias was assessed using Egger's linear regression tests, Begg's rank correlation tests, and funnel plots. The results showed that the patient with CAD had significantly lower circulating CTRP9 levels than the control group (Z = 3.26, P = 0.001). Subgroup analysis and meta-regression findings demonstrated that observed heterogeneity could be attributed to population distribution. Patient characteristics (year of publication, patients' age, and BMI), complications (diabetes and type 2 diabetes mellitus (T2DM)), clinical biochemical indicators, coronary artery lesion (stability of coronary atherosclerotic plaque, and the number of diseased coronary vessels), and classification of CAD were not identified as source of heterogeneity. The meta-analysis confirmed that circulating CTRP9 levels in CAD patients are significantly lower than those in patients without CAD. The association may be modified by the population distribution.

Vascular Extracellular Matrix in Atherosclerosis.

The extracellular matrix (ECM) plays a central role in the structural integrity and functionality of the cardiovascular system. Moreover, the ECM is involved in atherosclerotic plaque formation and stability. In fact, ECM remodeling affects plaque stability, cellular migration, and inflammatory responses. Collagens, fibronectin, laminin, elastin, and proteoglycans are crucial proteins during atherosclerosis development. This dynamic remodeling is driven by proteolytic enzymes such as matrix metalloproteinases (MMPs), cathepsins, and serine proteases. Exploring and investigating ECM dynamics is an important step to designing innovative therapeutic strategies targeting ECM remodeling mechanisms, thus offering significant advantages in the management of cardiovascular diseases. This review illustrates the structure and role of vascular ECM, presenting a new perspective on ECM remodeling and its potential as a therapeutic target in atherosclerosis treatments.

Neovascularization as a Leading Mechanism of Intraplaque Hemorrhage and Carotid Plaque Destabilization: A Narrative Review.

Intraplaque neovascularization (IPN) is considered a leading mechanism causing carotid plaque destabilization. We provide an objective and comprehensive summary of the biology, imaging techniques, and treatment options related to carotid IPN. Plaque neovascularization has been reported to originate mainly from the adventitial vasa vasorum as a response to hypoxia. The leakage and rupture of neovessels lead to the formation of extravasations and foci of inflammation that destabilize the plaque. Vascular endothelial growth factor and its receptors are key regulators of neoangiogenesis. Neovascularization can be analyzed by advanced computed tomography and magnetic resonance imaging. The basic tools for the ultrasound assessment of IPN are contrast-enhanced ultrasound, superb microvascular imaging, and ultrasound molecular imaging. A promising direction of research seems to be the identification of patients with advanced plaque neovascularization. A simple test assessing low-velocity flow in the IPN can detect patients at risk of stroke before they experience rupture of defective neovessels and intracerebral embolism. In addition to surgical treatment, the stabilization of carotid atherosclerotic plaque can be supported pharmacologically. Statins have the best-documented role in this respect. The ideal moment of intensified therapeutic intervention in patients with previously stable carotid plaque is its increased neovascularization. However, the time frame in which intracerebral embolization may occur is unknown, and therapeutic intervention may be too late. The formation of deficient neovessels can currently be non-invasively evaluated with ultrasound. Superb microvascular imaging may change the clinical approach for asymptomatic patients at risk of cerebral ischemia.

Association of ethanolamine desaturase TMEM189-plasmalogen axis with the stability of atherosclerotic plaque and cardiovascular events after acute coronary syndrome

Abstract Background Patients with acute coronary syndrome (ACS) are still at high risk of recurring major adverse cardiovascular events (MACE). Disorders of lipid metabolism play an important role in the pathogenesis and progression of ACS. However, whether there are lipid components that can play an early warning role on the recurrence of MACE in patients with ACS and its potential mechanism is not yet known. Purpose To explore the changes of lipid profile before recurrent MACE in ACS patients, screen lipid molecules related to recurrent MACE, and explore their effects on pathological changes of atherosclerotic plaque. Methods We selected 44 Chinese ACS patients from the ‘Peking and Rotterdam on Mission to Reduce Coronary Artery Disease’ (PRoMISS) study. Among them, patients with recurrent MACE within 1 year after first onset of ACS were defined as MACE group, and the control group was recruited through 1:1 matching. The lipid profile of patients was analyzed by liquid chromatography-mass spectrometry. The effects of the screened potential lipid biomarkers and their synthesis enzymes were evaluated by real-time PCR, Western blot, IHC staining and ELISA. The enzyme knockout mice and ApoE knockout mice were hybridized and bred to obtain double gene knockout mice to determine whether the enzyme deficiency affects atherosclerotic plaque. Results Lipomic analysis showed that in the MACE group, plasmalogens and their 12 subtypes continued to decrease three months before the MACE recurrence. The most critical ethanolamine desaturase in the synthesis pathway of plasmalogen, transmembrane protein 189 (TMEM189) was also significantly lower in the MACE group than in the control group, and the level of TMEM189 was positively correlated with plasmalogen. Analysis of single-cell RNA sequencing results in human carotid artery plaques shows that TMEM189 is enriched in endothelial cells and smooth muscle cells (VSMC). In VSMC, treatment with oxLDL reduced TMEM189 expression. Overexpression of TMEM189 significantly improved oxLDL induced reduction in collagen synthesis, expression of alpha-smooth muscle actin (alpha-SMA) and smooth muscle protein 22 (SM22). Overexpression of TMEM189 in VSMC, significantly reduced the content of plasmalogens in the cells. After 12 weeks of feeding with high-fat and high-cholesterol diet, it was found that the content of smooth muscle cells and vulnerability index in the aortic plaque in TMEM189 -/-ApoE -/- mice were significantly decreased than in ApoE -/- mice. Conclusions Before the recurrence of MACE in ACS patients, the synthesis pathway of plasmalogen was gradually damaged, and the reduction of plasmalogen was a potential marker for predicting the recurrence of MACE in ACS patients. The deficiency of TMEM189 - plasmalogen axis aggravates atherosclerosis, it can regulate the phenotype transformation of smooth muscle cells, inhibit their collagen synthesis, and thus affect plaque stability.

Inclisiran-containing low-density lipoprotein cholesterol reduction effectively stabilizes atherosclerotic plaques

Abstract Background/Introduction Near-infrared spectroscopy (NIRS) allows to quantify lipid composition of coronary artery lesions. High plaque lipid content has been associated with increased adverse cardiovascular event risk. Purpose Aim of this study was to evaluate atherosclerotic plaque lipid content reduction in association with low-density lipoprotein (LDL-C) lowering on the background of intensive hypolipidaemic therapy. Methods NIRS investigation was performed in stable coronary artery disease patients having 20-50% stenosis in the proximal or middle third of a coronary artery. Segment of interest was repeatedly evaluated after 15 months. Lipid-rich plaques (LRPs) were defined as lesions with maximum lipid-core burden index within 4 mm (maxLCBI4 mm) >250. Study participants were taking high-intensity statin (atorvastatin 40 or 80 mg and rosuvastatin 20 or 40 mg) with or without ezetimibe for a run-in period of 4-6 weeks. Afterwards, in patients not achieving LDL-C level <1.8 mmol/l, add-on inclisiran was started. Baseline LDL-C was defined as the level at the time of initiation of optimal lipid-lowering therapy. Statistical significance level of 0.05 was set. Results Data of 36 patients was analyzed. Mean baseline maxLCBI4 mm was 203.9, reduced by 39.8% (-81.1, 95%CI -129.2 to -33.0) at 15-month follow-up (P=0.003). In 15 patients LRPs were detected with mean baseline maxLCBI4 mm 363.5, which was decreased by 35.8% (-130.3, 95%CI -235.0 to -25.7) (P=0.020). Mean LDL-C level at baseline was 2.5 mmol/l among all participants, lowered by 32.0% (-0.8, 95%CI -1.1 to -0.5) after 15 months of treatment (P<0.001). 19 patients received inclisiran in addition to high-dose statin and optional ezetimibe therapy. In 24 patients achieving LDL-C target <1.8 mmol/l at 15-month follow-up, a significant maxLCBI4 mm reduction from 208.5 by 48.4% (-101.0, 95%CI -169.3 to -32.7) was established (P=0.010). Among those having LDL-C >1.8 mmol/l, maxLCBI4 mm reduction from 194.7 by 21.2% (-41.25, 95%CI -94.7 to 12.2) was observed, however the change was not statistically significant (P=0.114). Conclusion LDL-C lowering with high-intensity hypolipidaemic therapy, including escalation to proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibition, demonstrated atherosclerotic plaque stabilization by NIRS-detected lipid content decrease.

Alisol A inhibits and stabilizes atherosclerotic plaques by protecting vascular endothelial cells.

Dysfunction of endothelial cells represents a crucial aspect in the pathogenesis of atherosclerosis. The aim of this study was to explore the protective effects of alisol A on vascular endothelial cells and its possible mechanisms. An atherosclerosis model was established by feeding ApoE-/- mice with high-fat chow. Alisol A (150mg/kg/d) or atorvastatin (15mg/kg/d) was administered, and the levels of blood lipids were evaluated. The effect of the drugs on atherosclerotic plaques was observed by staining the aorta with Sudan IV. In vitro experiments were conducted using human aortic endothelial cells (HAECs) to assess the effects of alisol A on cell proliferation, migration, tubulation, secretion, and cellular integrity by CCK-8 assay, wound healing assay, angiogenesis assay, NO secretion, and release of LDH. Transcriptomics and molecular docking were used to explore the mechanism of plaque inhibition and stabilization by alisol A. Alisol A significantly reduced the aortic plaque area in ApoE-/- mice fed with high-fat chow. In vitro, alisol A had a protective effect on HAECs, which was reflected in the inhibition of vascular endothelial cell proliferation, promotion of NO secretion by vascular endothelial cells, inhibition of vascular endothelial cell migration and angiogenesis, and the maintenance of cell membrane integrity. Therefore, alisol A inhibited and stabilized atherosclerotic plaques and slowed down the process of atherosclerosis. Transcriptomics studies showed 4,086 differentially expressed genes (DEGs) in vascular endothelial cells after alisol A treatment. Enrichment analysis indicated that many genes involved in TNF signaling pathway were differentially expressed, and inflammatory genes were suppressed. The molecular docking results verified the hypothesis that alisol A has a low binding energy after docking with TNF target, and TNF could be a potential target of alisol A. Alisol A produced protection on vascular endothelial cells, achieving inhibition and stabilization of atherosclerotic plaques.

HUC-MSCs mitigate atherosclerosis induced by a high-fat diet in ApoE−/− mice by regulating the intestinal microbiota

BackgroundThe mechanism underlying human umbilical cord mesenchymal stem cells (hUC-MSCs) regulating the stability of atherosclerotic plaque was explored by establishing mice models of atherosclerosis induced by a high-fat diet and hUC-MSCs intervention. MethodsThe ApoE−/− mice atherosclerosis model was constructed using a high-fat diet, and mice were divided into a normal diet group (ND), high-fat diet group (HFD), hUC-MSCs treatment group (HFDM), while the blank control (BC) consisted of C57BL/6J mice. After successful establishment of the model, the feces, hearts, and aorta of mice were collected. Morphological features were detected using HE, oil red O, and Masson staining. Afterward, 16s rRNA gene sequences was used to detect the species and abundance of the intestinal flora in mice, and an atomic force microscope (AFM) was used to detect the Young's modulus of the fibrous cap of atherosclerotic plaques. Lastly, the expression level of the inflammatory factors NLRP3, IL-1β, and IL-18 were detected via immunohistochemistry and immunofluorescence assays. ResultsIn terms of morphological characteristics, the expression level of NLRP3, Young's modulus of the fibrous cap, and plaque stability were significantly reduced in HFD, whereas the ratio of Firmicutes to Bacteroidetes (F/B) was significantly increased. Interestingly, hUC-MSCs treatment reversed the above indices, thus enhancing plaque stability. ConclusionHFD led to dysregulation of intestinal flora homeostasis and induced aberrant expression levels of NLRP3, resulting in a decrease in the Young's modulus of plaques. However, hUC-MSCs treatment improved the biomechanical properties of plaque by modulating the intestinal flora and NLRP3, thereby elevating plaque stability and minimizing the risk of plaque rupture.

Taurine Reduces Atherosclerotic Plaque Area and Stability in Mice

Taurine Reduces Atherosclerotic Plaque Area and Stability in Mice

ECM Microenvironment in Vascular Homeostasis: New Targets for Atherosclerosis.

Alterations in vascular extracellular matrix (ECM) components, interactions, and mechanical properties influence both the formation and stability of atherosclerotic plaques. This review discusses the contribution of the ECM microenvironment in vascular homeostasis and remodeling in atherosclerosis, highlighting Cartilage oligomeric matrix protein (COMP) and its degrading enzyme ADAMTS7 as examples, and proposes potential avenues for future research aimed at identifying novel therapeutic targets for atherosclerosis based on the ECM microenvironment.

HCC-1 Accelerates Atherosclerosis by Inducing Endothelial Cell and Macrophage Pyroptosis and Serves as an Early Diagnostic Biomarker.

HCC-1 (hemofiltrate CC chemokine-1), a CC-type chemokine, exerts function to change intracellular calcium concentration, induce leukocyte, and manipulate enzyme release especially in monocytes. It has been reported that HCC-1 can predict the persistent acute kidney injury or suppress hepatocellular carcinoma by modulating cell cycle and promoting apoptosis; however, the effect of HCC-1 on atherosclerosis is poorly understood. Here, we aimed to clarify the function and mechanism of HCC-1 in atherosclerosis and whether it could serve as a novel biomarker for the diagnosis of atherosclerosis. HCC-1 expression in serum, atherosclerotic plaques, and normal arterial tissue from patients with atherosclerosis and control group was assessed by ELISA, immunohistochemistry and confocal microscope, and bioinformatic analysis. The atherosclerotic model of HCC-1 overexpressing and control mice was generated by tail vein injection of adeno-associated virus serotype 9-HCC-1 on an ApoE-/- background. Cell adhesion, polarization, and pyroptosis were evaluated in vitro. The relationship between HCC-1 concentration in serum and atherosclerosis was analyzed in patients with atherosclerosis. HCC-1 expression was positively correlated with the occurrence and stable-unstable switch of atherosclerosis under bioinformatic analysis, which is further supported by the results of increased HCC-1 expression in atherosclerosis patients both in serum and atherosclerotic plaque. adeno-associated virus serotype 9-HCC-1 mice had higher levels of inflammatory factors, increased macrophage accumulation and pyroptotic rate in plaque, and decreased atherosclerotic plaque stability. In vitro, HCC-1 promoted monocyte adhesion and M1 polarization and induced inflammation and pyroptosis both in endothelial cells and macrophages. HCC-1 expression was increased in patients with atherosclerosis, and HCC-1 overexpression accelerated atherosclerotic burden via an enhancement in monocyte recruitment, M1 polarization, and pyroptosis both in endothelial cells and macrophages. Our findings suggested that HCC-1 may serve as an early biomarker for the diagnosis of atherosclerosis, with the capacity to reflect the degree of stenosis.

A novel mechanism of ferroptosis inhibition-enhanced atherosclerotic plaque stability: YAP1 suppresses vascular smooth muscle cell ferroptosis through GLS1.

Atherosclerosis is a leading cause of cardiovascular diseases (CVDs), often resulting in major adverse cardiovascular events (MACEs), such as myocardial infarction and stroke due to the rupture or erosion of vulnerable plaques. Ferroptosis, an iron-dependent form of cell death, has been implicated in the development of atherosclerosis. Despite its involvement in CVDs, the specific role of ferroptosis in atherosclerotic plaque stability remains unclear. In this study, we confirmed the presence of ferroptosis in unstable atherosclerotic plaques and demonstrated that the ferroptosis inhibitor ferrostatin-1 (Fer-1) stabilizes atherosclerotic plaques in apolipoprotein E knockout (Apoe-/-) mice. Using bioinformatic analysis combining RNA sequencing (RNA-seq) with single-cell RNA sequencing (scRNA-seq), we identified Yes-associated protein 1 (YAP1) as a potential key regulator of ferroptosis in vascular smooth muscle cells (VSMCs) of unstable plaques. Invitro, we found that YAP1 protects against oxidized low-density lipoprotein (oxLDL)-induced ferroptosis in VSMCs. Mechanistically, YAP1 exerts its anti-ferroptosis effects by regulating the expression of glutaminase 1 (GLS1) to promote the synthesis of glutamate (Glu) and glutathione (GSH). These findings establish a novel mechanism where the inhibition of ferroptosis promotes the stabilization of atherosclerotic plaques through the YAP1/GLS1 axis, attenuating VSMC ferroptosis. Thus, targeting the YAP1/GLS1 axis to suppress VSMC ferroptosis may represent a novel strategy for preventing and treating unstable atherosclerotic plaques.

Abstract Tu015: Angiopoietin-like 4 Stabilizes Atherosclerotic Plaques by Modulating the Phenotypic Transition of Endothelial Cells and Vascular Smooth Muscle Cells

Atherosclerosis, the leading cause of death, is a vascular disease of chronic inflammation. We recently showed that angiopoietin-like 4 (ANGPTL4) promotes cardiac repair by suppressing pathological inflammation. Given the fundamental contribution of inflammation to atherosclerosis, we assessed the role of ANGPTL4 in the development of atherosclerosis and determined whether ANGPTL4 regulates atherosclerotic plaque stability. We injected ANGPTL4 protein twice a week into atherosclerotic Apoe-/- mice and analyzed the atherosclerotic lesion size, endothelial dysfunction, inflammation, and plaque stability. In atherosclerotic mice, ANGPTL4 reduced the atherosclerotic plaque size, endothelial dysfunction, and vascular inflammation. Furthermore, ANGPTL4 was associated with maintenance of vascular integrity and attenuated endothelial-to-mesenchymal transition (EndMT) in both HUVEC and atherosclerotic plaque of Apoe-/- mice. In the atherosclerotic lesions and fibrous caps, the number of α-SMA(+), SM22α(+), and SM-MHC(+) cells was higher, while the number of CD68(+) and Mac2(+) cells was lower in the ANGPTL4 group. Most importantly, the fibrous cap was significantly thicker in the ANGPTL4 group than in the PBS group. Smooth muscle cells (SMCs) isolated from atherosclerotic aortas showed significantly increased expression of CD68 and Krüppel-like factor 4 (KLF4), a modulator of the vascular SMC phenotype, along with downregulation of α-SMA, and these changes were attenuated by ANGPTL4 treatment. Furthermore, ANGPTL4 reduced TNFa-induced NADPH oxidase 1 (NOX1), a major source of reactive oxygen species, resulting in attenuation of KLF4-mediated SMC phenotypic changes. Our results reveal that ANGPTL4 treatment inhibits atherogenesis and suggest that targeting vascular stability and inflammation may serve as a novel therapeutic strategy to prevent and treat atherosclerosis. Even more importantly, ANGPTL4 treatment inhibited the phenotypic changes of SMCs into macrophage-like cells by downregulating NOX1 activation of KLF4, leading to the formation of more stable plaques.

CD8+ T Cells Drive Plaque Smooth Muscle Cell Dedifferentiation in Experimental Atherosclerosis.

Atherosclerosis is driven by the infiltration of the arterial intima by diverse immune cells and smooth muscle cells (SMCs). CD8+ T cells promote lesion growth during atherosclerotic lesion development, but their role in advanced atherosclerosis is less clear. Here, we studied the role of CD8+ T cells and their effects on SMCs in established atherosclerosis. CD8+ T cells were depleted in (SMC reporter) low-density lipoprotein receptor-deficient (Ldlr-/-) mice with established atherosclerotic lesions. Atherosclerotic lesion formation was examined, and single-cell RNA sequencing of aortic SMCs and their progeny was performed. Additionally, coculture experiments with primary aortic SMCs and CD8+ T cells were conducted. Although we could not detect differences in atherosclerotic lesion size, an increased plaque SMC content was noted in mice after CD8+ T-cell depletion. Single-cell RNA sequencing of aortic lineage-traced SMCs revealed contractile SMCs and a modulated SMC cluster, expressing macrophage- and osteoblast-related genes. CD8+ T-cell depletion was associated with an increased contractile but decreased macrophage and osteoblast-like gene signature in this modulated aortic SMC cluster. Conversely, exposure of isolated aortic SMCs to activated CD8+ T cells decreased the expression of genes indicative of a contractile SMC phenotype and induced a macrophage and osteoblast-like cell state. Notably, CD8+ T cells triggered calcium deposits in SMCs under osteogenic conditions. Mechanistically, we identified transcription factors highly expressed in modulated SMCs, including Runx1, to be induced by CD8+ T cells in cultured SMCs in an IFNγ (interferon-γ)-dependent manner. We here uncovered CD8+ T cells to control the SMC phenotype in atherosclerosis. CD8+ T cells promote SMC dedifferentiation and drive SMCs to adopt features of macrophage-like and osteoblast-like, procalcifying cell phenotypes. Given the critical role of SMCs in atherosclerotic plaque stability, CD8+ T cells could thus be explored as therapeutic target cells during lesion progression.

Flow shear force destabilizes carotid plaques by affecting CHOP and GRP78 proteins

BackgroundVarious factors, including blood, inflammatory, infectious, and immune factors, can cause ischemic stroke. However, the primary cause is often the instability of cervical arteriosclerosis plaque. It is estimated that 18-25% of ischemic strokes are caused by the rupture of carotid plaque.1 Plaque stability is crucial in determining patient prognosis. Developing a highly accurate, non-invasive, or minimally invasive technique to assess carotid plaque stability is crucial for diagnosing and treating stroke.Previous research by our group has demonstrated that the expression levels of CHOP (C/EBP homologous protein) and GRP78 (glucose-regulated protein 78) are correlated with the stability of atherosclerotic plaques.2 ObjectThis research assesses changes in GRP78 and CHOP expressions in human umbilical vein endothelial cells(HUVEC) following experiments within the hemodynamic influencing factors test system. Additionally, it includes conducting an empirical study on the impact of blood flow shear force on the stability of human carotid atherosclerotic plaques. The objective is to explore the implications of blood flow shear force on the stability of carotid atherosclerotic plaques. MethodThe hemodynamic influencing factors test bench system was configured with low (Group A, 4 dyns/cm²), medium (Group B, 8 dyns/cm²), and high shear force groups (Group C, 12 dyns/cm²). Relative expression levels of GRP78 and CHOP proteins in human umbilical vein endothelial cells were measured using Western blot analysis, and quantitative analysis of GRP78 and CHOP mRNA was conducted using RT-qPCR. Meanwhile, plaques from 60 carotid artery patients, retrieved via Carotid Endarterectomy (CEA), were classified into stable (S) and unstable (U) groups based on pathological criteria. Shear force at the carotid bifurcation was measured preoperatively using ultrasound. Western blot and RT-qPCR were used to analyze the relative expression levels of GRP78 and CHOP proteins and mRNA, respectively, in the plaque specimens from both groups. ResultExpression levels of GRP78, CHOP proteins, and their mRNAs were assessed in groups A, B, and C via Western blot and RT-qPCR. Results showed that in the low-shear group, all markers were elevated in group A compared to groups B and C. Statistical analysis revealed significantly lower shear forces at the carotid bifurcation in group U compared to group S. In group U plaques, GRP78 and CHOP expressions were significantly higher in group U than in group S. ConclusionBlood flow shear forces variably affect the expression of GRP78 and CHOP proteins, as well as their mRNA levels, in vascular endothelial cells. The lower the shear force and fluid flow rate, the higher the expression of GRP78 and CHOP, potentially leading to endoplasmic reticulum stress(ERS), which may destabilize the plaque.

Melatonin Ameliorates Atherosclerotic Plaque Vulnerability by Regulating PPARδ-Associated Smooth Muscle Cell Phenotypic Switching.

Vulnerable atherosclerotic plaque rupture, the leading cause of fatal atherothrombotic events, is associated with an increased risk of mortality worldwide. Peroxisome proliferator-activated receptor delta (PPARδ) has been shown to modulate vascular smooth muscle cell (SMC) phenotypic switching, and, hence, atherosclerotic plaque stability. Melatonin reportedly plays a beneficial role in cardiovascular diseases; however, the mechanisms underlying improvements in atherosclerotic plaque vulnerability remain unknown. In this study, we assessed the role of melatonin in regulating SMC phenotypic switching and its consequential contribution to the amelioration of atherosclerotic plaque vulnerability and explored the mechanisms underlying this process. We analyzed features of atherosclerotic plaque vulnerability and markers of SMC phenotypic transition in high-cholesterol diet (HCD)-fed apolipoprotein E knockout (ApoE-/-) mice and human aortic SMCs (HASMCs). Melatonin reduced atherosclerotic plaque size and necrotic core area while enhancing collagen content, fibrous cap thickness, and smooth muscle alpha-actin positive cell coverage on the plaque cap, which are all known phenotypic characteristics of vulnerable plaques. In atherosclerotic lesions, melatonin significantly decreased the synthetic SMC phenotype and KLF4 expression and increased the expression of PPARδ, but not PPARα and PPARγ, in HCD-fed ApoE-/- mice. These results were subsequently confirmed in the melatonin-treated HASMCs. Further analysis using PPARδ silencing and immunoprecipitation assays revealed that PPARδ plays a role in the melatonin-induced SMC phenotype switching from synthetic to contractile. Collectively, we provided the first evidence that melatonin mediates its protective effect against plaque destabilization by enhancing PPARδ-mediated SMC phenotypic switching, thereby indicating the potential of melatonin in treating atherosclerosis.

Application of adipose tissue-derived stem cell therapy with a clinically relevant dose does not significantly affect atherosclerotic plaque characteristics in a streptozotocin-induced hyperglycaemia mouse model

AimsDiabetes mellitus (DM) induces increased inflammation of atherosclerotic plaques, resulting in elevated plaque instability. Mesenchymal stem cell (MSC) therapy was shown to decrease plaque size and increase stability in non-DM animal models. We now studied the effect of MSC therapy in a streptozotocin-induced hyperglycaemia mouse model using a clinically relevant dose of adipose tissue-derived MSCs (ASCs). MethodsHyperglycaemia was induced in male C57BL/6 ApoE−/− mice (n=24) via intraperitoneal streptozotocin (STZ) injection (0.05 mg/g bodyweight) for 5 consecutive days. 16 weeks after the first STZ injection, the mice received either 100,000 ASCs (n=9) or vehicle (n=14) intravenously. The effects of ASC treatment on the size and stability of aortic root atherosclerotic plaques were determined 4 weeks post-treatment via (immuno)histochemical analyses. Furthermore, plasma monocyte subsets within 3 days pre- and 3 days post-treatment, and 4 weeks post-treatment, were studied. ResultsASC treatment did not significantly affect atherosclerotic plaque size or intra-plaque inflammation. Although ASC-treated mice had a higher percentage of intra-plaque fibrosis (42.5±3.3%) compared to vehicle-treated mice (37.6±6.8%, p=0.07), this did not reach significance. Additionally, although differences in the percentages of circulating pro- and anti-inflammatory monocytes were observed after ASC treatment compared to pre-treatment (p=0.005), their levels did not differ significantly at any time point compared to vehicle-treated mice. ConclusionsASC treatment with a clinically relevant dose did not significantly affect atherosclerotic plaque size or intra-plaque inflammation in a hyperglycaemia mouse model. Despite a borderline significant improvement in intraplaque fibrotic content, the potential of ASC treatment on atherosclerotic plaque stability in a diabetic environment remains to be determined.

20( S )-Protopanaxatriol Improves Atherosclerosis by Inhibiting Low-Density Lipoprotein Receptor Degradation in ApoE KO Mice.

Atherosclerosis (AS) is a chronic progressive disease caused by various factors and causes various cerebrovascular and cardiovascular diseases (CVDs). Reducing the plasma levels of low-density lipoprotein cholesterol is the primary goal in preventing and treating AS. Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in regulating low-density lipoprotein cholesterol metabolism. Panax notoginseng has potent lipid-reducing effects and protects against CVDs, and its saponins induce vascular dilatation, inhibit thrombus formation, and are used in treating CVDs. However, the anti-AS effect of the secondary metabolite, 20( S )-protopanaxatriol (20( S )-PPT), remains unclear. In this study, the anti-AS effect and molecular mechanism of 20( S )-PPT were investigated in vivo and in vitro by Western blotting, real-time polymerase chain reaction, enzyme-linked immunosorbent assay, immunofluorescence staining, and other assays. The in vitro experiments revealed that 20( S )-PPT reduced the levels of PCSK9 in the supernatant of HepG2 cells, upregulated low-density lipoprotein receptor protein levels, promoted low-density lipoprotein uptake by HepG2 cells, and reduced PCSK9 mRNA transcription by upregulating the levels of forkhead box O3 protein and mRNA and decreasing the levels of HNF1α and SREBP2 protein and mRNA. The in vivo experiments revealed that 20( S )-PPT upregulated aortic α-smooth muscle actin expression, increased the stability of atherosclerotic plaques, and reduced aortic plaque formation induced by a high-cholesterol diet in ApoE -/- mice (high-cholesterol diet-fed group). Additionally, 20( S )-PPT reduced the aortic expression of CD68, reduced inflammation in the aortic root, and alleviated the hepatic lesions in the high-cholesterol diet-fed group. The study revealed that 20( S )-PPT inhibited low-density lipoprotein receptor degradation via PCSK9 to alleviate AS.

Urolithin A promotes atherosclerotic plaque stability by limiting inflammation and hypercholesteremia in Apolipoprotein E-deficient mice.

Urolithin A (UroA), a dietary phytochemical, is produced by gut bacteria from fruits rich in natural polyphenols ellagitannins (ETs). The efficiency of ETs metabolism to UroA in humans depends on gut microbiota. UroA has shown a variety of pharmacological activities. In this study we investigated the effects of UroA on atherosclerotic lesion development and stability. Apolipoprotein E-deficient (ApoE-/-) mice were fed a high-fat and high-cholesterol diet for 3 months to establish atherosclerosis model. Meanwhile the mice were administered UroA (50 mg·kg-1·d-1, i.g.). We showed that UroA administration significantly decreased diet-induced atherosclerotic lesions in brachiocephalic arteries, macrophage content in plaques, expression of endothelial adhesion molecules, intraplaque hemorrhage and size of necrotic core, while increased the expression of smooth muscle actin and the thickness of fibrous cap, implying features of plaque stabilization. The underlying mechanisms were elucidated using TNF-α-stimulated human endothelial cells. Pretreatment with UroA (10, 25, 50 μM) dose-dependently inhibited TNF-α-induced endothelial cell activation and monocyte adhesion. However, the anti-inflammatory effects of UroA in TNF-α-stimulated human umbilicalvein endothelial cells (HUVECs) were independent of NF-κB p65 pathway. We conducted RNA-sequencing profiling analysis to identify the differential expression of genes (DEGs) associated with vascular function, inflammatory responses, cell adhesion and thrombosis in UroA-pretreated HUVECs. Human disease enrichment analysis revealed that the DEGs were significantly correlated with cardiovascular diseases. We demonstrated that UroA pretreatment mitigated endothelial inflammation by promoting NO production and decreasing YAP/TAZ protein expression and TEAD transcriptional activity in TNF-α-stimulated HUVECs. On the other hand, we found that UroA administration modulated the transcription and cleavage of lipogenic transcription factors SREBP1/2 in the liver to ameliorate cholesterol metabolism in ApoE-/- mice. This study provides an experimental basis for new dietary therapeutic option to prevent atherosclerosis.

LncRNA CDKN2B-AS1 regulates collagen expression

The long noncoding RNA CDKN2B-AS1 harbors a major coronary artery disease risk haplotype, which is also associated with progressive forms of the oral inflammatory disease periodontitis as well as myocardial infarction (MI). Despite extensive research, there is currently no broad consensus on the function of CDKN2B-AS1 that would explain a common molecular role of this lncRNA in these diseases. Our aim was to investigate the role of CDKN2B-AS1 in gingival cells to better understand the molecular mechanisms underlying the increased risk of progressive periodontitis. We downregulated CDKN2B-AS1 transcript levels in primary gingival fibroblasts with LNA GapmeRs. Following RNA-sequencing, we performed differential expression, gene set enrichment analyses and Western Blotting. Putative causal alleles were searched by analyzing associated DNA sequence variants for changes of predicted transcription factor binding sites. We functionally characterized putative functional alleles using luciferase-reporter and antibody electrophoretic mobility shift assays in gingival fibroblasts and HeLa cells. Of all gene sets analysed, collagen biosynthesis was most significantly upregulated (Padj=9.7 × 10− 5 (AUC > 0.65) with the CAD and MI risk gene COL4A1 showing strongest upregulation of the enriched gene sets (Fold change = 12.13, Padj = 4.9 × 10− 25). The inflammatory “TNFA signaling via NFKB” gene set was downregulated the most (Padj=1 × 10− 5 (AUC = 0.60). On the single gene level, CAPNS2, involved in extracellular matrix organization, was the top upregulated protein coding gene (Fold change = 48.5, P < 9 × 10− 24). The risk variant rs10757278 altered a binding site of the pathogen responsive transcription factor STAT1 (P = 5.8 × 10− 6). rs10757278-G allele reduced STAT1 binding 14.4% and rs10757278-A decreased luciferase activity in gingival fibroblasts 41.2% (P = 0.0056), corresponding with GTEx data. CDKN2B-AS1 represses collagen gene expression in gingival fibroblasts. Dysregulated collagen biosynthesis through allele-specific CDKN2B-AS1 expression in response to inflammatory factors may affect collagen synthesis, and in consequence tissue barrier and atherosclerotic plaque stability.

Short-term glycemic variability and intracranial atherosclerotic plaque stability assessed by high-resolution MR vessel wall imaging in type 2 diabetes mellitus

ObjectiveTo investigate the relationship between short-term glycemic variability in patients with T2DM and the vulnerability of intracranial atherosclerotic plaques using HR-MR-VWI. Materials and methodsIn total, 203 patients with acute ischemic stroke (AIS)/transient ischemia (TIA) combined with T2DM were enrolled. All of them underwent HR-MR-VWI during the period between July 2020 and July 2023. 203 patients were divided into groups with higher (1,5-AG≤ 30.7 μmol/L) and lower (1,5-AG> 30.7 μmol/L) short-term glycemic variability. Patients were also divided into the T1WI and non-T1WI hyperintensity groups. Associated factors(FBG, HbA1c, and 1,5-AG)for the T1WI hyperintensity were analyzed by binary logistic regression. We used the area under the curve (AUC), while the sensitivity and specificity were calculated at the optimal threshold. The Delong test was employed to compare the quality of the AUC of the predictors. ResultsThe group with higher short-term glycemic variability had a higher incidence of the hyperintensity on T1WI, higher degree of enhancement, higher degree of stenosis and smaller lumen area (P < 0.05). The T1WI hyperintensity group had higher HbA1c levels, higher hemoglobin levels and lower 1,5-AG levels(P < 0.05). 1,5-AG (OR = 0.971, 95 % CI: 0.954∼0.988, P = 0.001), HbA1c (OR=1.305, 95 % CI: 1.065∼1.598, P = 0.01) and male sex (OR = 2.048, 95 % CI: 1.016∼4.128, P = 0.045)/(OR=2.102, 95 % CI: 1.058∼4.177, P = 0.034) were independent risk factors for the hyperintensity on T1WI. 1,5-AG demonstrated enhanced performance and yielded the highest AUC of the receiver operator characteristic curve (AUC = 0.726), with sensitivity and specificity values of 0.727 and 0.635 respectively. Conclusion1,5-AG, HbA1c and male sex are independent predictors of intracranial plaques with T1WI hyperintensity, the greater short-term glycemic variability, the higher incidence of vulnerable plaques.