Atherosclerotic Plaque Stability Research Articles

MiR-449a disturbs atherosclerotic plaque stability in streptozotocin and high-fat diet-induced diabetic mice by targeting CEACAM1

BackgroundEmerging evidence indicates carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is involved in the development of atherosclerosis (AS). However, the roles and functions of CEACAM1 in AS remain unknown. Therefore, this study aims to investigate the roles and molecular functions of CEACAM1 in AS.MethodsWe constructed a diabetes mellitus (DM) + high-fat diet (HFD) mouse model based on the streptozotocin (STZ)-induced apolipoprotein E-knockdown (ApopE−/−) mouse to investigate the roles and regulatory mechanism of miR-449a/CEACAM1 axis. The mRNA expression and protein levels in this study were examined using quantity PCR, western blot, immunofluorescence (IF), enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry (IHC), respectively. And the lipid deposition and collagen content were detected using Oil Red O and Sirius Red staining. Cell apoptosis, migration, invasion, and tuber formation were detected by Annexin-V FITC/PI, wound healing, transwell, and tuber formation assays, respectively. The relationship between miR-449a and CEACAM1 was determined by a dual-luciferase reporter gene assay.ResultsmiR-449a and MMP-9 were upregulated, and CEACAM1 was downregulated in the DM + HFD MOUSE model. Upregulation of CEACAM1 promoted atherosclerotic plaque stability and inhibited inflammation in the DM + HFD mouse model. And miR-449a directly targeted CEACAM1. Besides, miR-449a interacted with CEACAM1 to regulate atherosclerotic plaque stability and inflammation in DM-associated AS mice. In vitro, the rescue experiments showed miR-449a interacted with CEACAM1 to affect apoptosis, migration, invasion, and tuber formation ability in high glucose (HG)-induced HUVECs.ConclusionThese results demonstrated that miR-449a promoted plaque instability and inflammation in DM and HFD-induced mice by targeting CEACAM1.

Abstract 3131: An Investigation Into The Interplay Between Epha2 And Inflammatory Responses In Vascular Smooth Muscle Cells

Cardiovascular disease continues to be a leading cause of death in the United States. A major underlying cause of cardiovascular events such as stroke and myocardial infarction is atherosclerosis. Notable mediators in the pathogenesis of atherosclerosis include vascular smooth muscle cells (VSMCs) and inflammation. During atherogenesis, the SMCs undergo phenotypic change from a quiescent state to a synthetic, proliferative state. The synthetic SMCs drive neointima formation which influences the stenosis characteristic of atherosclerosis. Signaling of a receptor tyrosine kinase called EphA2 has been previously determined to have a role in SMC phenotypic change. It was found that a global and SMC specific knockdown (KD) of EphA2 resulted in decreased plaque formation, suggesting the role of EphA2 expression in promoting SMC proliferation. Inflammation is another important factor that contributes to plaque formation. Oxidized low-density lipoproteins (LDLs) have been found to also cause VSMCs to undergo change to a synthetic phenotype, one that resembles macrophages. Cytokines released by macrophages orchestrate the pro-inflammatory response that affects the development and stability of atherosclerotic plaques. Considering the important role inflammation also serves in the progression of plaque formation, we investigated the relationship between EphA2 and VSMC-mediated inflammatory response. Cre-flox breeding was performed to achieve EphA2 knockout (KO) specifically in VSMCs of 8-10-wk old ApoE -/- mice. Successful KO was confirmed with PCR and Western blot. Si-RNA was transfected to achieve KD in human SMCs. The cells were then treated with cytokines, including IL-1 beta, TNF-alpha, IL-6, IL-33, and interferon-gamma, followed by RT-qPCR analysis to measure expression of inflammatory genes. To assess cytokine secretion, we utilized a cytokine array to measure presence of the cytokines of interest. Protein analysis to quantify expression of inflammatory proteins in KO vs control cells was done via Western blot. We observed that the loss of EphA2 causes significant reductions in the production of several proinflammatory cytokines and chemokines, indicating its role in mediating VSMC inflammatory responses.

Abstract 141: Lesion Site-specific Mass Spectrometry Analysis Of Human Carotid Plaques Reveals Strongest Association Of The Necrotic Core And Fibrous Cap Proteome With Plaque Stability

The spatial impact of the plaque proteome on atherosclerotic plaque stability remains largely unknown. In this study, we performed mass spectrometry-based proteomics of various carotid plaque subregions (media, necrotic core, fibrous cap), using guided laser capture micro-dissections (LCM) of FFPE tissue in a matched cohort of 112 patients undergoing carotid endarterectomy (CEA) to study the impact of the plaque proteome, not only on lesion stability, but also on subregional changes of expression. Further, we performed a histological analysis according to the AHA classification and combined the data with clinical information on risk factors, comorbidities, and patient symptoms. Our LCM mass spectrometry analysis yielded over 3000 proteins for each plaque subregion. Gene set enrichment analysis revealed a high abundance of extracellular proteins and vice versa for cellular proteins. Subregion analysis identified the necrotic core and the fibrous cap to display the most significant changes in proteins associated with the stability status, whereas the media proteome remained largely unaffected. Primary and secondary enrichment analysis revealed synthetic vascular smooth muscle cells as the major source of these proteins. In addition, we analyzed the serum proteome of more than 500 patients (including non-athero control patients) with overlap from the tissue cohort. We combined these with histological evaluation and a complete set of clinical data. We used these two datasets for developing tissue and serological markers associated with plaque stability. In summary, our approach utilized LCM mass spectrometry, histological evaluation, and clinical data to analyze the association of different lesion subregions on plaque stability. We are able to demonstrate a high through-put method to identify potential protein markers relevant to atherosclerotic plaque stability.

Abstract 1055: Atxn2 Inhibition Enhances The Contractile Phenotype Of Vascular Smooth Muscle Cells

Introduction: Phenotype switching of vascular smooth muscle cells (VSMC) between their synthetic and differentiated contractile state is a critical player in controlling the stability of atherosclerotic plaque. Hundreds of genes associated with coronary artery disease (CAD) have been identified through Genome-Wide Association Studies (GWAS). However, the involvement of most identified genes in the pathogenesis of atherosclerosis remains unknown. ATXN2 was identified as a CAD GWAS candidate gene with unclear biological role in atherosclerosis. ATXN2 regulates the endocytic internalization of the EGF receptor, which is essential in the VSMC phenotype switching. Therefore, we aimed to investigate whether ATXN2 is involved in the VSMC phenotype switching. Methods and Results: Using immunofluorescence, we demonstrated that ATXN2 siRNA knockdown resulted in a 5-fold increase in the ratio of the contractile phenotype of human aortic VSMC that expressed a high alpha-smooth muscle cell actin content compared to controls. In addition, we found that ATXN2 inhibition reduced the VSMC proliferation and migration as assessed by the EdU cell proliferation assay and the wound healing migration assay, respectively. Next, we confirmed that ATXN2 siRNA knockdown increased the mRNA expression of contractile markers: calponin, alpha-smooth muscle cell actin, and smooth muscle protein 22 alpha using quantitative PCR. We found that the enhancing effect of VSMC contractile phenotype by ATXN2 siRNA was less pronounced in the absence of TGF-beta. Indeed, the enhancing effect of ATXN2 inhibition on the VSMC contractile phenotype was abolished by SMAD4 gene silencing, suggesting the involvement of TGF-beta signaling. Furthermore, we validated the expression signature of the ATXN2 gene in different phenotypic subsets of VSMC from human atherosclerotic plaque samples using single-cell RNA sequencing analysis. We detected that ATXN2 expression was negatively correlated with the contractile VSMC subset. In conclusion, our findings suggests ATXN2, CAD GWAS candidate gene, as a novel potential therapeutic target to enhance the VSMC contractile phenotype for stabilizing atherosclerotic plaque in coronary artery disease patients.

Abstract 1038: Map Kinase Phosphatase-5 Deficiency Impairs Macrophage Efferocytosis And Exacerbates Late-stage Atherosclerosis

Objective: Atherosclerosis is a vascular inflammatory disease that is a major cause of death in Westernized society. Late-stage atherosclerosis is characterized by a vulnerable plaque, which consists of a thin fibrous cap and a large necrotic core that is prone to rupture. Efferocytosis is a repair mechanism that clears apoptotic cells within the plaque, contributing to plaque stability. In late-stage atherosclerotic conditions efferocytosis is impaired and promotes rupture. MAPK phosphatase-5 (MKP5) regulates various processes such as immune diseases and cardiac fibrosis. The goal of this project is to identify the role of MKP5 in late-stage atherosclerotic plaque stability. Approach/Results: To induce late-stage atherosclerosis, male and female LDLR -/- and LDLR -/- MKP5 -/- mice were subjected to transverse aortic constriction (TAC) surgery and fed on a highfat diet (western diet with 1.25% cholesterol) for 12 weeks. Our data show that MKP5 deficiency exacerbates plaque instability, through decreased cap thickness, decreased collagen content, increased lesion size and necrosis in the plaque, and severe coronary lesions. Furthermore, there are higher instances of intraplaque hemorrhage in the right carotids of the LDLR -/- MKP5 -/- mice. Mechanistically, TUNEL staining of the aortic roots was performed to determine efferocytotic capabilities in vivo. Results showed fewer macrophage-associated TUNEL-positive cells in LDLR -/- MKP5 -/- aortic roots. To further investigate this mechanism, bone-marrow-derived macrophages (BMDMs) isolated from WT and MKP5 -/- mice were incubated with apoptotic Jurkat cells and then analyzed to determine the engulfment of apoptotic cells by the macrophages via confocal microscopy and flow cytometry. Results showed that efferocytosis was significantly inhibited in MKP5 -/- BMDMs. Bulk-RNA sequencing analysis revealed that the expression of efferocytotic genes in MKP5 -/- BMDMs was decreased at basal and oxLDL stimulated conditions. Conclusion: Results suggest that MKP5 deficiency impairs efferocytosis, exacerbates atherosclerosis, and causes late-stage plaque to move toward unstable phenotypes.

Abstract 127: Pushing The Limits Of Macrophage Efferocytosis In Atherosclerosis

Introduction: Enhancing macrophage efferocytosis represents a potential therapeutic strategy for residual risk reduction in atherosclerotic cardiovascular disease. Our genome-wide CRISPR screening discovered Pdcd6ip as a top-ranked negative regulator of efferocytosis. PDCD6IP, also known as ALIX, is an accessory protein of ESCRT machinery, regulating multivesicular body formation and cytokinesis. Yet, how PDCD6IP may act as a molecular break on macrophage efferocytosis has not been characterized. Goals: Determine if knockout of Pdcd6ip enhances macrophage efferocytosis and protects against atherosclerosis pathogenesis. Methods: In vitro efferocytosis assays were performed in bone marrow-derived macrophages (BMDMs) and human monocyte-derived macrophages (HMDM). In vivo studies involved Ldlr -/- recipient mice with bone marrow transplantation (BMT) of WT and Pdcd6ip -/- BMs. Results: Pdcd6ip -/- induces cytokinesis arrest in proliferating BMDMs, resulting in an increased formation of binuclear polyploid BMDMs. Binuclear BMDMs have a remarkable capacity to engulf multiple apoptotic cells (ACs), termed continuing efferocytosis. While both mononuclear and binuclear Pdcd6ip -/- BMDMs demonstrate enhanced efferocytosis in primary efferocytosis, binuclear Pdcd6ip -/- BMDMs show enhanced continuing efferocytosis compared to binuclear WT BMDMs. Pdcd6ip -/- BMDMs have increased phagolysosomal acidification of the engulfed ACs, implicating proper degradation. Pdcd6ip -/- did not alter the engulfment of live cells, beads, or zymosan, supporting specific regulation of AC clearance. We hypothesize that Pdcd6ip -/- will be protective in atherosclerosis. Indeed, BMT of Pdcd6ip -/- BMs into Ldlr -/- mice led to increased atherosclerotic plaque stability after 22 weeks of Western diet feeding, characterized by decreased necrotic core area and increased fibrous cap thickness, without affecting the overall lesion area. Knockdown of PDCD6IP in HMDMs also led to an increased formation of binuclear HMDMs and enhanced continuing efferocytosis. Conclusion: We discovered a new role of PDCD6IP as a molecular break of macrophage efferocytosis. Inhibiting macrophage PDCD6IP may represent a therapeutic opportunity in atherosclerosis.

Abstract 133: Trem2 Agonist Promotes Atherosclerotic Plaque Stability

Atherosclerosis is an inflammatory disease of the arteries that is a major cause of cardiovascular disease (CVD). Atherosclerotic plaque progression is driven by the deposition of low-density lipoprotein (LDL) within the intima, which is internalized by macrophages, forming lipid-engorged foamy cells. Importantly, the accumulation of excess LDL within foamy macrophages is cytotoxic, driving apoptosis, which ultimately promotes necrotic core formation and increased potential for plaque rupture. Recently, our lab identified the lipid receptor Trem2 to be highly expressed by foamy macrophages and regulate cell survival, proliferation, and cholesterol efflux. Thus, we tested whether agonizing Trem2 could drive macrophage survival to promote plaque stability. Using a Trem2 agonist antibody (AL002a, Alector Inc.), we treated Ldlr -/- mice that had preexisting atherosclerotic plaque with either isotype control or AL002a antibody for 8 weeks. AL002a treated mice showed an increase in plaque size in the aortic arch and aortic sinus. Plaque analysis determined that this was primarily due to an expansion of macrophage cellularity, through promotion of cell survival and proliferation. Furthermore, we found that plaques from AL002a treated mice had decreased necrotic core formation, increased fibrous cap size, and increased collagen deposition, consistent with improved stability. Single cell RNA-sequencing of atherosclerotic aortae from isotype and AL002a treated mice found that Trem2-agonism dramatically reshaped transcriptomes of both foamy macrophages and smooth muscle cells, including upregulation of oxidative phosphorylation (OXPHOS), cholesterol efflux, collagen, and pro-survival genes while downregulating inflammatory pathways. In vitro studies using bone marrow derived macrophages confirmed AL002a’s ability to drive cell survival, proliferation, OXPHOS, and cholesterol efflux in culture. Finally, we determined that AL002a’s ability to reshape aspects of foamy macrophage function was dependent on its ability to activate the kinase SYK downstream of Trem2. Overall, this data identifies Trem2 as a viable therapeutic target for promoting atherosclerotic plaque stability and improving outcomes in CVD.

Lipid-Lowering Therapy after Acute Coronary Syndrome.

Achieving guideline-recommended low-density lipoprotein cholesterol (LDL-C) targets remains a significant challenge in clinical practice. This review assesses the barriers to reaching LDL-C goals and explores the potential solutions to these issues. When aiming for the recommended LDL-C goal, strategies like "lower is better" and "strike early and strong" should be used. The evidence supports the safety and efficacy of intensive lipid-lowering therapy post-acute coronary syndrome (ACS), leading to improved long-term cardiovascular health and atherosclerotic plaque stabilization. Despite the availability of effective lipid-lowering therapies, such as high-intensity statins, ezetimibe, the combination of both, bempedoic acid, and proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors, a substantial proportion of patients do not meet their LDL-C targets. Contributing factors include systemic healthcare barriers, healthcare provider inertia, patient non-adherence, and statin intolerance. Statin intolerance, often rather statin reluctance, is a notable obstacle due to perceived or expected side effects, which can lead to discontinuation of therapy. In conclusion, while there are obstacles to achieving optimal LDL-C levels post-ACS, these can be overcome with a combination of patient-centric approaches, clinical vigilance, and the judicious use of available therapies. The safety and necessity of reaching lower LDL-C goals to improve outcomes in patients post-ACS are well-supported by current evidence.

YKL40 potentially via activating the P38 pathway in decreasing atherosclerotic plaque stability.

YKL40 potentially via activating the P38 pathway in decreasing atherosclerotic plaque stability.

Elucidating the mechanisms of formononetin in modulating atherosclerotic plaque formation in ApoE-/- mice

BackgroundAtherosclerosis(AS) poses a pressing challenge in contemporary medicine. Formononetin (FMN) plays a crucial role in its prevention and treatment. However, the detailed impact of FMN on the stability of atherosclerotic plaques and its underlying mechanisms remain to be elucidated.MethodsAn intervention consisting of FMN was given along with a high-fat food regimen in the ApoE-/- mouse model. The investigation included the evaluation of the degree of atherosclerotic lesion, the main components of the plaque, lipid profiles, particular markers indicating M1/M2 macrophage phenotypes, the quantities of factors related to inflammation, the infiltration of macrophages, and the identification of markers linked to the α7nAChR/JAK2/STAT3 axis effect molecules.ResultsThe evaluation of aortic morphology in ApoE-/-mice revealed that FMN significantly improved the plaque area, fibrous cap protrusion, lipid deposition, and structural alterations on the aortic surface, among other markers of atherosclerosis,and there is concentration dependence. Furthermore, the lipid content of mouse serum was assessed, and the results showed that the low-, medium-, and high-dosage FMN groups had significantly lower levels of LDL-C, ox-LDL, TC, and TG. The results of immunohistochemical staining indicated that the low-, medium-, and high-dose FMN therapy groups had enhanced CD206 expression and decreased expression of CD68 and iNOS. According to RT-qPCR data, FMN intervention has the potential to suppress the expression of iNOS, COX-2, miR-155-5p, IL-6, and IL-1β mRNA, while promoting the expression of IL-10, SHIP1, and Arg-1 mRNA levels. However, the degree of inhibition varied among dosage groups. Western blot investigation of JAK/STAT signaling pathway proteins and cholinergic α7nAChR protein showed that p-JAK2 and p-STAT3 protein expression was suppressed at all dosages, whereas α7nAChR protein expression was enhanced.ConclusionsAccording to the aforementioned findings, FMN can reduce inflammation and atherosclerosis by influencing macrophage polarization, blocking the JAK/STAT signaling pathway, and increasing α7nAChR expression.

Potential Benefits and Risks Associated with the Use of Statins.

HMG-CoA reductase inhibitors, commonly known as statins, are the primary treatment choice for cardiovascular diseases, which stand as the leading global cause of mortality. Statins also offer various pleiotropic effects, including improved endothelial function, anti-inflammatory properties, reduced oxidative stress, anti-thrombotic effects, and the stabilization of atherosclerotic plaques. However, the usage of statins can be accompanied by a range of adverse effects, such as the development of type 2 diabetes mellitus, muscular symptoms, liver toxicity, kidney diseases, cataracts, hemorrhagic strokes, and psychiatric complications. These issues are referred to as statin-associated symptoms (SAS) and are relatively infrequent in clinical trials, making it challenging to attribute them to statin use definitively. Therefore, these symptoms can lead to significant problems, necessitating dose adjustments or discontinuation of statin therapy. This review aims to provide a comprehensive overview of the mechanism of action, potential advantages, and associated risks of statin utilization in clinical settings.

S-propargyl-cysteine promotes the stability of atherosclerotic plaque via maintaining vascular muscle contractile phenotype.

Introduction: Plaque rupture in atherosclerosis contributes to various acute cardiovascular events. As a new sulfide-containing donor, S-propargyl-cysteine (SPRC) has been reported to play a beneficial role in cardioprotection, potentially through its anti-inflammatory, anti-oxidative and anti-atherogenic activities. Our previous study observed an increase in eNOS phosphorylation in endothelial cells. However, it remains unclear whether SPRC influences vascular smooth muscle cells (VSMCs) within the plaque and if this effect contributes to plaque stabilization. Methods: An atherosclerotic unstable plaque mouse model was established by subjecting ApoE-/- mice to tandem stenosis of the right carotid artery along with a Western diet. Daily SPRC administration was conducted for 13weeks. Plaque morphology and stability were assessed using MRI scanning and histopathological staining. In our in vitro studies, we stimulated human artery vascular smooth muscle cells (HAVSMCs) with platelet-derived growth factor-BB (PDGF-BB), both with and without 100μM SPRC treatment. Cell phenotype was assessed using both Western blot and Real-time PCR. Cell proliferation was assessed using the BrdU cell proliferation kit and immunofluorescence of Ki-67, while cell migration was measured using scratch wound healing and transwell assay. MiR-143-3p overexpression and knockdown experiments were used to investigate whether it mediates the effect of SPRC on VSMC phenotype. Results and Discussion: SPRC treatment reduced plasma lipid levels, increased collagen content and decreased cell apoptosis in atherosclerotic plaques, indicating improved plaque stability. Both in vivo and in vitro studies elucidated the role of SPRC in preserving the contractile phenotype of VSMCs through up-regulation of miR-143-3p expression. Furthermore, SPRC suppressed the pro-proliferation and pro-migration effects of PDGF-BB on HAVSMCs. Overall, these findings suggest that the inhibitory effect of SPRC on phenotype switch from contractile to synthetic VSMCs may contribute to its beneficial role in enhancing plaque stability.

No evidence of coronary plaque stabilization by allopurinol in patients with acute coronary syndrome

BackgroundAllopurinol, a xanthine inhibitor that lowers uric acid concentration, has been proven to reduce inflammation and oxidative stress in patients with cardiovascular disease. However, it is unknown whether these beneficial effects translate into favorable plaque modification in acute coronary syndromes (ACS). This study aimed to investigate whether allopurinol could improve coronary plaque stabilization using coronary computed tomography angiography (CCTA). MethodsThis was a prospective, single-center, randomized, double-blind clinical trial began in March 2019. A total of 162 ACS patients aged 18–80 years with a blood level of high-sensitivity C-reactive protein (hsCRP) ​> ​2 ​mg/L were included. The subjects were randomly assigned in a 1:1 ratio to receive either allopurinol sustained-release capsules (at a dose of 0.25 ​g once daily) or placebo for 12 months. The plaque analysis was performed at CCTA. The primary efficacy endpoint was the change in low-attenuation plaque volume (LAPV) from baseline to the 12-month follow-up. ResultsAmong 162 patients, 54 in allopurinol group and 51 in placebo group completed the study. The median follow-up duration was 14 months in both groups. Compared with placebo, allopurinol therapy did not significantly alter LAPV (−13.4 ​± ​3.7 ​% vs. −17.8 ​± ​3.6 ​%, p ​= ​0.390), intermediate attenuation plaque volume (−16.1 ​± ​3.0 ​% vs. −16.2 ​± ​2.9 ​%, p ​= ​0.992), dense calcified plaque volume (12.2 ​± ​13.7 ​% vs. 9.7 ​± ​13.0 ​%, p ​= ​0.894), total atheroma volume (−15.2 ​± ​3.2 ​% vs. −16.4 ​± ​3.1 ​%, p ​= ​0.785), remodeling index (2.0 ​± ​3.9 ​% vs. 5.4 ​± ​3.8 ​%, p ​= ​0.536) or hsCRP levels (−73.6 [−91.6–17.9] % vs. −81.2 [−95.4–47.7] %, p ​= ​0.286). ConclusionsOur findings suggest that allopurinol does not improve atherosclerotic plaque stability or inflammation in ACS.

Statins-Beyond Their Use in Hypercholesterolemia: Focus on the Pediatric Population.

Statins are a class of medications primarily used in adults to lower cholesterol levels and reduce the risk of cardiovascular events. However, the use of statins in children is generally limited and carefully considered despite the well-documented anti-inflammatory, anti-angiogenic, and pro-apoptotic effects, as well as their effect on cell signaling pathways. These multifaceted effects, known as pleiotropic effects, encompass enhancements in endothelial function, a significant reduction in oxidative stress, the stabilization of atherosclerotic plaques, immunomodulation, the inhibition of vascular smooth muscle proliferation, an influence on bone metabolism, anti-inflammatory properties, antithrombotic effects, and a diminished risk of dementia. In children, recent research revealed promising perspectives on the use of statins in various conditions including neurological, cardiovascular, and oncologic diseases, as well as special situations, such as transplanted children. The long-term safety and efficacy of statins in children are still subjects of ongoing research, and healthcare providers carefully assess the individual risk factors and benefits before prescribing these medications to pediatric patients. The use of statins in children is generally less common than in adults, and it requires close monitoring and supervision by healthcare professionals. Further research is needed to fully assess the pleiotropic effects of statins in the pediatric population.

Fufang Zhenzhu Tiaozhi (FTZ) suppression of macrophage pyroptosis: Key to stabilizing rupture-prone plaques

BackgroundResearch on the Chinese herbal formula Fufang Zhenzhu Tiaozhi (FTZ) has demonstrated its effectiveness in treating hyperlipidemia and glycolipid metabolic disorders. Additionally, FTZ has shown inhibitory effects on oxidative stress, regulation of lipid metabolism, and reduction of inflammation in these conditions. However, the precise mechanisms through which FTZ modulates macrophage function in atherosclerosis remain incompletely understood. Therefore, this study aims to investigate whether FTZ can effectively stabilize rupture-prone plaques by suppressing macrophage pyroptosis and impeding the development of M1 macrophage polarization in ApoE−/− mice. MethodsTo assess the impact of FTZ on macrophage function and atherosclerosis in ApoE−/− mice, we orally administered FTZ at a dosage of 1.2 g/kg body weight daily for 14 weeks. Levels of interleukin-18 and interleukin-1β were quantified using ELISA kits to gauge FTZ's influence on inflammation. Total cholesterol content was measured with a Cholesterol Assay Kit to evaluate FTZ's effect on lipid metabolism. Aortic tissues were stained with Oil Red O, and immunohistochemistry techniques were applied to assess atherosclerotic lesions and plaque stability. To evaluate the effects of FTZ on macrophage pyroptosis and oxidative damage, immunofluorescence staining was utilized. Additionally, we conducted an analysis of protein and mRNA expression levels of NLRP3 inflammasome-related genes and macrophage polarization-related genes using RT-PCR and western blotting techniques. ResultsThis study illustrates the potential therapeutic effectiveness of FTZ in mitigating the severity of atherosclerosis and improving serum lipid profiles by inhibiting inflammation. The observed enhancements in atherosclerosis severity and inflammation can be attributed to the suppression of NLRP3 inflammasome activity and M1 polarization by FTZ. ConclusionThe current findings indicate that FTZ provides protection against atherosclerosis, positioning it as a promising candidate for novel therapies targeting atherosclerosis and related cardiovascular diseases.

Atherosclerotic plaque stabilization and regression: a review of clinical evidence.

Atherosclerotic plaque results from a complex interplay between lipid deposition, inflammatory changes, cell migration and arterial wall injury. Over the past two decades, clinical trials utilizing invasive arterial imaging modalities, such as intravascular ultrasonography, have shown that reducing levels of atherogenic lipoproteins, mainly serum LDL-cholesterol (LDL-C), to very low levels can safely reduce overall atherosclerotic plaque burden and favourably modify plaque composition. Classically, this outcome has been achieved with intensive statin therapy. Since 2016, newer and potent lipid-lowering strategies, such as proprotein convertase subtilisin-kexin type 9 inhibition, have shown incremental effects on plaque regression and risk of clinical events. Despite maximal reduction in plasma LDL-C levels, considerable residual cardiovascular risk remains in some patients. Therefore, there is a need to study therapeutic approaches that address residual risk beyond LDL-C reduction to promote plaque stabilization or regression. Contemporary imaging modalities, such as coronary computed tomography angiography, enable non-invasive assessment of the overall atherosclerotic plaque burden as well as of certain local plaque characteristics. This technology could allow further study of plaque stabilization and regression using novel therapeutic approaches. Non-invasive plaque assessment might also offer the potential to guide personalized management strategies if validated for this purpose.

The value of Lp(a) and TG/HDLC in peripheral blood to assess the stability of carotid plaque in patients with ischemic stroke.

The objective of this study was to investigate the relationship between lipoprotein (a) (Lp(a)), triglyceride/high-density lipoprotein cholesterol (TG/HDL-C), and the stability of carotid atherosclerotic plaque in patients with acute ischemic stroke. A total of 142 patients with acute ischemic stroke were selected and divided into group A (59 cases of stable plaque formation) and group B (83 cases of unstable plaque formation) according to the characteristics of carotid artery plaque formation detected by carotid color Doppler ultrasound. The serum Lp(a), lipid metabolism indexes, peripheral blood routine indexes, and related serum inflammatory factors indexes were compared between the two groups. Receiver operating characteristic curve and multivariate logistic regression model were used to analyze the relationship between each index and the formation of carotid unstable plaque. There were no significant differences in serum total cholesterol (TC), HDL-C, and low-density lipoprotein cholesterol (LDL-C) between groups A and B (p > .05). The values of Lp(a), TG, and TG/HDL-C in group B were higher than those in group A, and the differences were statistically significant (p < .05). There were no significant differences in serum TC, HDL-C, and LDL-C between groups A and B (p > .05). The values of Lp(a), TG, and TG/HDL-C in group B were higher than those in group A, and the differences were statistically significant (p < .05). The values of HBA1C, Lp-PLA2, CRP, CysC, Hcy, TNF-α, neutrophils, and NLR in group B were higher than those in group A, and the differences were statistically significant (p < .05). There was no significant difference in FPG, systolic blood pressure, diastolic blood pressure, Hb, white blood cells, platelets, and lymphocytes between groups A and B (p > .05). The results of logistic regression model showed that the increase of Lp(a), TG/HDL-C, HBA1C, Lp-PLA2, CRP, CysC, Hcy, and NLR could increase the risk of carotid artery unstable plaque in patients with ischemic stroke (p < .05). Lp(a) and TG/HDL-C have certain value in evaluating the stability of carotid atherosclerotic plaque in patients with acute ischemic stroke, and the increased levels of LP (a) and TG/HDL-C will significantly increase the risk of carotid unstable plaque in patients.

White blood cell counts can predict 4-year cardiovascular disease risk in patients with stable coronary heart disease: a prospective cohort study.

The prevalence of cardiovascular disease has increased sharply in the Asian population, and evaluation of the risk of cardiovascular events with stable coronary heart disease remains challenging. The role of white blood cell (WBC) count in assisting clinical decision-making in this setting is still unclear. This study sought to evaluate the prognostic meaning of WBC count among patients with stable coronary heart disease. This study included Asian participants (n = 1,933) from the prospective STABILITY (Stabilization of Atherosclerotic Plaque by Initiation of Darapladib Therapy) trial, which involved 15,828 patients with stable coronary heart disease with 3-5 years of follow-up on optimal secondary preventive treatment. WBC count was measured at baseline. Associations between WBC count and cardiovascular outcomes were evaluated by Cox regression analyses with multivariable adjustments. Hematologic emergencies in patients may introduce potential bias. In the lower WBC count quartiles, patients had lower-risk clinical profiles. Higher WBC counts were associated with greater event probabilities for cardiovascular death, major cardiovascular events, or all-cause death. In Cox regression models, WBC counts were an independent predictor of major adverse cardiovascular events (OR = 2.445, 95% CI 1.427-4.190, P = 0.001) for the primary outcomes. For the secondary outcomes, including the composite of all-cause death, cardiovascular death, myocardial infarction, stroke, and hospitalization for heart failure, WBC counts were significantly predictive of events with similar magnitude (OR = 1.716, 95% CI 1.169-2.521, P = 0.006). In patients with stable coronary heart disease, higher WBC counts were associated with a heightened risk for the primary or secondary outcomes. https://clinicaltrials.gov/; Unique identifier NCT00799903.

NSUN6 and HTR7 disturbed the stability of carotid atherosclerotic plaques by regulating the immune responses of macrophages.

Ischemic stroke associated with atherosclerosis is globally named atherothrombotic stroke. Presently, the underlying pathogenic genes promoting carotid atherosclerotic plaques transfer from a stable to unstable state remains elusive. This study aims to find the hub genes disturbing the stability of plaques and explore the primary cells affected by these hub genes. The optimal hub genes from five datasets for unstable plaques were identified by overlapping genes derived from Boruta and LASSO algorithms. The hub genes' expression levels in stroke patients were confirmed through RT-qPCR. Visualization of the hub genes' expression across various cell clusters was achieved with the aid of the Seurat R package. Then, hub genes were overexpressed or knock-down by lentivirus and siRNA, respectively. The inflammatory factors in the culture medium were detected using an ELISA assay. Eight genes (APOD, ASXL1, COL4A5, HTR7, INF2, NSUN6, PDSS2, and RBBP7) were identified and confirmed by RT-qPCR. The prognostic model was built upon this eight-gene composite foundation, and the area under the curve was 0.98. Based on CIBERSORT findings, unstable plaques displayed a higher macrophage proportion compared to stable ones (P < 0.05). These eight genes also correlated with infiltrated immune cells, especially macrophages. Then, according to single-cell RNA-seq analysis, we found that the eight hub genes mainly expressed in macrophages. The cellular localization of two hub genes (NSUN6 and HTR7) with high distinguishability was confirmed, and gene set enrichment analysis also clarified the possible biological pathways regulated by them. The findings from the in vitro investigation revealed that TNF-α and IL-6 were reduced in macrophages with NSUN6 overexpression or HTR7 knockdown. Eight hub genes, especially NSUN6 and HTR7, were found to promote the progression of plaques by regulating the immune responses of macrophages.

Cholesterol trafficking, lysosomal function, and atherosclerosis.

Despite years of study and major research advances over the past 50 years, atherosclerotic diseases continue to rank as the leading global cause of death. Accumulation of cholesterol within the vascular wall remains the main problem and represents one of the early steps in the development of atherosclerotic lesions. There is a complex relationship between vesicular cholesterol transport and atherosclerosis, and abnormalities in cholesterol trafficking can contribute to the development and progression of the lesions. The dysregulation of vesicular cholesterol transport and lysosomal function fosters the buildup of cholesterol within various intracytoplasmic compartments, including lysosomes and lipid droplets. This, in turn, promotes the hallmark formation of foam cells, a defining feature of early atherosclerosis. Multiple cellular processes, encompassing endocytosis, exocytosis, intracellular trafficking, and autophagy, play crucial roles in influencing foam cell formation and atherosclerotic plaque stability. In this review, we highlight recent advances in the understanding of the intricate mechanisms of vesicular cholesterol transport and its relationship with atherosclerosis and discuss the importance of understanding these mechanisms in developing strategies to prevent or treat this prevalent cardiovascular disease.