Reduced Plaque Size Research Articles

Leukemia inhibitory factor receptor inhibition by EC359 reduces atherosclerotic stenosis grade in Ldlr−/− mice

Cytokines are involved in all stages of atherosclerosis, generally contributing to disease progression. Previously, members of the Interleukin (IL)-6 cytokine family, such as IL-6, oncostatin M, and cardiotrophin-1, have been extensively studied in atherosclerosis. However, the role of leukemia inhibitory factor (LIF), member of the IL-6 family, and its receptor (LIFR), remains to be further elucidated. Therefore, the aim of this study is to provide insight in LIF receptor signalling in atherosclerosis development.Single-cell RNA sequencing analysis of human carotid artery plaques revealed that mast cells highly express LIF, whereas LIFR was specifically expressed on activated endothelial cells. A similar expression pattern of Lifr was observed in mouse atherosclerotic plaques. Next, female Western-type diet fed Ldlr−/− mice were treated with LIF receptor inhibitor EC359 (5 mg/kg s.c., n = 15) or control solvent (n = 15) three times per week for eight weeks. Stenosis grade was reduced in the aortic root of EC359 treated mice compared to control mice, but treatment did not affect plaque composition. Serum cholesterol levels were significantly reduced in EC359 treated mice, likely attributed to a reduction in VLDL cholesterol levels. Furthermore, LIF receptor inhibition reduced Pecam1 and Vcam1 expression in the aorta. Consequently, immune cell infiltration was reduced in aortic plaques of EC359 treated mice compared to control mice.Conclusively, we demonstrated that LIF receptor is a potential therapeutic target in atherosclerosis by reducing plaque size, attributed to lower serum cholesterol levels, reduced endothelial activation and less immune cell infiltration in the plaque.

The Impact of RNA Interference on Atherosclerotic Plaque Progression: A Meta-Analysis of Preclinical Studies

Background: Atherosclerosis, a chronic inflammatory disease characterized by the buildup of plaque within arteries, remains a leading cause of cardiovascular morbidity and mortality. RNA interference (RNAi) has emerged as a promising therapeutic strategy for atherosclerosis by targeting genes involved in plaque formation and progression. This meta-analysis aimed to evaluate the efficacy of RNAi in preclinical models of atherosclerosis. Methods: A systematic search of PubMed, Embase, and Web of Science databases was conducted from January 2013 to December 2023 to identify preclinical studies investigating the impact of RNAi on atherosclerotic plaque progression. Studies utilizing various RNAi modalities (siRNA, miRNA mimics/inhibitors, shRNA) targeting different genes involved in atherosclerosis were included. The primary outcome was plaque size reduction. Secondary outcomes included changes in plaque composition, lipid profiles, and inflammatory markers. A random-effects model was used to pool data and calculate standardized mean differences (SMD) with 95% confidence intervals (CI). Heterogeneity was assessed using the I² statistic. Results: Seven preclinical studies met the inclusion criteria, encompassing a total of 210 animals. RNAi interventions significantly reduced atherosclerotic plaque size compared to controls (SMD -1.51; 95% CI -2.36 to -0.66; p<0.00001; I²=12%). Analysis of secondary outcomes revealed favorable effects of RNAi on plaque composition, with a significant decrease in lipid content and an increase in collagen content. Furthermore, RNAi significantly improved lipid profiles by reducing total cholesterol and LDL-cholesterol levels. A significant reduction in inflammatory markers, such as TNF-α and IL-6, was also observed. Conclusion: This meta-analysis provides compelling evidence supporting the therapeutic potential of RNAi in attenuating atherosclerotic plaque progression in preclinical models. RNAi effectively reduced plaque size, improved plaque stability, and modulated lipid metabolism and inflammation.

Functional and morphological improvement of significant non-culprit coronary artery stenosis by LDL-C reduction with a PCSK9 antibody: Rationale and design of the randomized FITTER trial

Non-culprit coronary artery lesions are commonly present in patients presenting with an acute coronary syndrome (ACS). Additional stenting of non-culprit lesions in addition to the culprit lesion intends to prevent secondary events caused by these lesions. At the same time, multiple trials have demonstrated the potential of proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors in reducing plaque size and changing plaque composition of non-culprit lesions. Whether intensive low-density lipoprotein cholesterol (LDL-C) reduction with PCSK9 inhibitor evolocumab improves non-culprit vessel hemodynamics, reduces the risk of plaque rupture of important non-culprit lesions, and might obviate the need for additional stenting has not been investigated. The “Functional Improvement of non‐infarcT related coronary artery stenosis by Extensive LDL‐C Reduction with a PCSK9 Antibody” (FITTER) trial is a multi-center, randomized, double-blind, placebo-controlled clinical trial for patients presenting with ACS and multivessel disease (MVD). After treatment of the culprit lesion, fractional flow reserve (FFR) is performed in non-culprit vessels amenable for percutaneous coronary intervention (PCI). Coronary intervention in patients with hemodynamically important non-critical lesions (FFR: 0.67–0.85) is staged after baseline imaging using near-infrared spectroscopy (NIRS) and intravascular ultrasound (IVUS). Eligible patients are randomized and treated for 12 weeks with either evolocumab or placebo, in addition to high-intensity statin therapy. Follow-up angiography with repeat FFR and IVUS-NIRS is scheduled at 12 weeks. Staged PCI is performed at the operator's discretion.The FITTER trial is the first study to evaluate the effect of maximal LDL-C reduction by the PCSK9 inhibitor evolocumab on invasively measured FFR, plaque size, and plaque composition in hemodynamically important non-culprit lesions, during a treatment period of just 12 weeks after an ACS. Currently, all patients have been included (August 2023) and data analysis is ongoing. Trial registration numberclinicaltrials.gov NCT04141579.

Elastin-specific MR probe for visualization and evaluation of an interleukin-1β targeted therapy for atherosclerosis

Atherosclerosis is a chronic inflammatory condition of the arteries and represents the primary cause of various cardiovascular diseases. Despite ongoing progress, finding effective anti-inflammatory therapeutic strategies for atherosclerosis remains a challenge. Here, we assessed the potential of molecular magnetic resonance imaging (MRI) to visualize the effects of 01BSUR, an anti-interleukin-1β monoclonal antibody, for treating atherosclerosis in a murine model. Male apolipoprotein E-deficient mice were divided into a therapy group (01BSUR, 2 × 0.3 mg/kg subcutaneously, n = 10) and control group (no treatment, n = 10) and received a high-fat diet for eight weeks. The plaque burden was assessed using an elastin-targeted gadolinium-based contrast probe (0.2 mmol/kg intravenously) on a 3 T MRI scanner. T1-weighted imaging showed a significantly lower contrast-to-noise (CNR) ratio in the 01BSUR group (pre: 3.93042664; post: 8.4007067) compared to the control group (pre: 3.70679168; post: 13.2982156) following administration of the elastin-specific MRI probe (p < 0.05). Histological examinations demonstrated a significant reduction in plaque size (p < 0.05) and a significant decrease in plaque elastin content (p < 0.05) in the treatment group compared to control animals. This study demonstrated that 01BSUR hinders the progression of atherosclerosis in a mouse model. Using an elastin-targeted MRI probe, we could quantify these therapeutic effects in MRI.

Telomere stabilization by metformin mitigates the progression of atherosclerosis via the AMPK-dependent p-PGC-1α pathway.

Telomere dysfunction is a well-known molecular trigger of senescence and has been associated with various age-related diseases, including atherosclerosis. However, the mechanisms involved have not yet been elucidated, and the extent to which telomeres contribute to atherosclerosis is unknown. Therefore, we investigated the mechanism of metformin-induced telomere stabilization and the ability of metformin to inhibit vascular smooth muscle cell (VSMC) senescence caused by advanced atherosclerosis. The present study revealed that metformin inhibited the phenotypes of atherosclerosis and senescence in VSMCs. Metformin increased the phosphorylation of AMPK-dependent PGC-1α and thus increased telomerase activity and the protein level of TERT in OA-treated VSMCs. Mechanistically, the phosphorylation of AMPK and PGC-1α by metformin not only enhanced telomere function but also increased the protein level of TERT, whereas TERT knockdown accelerated the development of atherosclerosis and senescent phenotypes in OA-treated VSMCs regardless of metformin treatment. Furthermore, the in vivo results showed that metformin attenuated the formation of atherosclerotic plaque markers in the aortas of HFD-fed ApoE KO mice. Although metformin did not reduce plaque size, it inhibited the phosphorylation of the AMPK/PGC-1α/TERT signaling cascade, which is associated with the maintenance and progression of plaque formation, in HFD-fed ApoE KO mice. Accordingly, metformin inhibited atherosclerosis-associated phenotypes in vitro and in vivo. These observations show that the enhancement of telomere function by metformin is involved in specific signaling pathways during the progression of atherosclerosis. These findings suggest that telomere stabilization by metformin via the AMPK/p-PGC-1α pathway might provide a strategy for developing therapeutics against vascular diseases such as atherosclerosis.

M2 Macrophage-Derived Exosomes Inhibit Atherosclerosis Progression by Regulating the Proliferation, Migration, and Phenotypic Transformation of Smooth Muscle Cells.

Vascular smooth muscle cell (VSMC) intimal migration, proliferation, and phenotypic transformation from a contractile to a synthetic state are hallmarks of the progression of atherosclerotic plaques. This study aims to explore the effects of exosomes derived from M2 macrophages (ExoM2) on the pathological changes of VSMCs in atherosclerosis (AS). Cell Counting Kit-8 (CCK8) and wound healing assays were used to examine the impact of ExoM2 on platelet-derived growth factor-BB (PDGF-BB)-induced VSMC proliferation and migration, respectively. Western blotting was employed to analyze changes in the expression levels of contractile markers (e.g., alpha-smooth muscle actin [α-SMA]) and synthetic ones (e.g., osteopontin [OPN]) in VSMCs with or without ExoM2 treatment. ApoE-⁣/- mice on a high fat diet were utilized to observe the effects of ExoM2 on plaque progression and stability. Serial histopathological analysis was performed to elucidate the cellular mechanisms underlying the atheroprotective effects of ExoM2. Compared with controls, ExoM2 significantly inhibited PDGF-BB-induced VSMC proliferation, migration, and phenotypic transformation in vitro. In ApoE-⁣/- mice, ExoM2 treatment led to a marked reduction in plaque size, necrotic core area, the CD68/α-SMA ratio, and matrix metalloproteinase 9 (MMP9) and OPN levels, while enhancing plaque stability. ExoM2 inhibit AS progression by regulating VSMC proliferation, migration, and phenotypic transformation.

PIEZO1 targeting in macrophages boosts phagocytic activity and foam cell apoptosis in atherosclerosis

The rising incidences of atherosclerosis have necessitated efforts to identify novel targets for therapeutic interventions. In the present study, we observed increased expression of the mechanosensitive calcium channel Piezo1 transcript in mouse and human atherosclerotic plaques, correlating with infiltration of PIEZO1-expressing macrophages. In vitro administration of Yoda1, a specific agonist for PIEZO1, led to increased foam cell apoptosis and enhanced phagocytosis by macrophages. Mechanistically, PIEZO1 activation resulted in intracellular F-actin rearrangement, elevated mitochondrial ROS levels and induction of mitochondrial fragmentation upon PIEZO1 activation, as well as increased expression of anti-inflammatory genes. In vivo, ApoE−/− mice treated with Yoda1 exhibited regression of atherosclerosis, enhanced stability of advanced lesions, reduced plaque size and necrotic core, increased collagen content, and reduced expression levels of inflammatory markers. Our findings propose PIEZO1 as a novel and potential therapeutic target in atherosclerosis.

Biofilm Formation, Motility, and Virulence of Listeria monocytogenes Are Reduced by Deletion of the Gene lmo0159, a Novel Listerial LPXTG Surface Protein.

Listeria monocytogenes (L. monocytogenes) is a foodborne pathogen that causes listeriosis in humans and other animals. Surface proteins with the LPXTG motif have important roles in the virulence of L. monocytogenes. Lmo0159 is one such protein, but little is known about its role in L. monocytogenes virulence, motility, and biofilm formation. Here, we constructed and characterized a deletion mutant of lmo0159 (∆lmo0159). We analyzed not only the capacity of biofilm formation, motility, attachment, and intracellular growth in different cell types but also LD50; bacterial load in mice's liver, spleen, and brain; expression of virulence genes; and survival time of mice after challenge. The results showed that the cross-linking density of the biofilm of ∆lmo0159 strain was lower than that of WT by microscopic examination. The expression of biofilm-formation and virulence genes also decreased in the biofilm state. Subsequently, the growth and motility of ∆lmo0159 in the culture medium were enhanced. Conversely, the growth and motility of L. monocytogenes were attenuated by ∆lmo0159 at both the cellular and mouse levels. At the cellular level, ∆lmo0159 reduced plaque size; accelerated scratch healing; and attenuated the efficiency of adhesion, invasion, and intracellular proliferation in swine intestinal epithelial cells (SIEC), RAW264.7, mouse-brain microvascular endothelial cells (mBMEC), and human-brain microvascular endothelial cells (hCMEC/D3). The expression of virulence genes was also inhibited. At the mouse level, the LD50 of the ∆lmo0159 strain was 100.97 times higher than that of the WT strain. The bacterial load of the ∆lmo0159 strain in the liver and spleen was lower than that of the WT strain. In a mouse model of intraperitoneal infection, the deletion of the lmo0159 gene significantly prolonged the survival time of the mice, suggesting that the lmo0159 deletion mutant also exhibited reduced virulence. Thus, our study identified lmo0159 as a novel virulence factor among L. monocytogenes LPXTG proteins.

Decreased atherosclerosis and abdominal aortic aneurysm in mice deficient in polymeric immunoglobulin receptor

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): La Caixa Banking Foundation and Comunidad de Madrid Background Polymeric immunoglobulin receptor (PIGR) is a transmembrane protein involved in the transcytosis of polymeric immunoglobulins that is widely expressed in mucosal epithelial cells. Recent data from our group described increased PIGR levels in plasma and tissue of atherosclerotic subjects. However, the role for PIGR in the mechanisms involved in vascular remodeling has not been addressed. Purpose To analyse the role for PIGR in experimental mouse models of atherosclerosis and abdominal aortic aneurysm (AAA). Methods We analyzed PIGR expression, secretion and distribution in early atherosclerotic plaques and AAA tissues compared to healthy aortic samples, as well as in macrophages derived from the human monocytic THP-1 cell line. Next, we studied the effect of global PIGR deficiency in experimental atherosclerosis (Ldlr-/- Pigr-/- mice fed with atherogenic diet for 10 weeks). In addition, the contribution of hematopoietic PIGR was assessed in both experimental atherosclerosis and AAA (Ldlr-/- mice fed with atherogenic diet and 1 µg/Kg/min angiotensin II infusion for 28 days) by bone marrow transplantation experiments. Finally, bone marrow-derived macrophages (BMDMs) from Pigr-/- and control mice were isolated to assess their inflammatory expression profile and modulation of polarization in response to oxidized LDL (ox-LDL), as well as their ability to promote foam cell formation. Results PIGR expression was increased in the intima of early human atherosclerotic lesions and showed colocalization with CD68+ expression. PIGR expression was also increased during the in vitro differentiation of THP-1 monocytes to macrophages with phorbol myristate acetate. Both global and hematopoietic-specific PIGR deficient mice displayed reduced plaque size in the aortic sinuses as well as a strong decrease in macrophage infiltration (CD68+ cells), while no changes in smooth muscle cells (α-SMA+ cells) and collagen content were observed. Regarding AAA, increased PIGR levels were found in human AAA tissues, associated with infiltrating CD68+ cells. PIGR deficiency in hematopoietic cells also showed significantly reduced AAA dilatation and reduced macrophage infiltration. In vitro experiments with BMDMs obtained from Pigr-/- mice and stimulated with ox-LDL showed reduced proinflammatory gene expression (TNF-a and IL-6) and M1 markers (iNOS and CD11c) as well as diminished foam cell formation. Conclusions PIGR deficiency in hematopoietic cells decreases atherosclerosis and AAA progression by modulating macrophage inflammatory responses, suggesting a novel proinflammatory role for PIGR in vascular pathologies. Future experiments will test the potential therapeutic effect of modulating PIGR to dampen the inflammatory profile in vascular diseases.Results in human and mouse studies

Mild phototherapy mediated by IR780-Gd-OPN nanomicelles suppresses atherosclerotic plaque progression through the activation of the HSP27-regulated NF-κB pathway

Reducing plaque lipid content and enhancing plaque stability without causing extensive apoptosis of foam cells are ideal requirements for developing a safe and effective treatment of atherosclerosis. In this study, we synthesized IR780-Gd-OPN nanomicelles by conjugating osteopontin (OPN) and loading a gadolinium-macrocyclic ligand (Gd-DOTA) onto near-infrared dye IR780-polyethylene glycol polymer. The nanomicelles were employed for mild phototherapy of atherosclerotic plaques and dual-mode imaging with near-infrared fluorescence and magnetic resonance. In vitro results reveal that the mild phototherapy mediated by IR780-Gd-OPN nanomicelles not only activates heat shock protein (HSP) 27 to protect foam cells against apoptosis but also inhibits the nuclear factor kappa-B (NF-κB) pathway to regulate lipid metabolism and macrophage polarization, thereby diminishing the inflammatory response. In vivo results further validate that mild phototherapy effectively reduces plaque lipid content and size while simultaneously enhancing plaque stability by regulating the ratio of M1 and M2-type macrophages. In summary, this study presents a promising approach for developing a safe and highly efficient method for the precise therapeutic visualization of atherosclerosis. Statement of significanceThe rupture of unstable atherosclerotic plaques is a major cause of high mortality rates in cardiovascular diseases. Therefore, the ideal outcome of atherosclerosis treatment is to reduce plaque size while enhancing plaque stability. To address this challenge, we designed IR780-Gd-OPN nanomicelles for mild phototherapy of atherosclerosis. This treatment can effectively reduce plaque size while significantly improving plaque stability by increasing collagen fiber content and elevating the ratio of M2/M1 macrophages, which is mainly attributed to the inhibition of the NF-κB signaling pathway by mild phototherapy-activated HSP27. In summary, our proposed mild phototherapy strategy provides a promising approach for safe and effective treatment of atherosclerosis.

Abstract 3063: Canonical And Noncanonical Heat Shock Factor 1 Signaling In Smooth Muscle Cells Promote Atherosclerotic Plaque Burden

Smooth muscle cells (SMCs) in atherosclerotic plaques undergo complex phenotypic modulation, characterized by migration from the aortic medial layer to the intima, proliferation, de-differentiation, and variable increases in molecular markers characteristic of macrophages, fibroblasts, osteogenic cells and mesenchymal stem cells. This transition is driven in part by cholesterol entering the endoplasmic reticulum (ER) and activating ER stress. The importance of ER stress-driven PERK signaling in atherosclerosis is underscored by the observation that SMC-specific deletion of Perk reduces lesion size by ~70% in hypercholesterolemic (HC) mice. HC mice harboring either an SM α-actin ( ACTA2 ) missense variant or SMC-specific deletion of the centrosomal scaffolding protein pericentrin ( PCNT ) have increased augmented plaque burden compared to their HC wildtype (WT) counterparts, despite having comparable serum lipid levels. We went on to demonstrate that both these genetic alterations increase SMC cytosolic stress and activation of canonical heat shock factor 1 (HSF1) signaling, which in turn increases HMG-CoA reductase (HMGCR) activity and cholesterol biosynthesis, ER stress, PERK signaling, SMC phenotypic modulation and finally, augments the plaque burden. We also noted another distinct pro-atherogenic form of HSF1 activation in SMCs: SMCs in culture exposed to high levels of exogenous cholesterol activate PERK signaling, but also consistently activate HSF1, and blocking PERK signaling prevents HSF1 activation, suggesting “noncanonical” HSF1 activation. While global loss of Hsf1 in HC mice reduces plaque burden, we generated tamoxifen-inducible SMC-specific Hsf1 -deficient mice ( Hsf1 SMC-/- ) to assess the role of HSF1 signaling in SMCs of HC mice. Hyperlipidemia was induced in these mice by a single injection of AAV- PCSK9 DY followed by a high fat diet for 12 weeks. En face aortic Oil Red O staining demonstrated that Hsf1 SMC-/- mice had 67% reduction in plaque size in the whole aorta (p=0.0007), compared to HC WT mice, similar to the results seen with SMC-specific Perk deficiency. These data suggest distinct roles of canonical and noncanonical HSF1 signaling in SMCs that contribute to atherosclerotic plaque burden.

Abstract 2075: Vascular Smooth Muscle Cell Derived Pdgfd Promotes Vascular Remodeling And Plaque Progression In Atherosclerosis

We have recently identified rs2019090 and PDGFD as the functional variant and gene mediating CAD risk at the 11q22.3 locus, with our initial analysis using a global knockout (KO) model showing that this gene may promote phenotypic changes in smooth muscle cells (SMCs) in the plaque and contribute to neointimal vascular calcification. Nonetheless, the specific cell-type and phenotypic states through which PDGFD may confer disease risk remains unexplored.To delineate the impact of SMC-derived PDGFD signalling on cell state transitions and plaque progression in atherosclerosis, we have developed a novel SMC-specific lineage tracing and Pdgfd KO mouse ( Pdgfd ΔSMC/ΔSMC , Myh11 CreERT2 , ROSA tdT/+ , ApoE -/- ) allowing us to confidently define the impact of SMC-derived Pdgfd in the vascular SMC lineage as well as in neighbouring populations. Leveraging our lineage tracing KO mutants on a hypercholesterolemic diet, we have employed single cell RNA sequencing (scRNAseq) and histological analyses to characterize the cellular and molecular effect of Pdgfd in vascular disease. SMC-specific Pdgfd deletion resulted in alterations in the distribution of transitioning SMC populations, as well as previously unexplored gene expression differences within these cell types. This was accompanied by a significant reduction in atherosclerotic burden and plaque size across the aorta, including aortic root as well as descending abdominal aorta. Histological analysis revealed decreased monocyte recruitment, show by quantifying CD68+ cells within the plaque. To explore this further we interrogated the scRNAseq dataset to identify major pathways of cell-cell communication and the impact of altered Pdgfd signalling across different cell types. Overall, these data reveal that SMC-derived PDGFD substantially alters lesional SMC cell phenotype transitions, as well as inflammatory cell recruitment, implicating it as a major regulator of atherosclerotic disease progression.

Interleukin-1 receptor accessory protein blockade limits the development of atherosclerosis and reduces plaque inflammation.

The interleukin-1 receptor accessory protein (IL1RAP) is a co-receptor required for signalling through the IL-1, IL-33, and IL-36 receptors. Using a novel anti-IL1RAP-blocking antibody, we investigated the role of IL1RAP in atherosclerosis. Single-cell RNA sequencing data from human atherosclerotic plaques revealed the expression of IL1RAP and several IL1RAP-related cytokines and receptors, including IL1B and IL33. Histological analysis showed the presence of IL1RAP in both the plaque and adventitia, and flow cytometry of murine atherosclerotic aortas revealed IL1RAP expression on plaque leucocytes, including neutrophils and macrophages. High-cholesterol diet fed apolipoprotein E-deficient (Apoe-/-) mice were treated with a novel non-depleting IL1RAP-blocking antibody or isotype control for the last 6 weeks of diet. IL1RAP blockade in mice resulted in a 20% reduction in subvalvular plaque size and limited the accumulation of neutrophils and monocytes/macrophages in plaques and of T cells in adventitia, compared with control mice. Indicative of reduced plaque inflammation, the expression of several genes related to leucocyte recruitment, including Cxcl1 and Cxcl2, was reduced in brachiocephalic arteries of anti-IL1RAP-treated mice, and the expression of these chemokines in human plaques was mainly restricted to CD68+ myeloid cells. Furthermore, in vitro studies demonstrated that IL-1, IL-33, and IL-36 induced CXCL1 release from both macrophages and fibroblasts, which could be mitigated by IL1RAP blockade. Limiting IL1RAP-dependent cytokine signalling pathways in atherosclerotic mice reduces plaque burden and plaque inflammation, potentially by limiting plaque chemokine production.

Recent H9N2 avian influenza virus lost hemagglutination activity due to a K141N substitution in hemagglutinin.

The H9N2 avian influenza virus (AIV) represents a significant risk to both the poultry industry and public health. Our surveillance efforts in China have revealed a growing trend of recent H9N2 AIV strains exhibiting a loss of hemagglutination activity at 37°C, posing challenges to detection and monitoring protocols. This study identified a single K141N substitution in the hemagglutinin (HA) glycoprotein as the culprit behind this diminished hemagglutination activity. The study evaluated the evolutionary dynamics of residue HA141 and studied the impact of the N141K substitution on aspects such as virus growth, thermostability, receptor-binding properties, and antigenic properties. Our findings indicate a polymorphism at residue 141, with the N variant becoming increasingly prevalent in recent Chinese H9N2 isolates. Although both wild-type and N141K mutant strains exclusively target α,2-6 sialic acid receptors, the N141K mutation notably impedes the virus's ability to bind to these receptors. Despite the mutation exerting minimal influence on viral titers, antigenicity, and pathogenicity in chicken embryos, it significantly enhances viral thermostability and reduces plaque size on Madin-Darby canine kidney (MDCK) cells. Additionally, the N141K mutation leads to decreased expression levels of HA protein in both MDCK cells and eggs. These findings highlight the critical role of the K141N substitution in altering the hemagglutination characteristics of recent H9N2 AIV strains under elevated temperatures. This emphasizes the need for ongoing surveillance and genetic analysis of circulating H9N2 AIV strains to develop effective control and prevention measures.IMPORTANCEThe H9N2 subtype of avian influenza virus (AIV) is currently the most prevalent low-pathogenicity AIV circulating in domestic poultry globally. Recently, there has been an emerging trend of H9N2 AIV strains acquiring increased affinity for human-type receptors and even losing their ability to bind to avian-type receptors, which raises concerns about their pandemic potential. In China, there has been a growing number of H9N2 AIV strains that have lost their ability to agglutinate chicken red blood cells, leading to false-negative results during surveillance efforts. In this study, we identified a K141N mutation in the HA protein of H9N2 AIV to be responsible for the loss of hemagglutination activity. This finding provides insight into the development of effective surveillance, prevention, and control strategies to mitigate the threat posed by H9N2 AIV to both animal and human health.

The Efficacy and Safety of Low‐Intensity Pulsed Ultrasound Therapy for the Treatment of Peyronie’s Disease (A Report of 106 Cases): A Retrospective Study

Objective: To evaluate the efficacy and safety of low‑intensity pulsed ultrasound (LIPUS) for the treatment of Peyronie’s disease (PD).Methods: To analyze the treatment effect of 106 PD patients, 56 patients treated with LIPUS as therapy group and 50 patients with oral L‐carnitine and vitamin E as control group. 56 patients were treated with LIPUS three times per week for four consecutive weeks (12 times totally). Penile erectile pain, erectile function, the size of penile plaques, and penile curvature were assessed for all of these 106 PD patients before treatment and at 4‐ and 24‐week follow‐ups to evaluate the treatment effect. Erectile pain was assessed by visual analog scale (VAS), while erectile function using the International Index of Erectile Function scale (IIEF‐5), and penile plaque size was measured by ultrasound. After alprostadil was injected into the penile body to induce a full erection, the penis was measured with a protractor at the maximum bending degree.Results: After 4‐weeks follow‐up, the VAS scores of the LIPUS group were significantly lower than the baseline (P &lt; 0.01), while no significant differences were found in the control group (P &gt; 0.05). After 24‐weeks follow‐up, the VAS scores of the control group reduced significantly but also were significantly higher than the LIPUS group (P &lt; 0.01). The IIEF‐5 scores of the LIPUS group were significantly higher than the baseline (P &lt; 0.01). While the IIEF‐5 scores of the control group showed no significant differences both after 4‐ and 24‐weeks follow‐up (P &gt; 0.05). The penile plaque size showed no significant differences after 4‐ and 24‐weeks follow‐up both in the LIPUS group and the control group (P &gt; 0.05). The penile curvature degrees showed a significant reduction after 4‐weeks follow‐up in LIPUS group (P &lt; 0.01). While the penile curvature degrees of the control group showed no significant differences both after 4‐ and 24‐weeks follow‐up (P &gt; 0.05). No complications, such as penile pain, hematoma, or skin ecchymosis, were observed during the entire process of LIPUS treatment, nor did they appear during follow‐up.Conclusions: In patients with acute PD, LIPUS could relieve erectile pain and improve erectile dysfunction. Moreover, it appears not to reduce plaque size and not to have a clinically significant effect on penile curvature. Furthermore, LIPUS also shows a good safety profile during the management of PD and should be recommended for the clinical management of PD.

IL-1 alpha is myristoylated for monocyte surface translocation and promotes atherosclerosclerosis in mice

Abstract Background The role of Interleukin-1 alpha (IL-1α) and beta (IL-1β) in the pathogenesis of atherosclerosis in vivo has been controversial for years. In contrast to IL-1β, which is dependent on the NLR family pyrin domain containing 3 (NLRP3) inflammasome, IL-1α can be tethered to the plasma membrane (csIL-1α) where it exerts pro-inflammatory effects. We characterised the role of IL-1α and the Nlrp3 inflammasome in a nongenetic hyperlipidemic mouse model and examined the function of csIL-1α in human monocytes. Methods Atherosclerosis was induced by a single injection of mutant PCSK9-AAV8 virus in wild-type (WT, referred to as PCSK9-AAV), Il1a-/-, Nlrp3-/-, and Il1b-/- mice, in combination with high-fat western diet (21% fat, 0.2% cholesterol) for 12 weeks. Plaque size and lipid content, were histologically analysed. The serum proteome was studied using a targeted multiplex PEA (Olink Proteomics). Human primary monocytes and human umbilical vein endothelial cells (HUVECs) were used to examine csIL-1α-IL1R interaction by analyzing VCAM1 expression and monocyte adhesion. LPS was used to induce IL-1α protein synthesis and membrane translocation in vitro. Myristoylation was measured via flow cytometry. Results Il1a-/- animals showed significantly reduced atherosclerotic lesion size (-62% vs. PCSK9-AAV) and fat accumulation within the plaque (-64% vs. PCSK9-AAV). Knockout of Nlrp3 and IL-1β did not reduce plaque size or lipid content, indicating an Nlrp3-independent mechanism of the protective effect of Il1a-/-. Circulating pro-inflammatory cytokines such as IL-1α, IL-1β, and IL-6 were downregulated in serum of Il1a-/-, Nlrp3-/-, and Il1b-/- mice compared to PCSK9-AAV mice suggesting that the development of atherosclerotic plaques is independent of circulating inflammatory cytokines at an early stage. Therefore, we investigated the role of csIL-1α on human monocytes. Stimulation with LPS showed enrichment of csIL-1α by 13.2% (p&amp;lt;0.05) on monocytes without releasing IL-1α. Treatment of HUVECs with LPS-stimulated monocytes resulted in increased VCAM1 expression (+22.2%, p&amp;lt;0.05) and monocyte adhesion (1.9 fold, p&amp;lt;0.05) through a csIL-1α-IL1R interaction which was abolished by the addition of neutralising antibodies for IL1R1 and IL-1α (p&amp;lt;0.05, vs. LPS-treated monocytes). To investigate whether the IL-1α translocation is mediated by protein myristoylation, human monocytes were pre-incubated with the n-myristoyl-transferase inhibitor IMP-1088 before LPS stimulation, resulting in a reduction of csIL-1α expression (p&amp;lt;0.05, vs. LPS-treated monocytes). Conclusion This study demonstrated a protective effect of Il1a deficiency on the development of atherosclerosis. CsIL-1α mediates monocyte-to-endothelial adhesion by increasing VCAM1 expression through endothelial IL1R1 signaling. The data underscore the importance of the juxtacrine signaling of IL-1α and its role in the development of atherosclerosis independent of circulating cytokine levels.

Abstract 16155: Achieving Coronary Artery Disease Regression: A Triumph of Lifestyle Modification and Optimal Medical Therapy

Background: Coronary artery disease (CAD) is the leading cause of death worldwide. Early recognition of risk factors and targeting them using lifestyle modifications and pharmacological measures is vital for secondary prevention. This case demonstrates the effectiveness of comprehensive management in achieving marked regression of CAD. Case presentation: A 45-year-old female with a history of type II diabetes mellitus, hypertension, and hyperlipidemia with markedly elevated lipoprotein (A) of 233 nmol/L presented with chronic coronary syndrome (CCS). She had a strong family history of CAD and denied smoking, alcohol use, and recreational drug use. Transthoracic echocardiography revealed preserved left ventricular systolic function with an ejection fraction of 50%, regional wall motion abnormalities in the form of inferior wall hypokinesia, and mild mitral regurgitation. She underwent coronary angiography (CA) which revealed significant three-vessel CAD. Figure 1 Decision making: She underwent successful percutaneous coronary intervention (PCI) to the RCA and PDA with two overlapping drug-eluted stents and plain balloon angioplasty to the PL branch. The patient adhered to strict lifestyle modification and was prescribed optimal anti-ischemic and anti-diabetic medications. She lost 22 lbs over two years, her LDL went down from 178 to 14 mg/dl and A1c from 12.2 to 6.8. During follow up, she started to complain of dyspnea on exertion for two weeks. Despite the marked laboratory improvement and despite she was found to be anemic at that time which can explain her symptoms, decision was to proceed to CA given her high pretest probability. A repeat CA showed marked regression in her CAD. Figure 2 Conclusion: Regression of coronary artery disease is a reality giving hope for not just stopping disease progression but achieving reduction in plaque size with optimal medical therapy, where pharmacological measures and lifestyle modifications work synergistically.

Nanodiamond Decorated PEO Oxide Coatings on NiTi Alloy.

Cardiovascular diseases (CVDs) remain a leading cause of death in the European population, primarily attributed to atherosclerosis and subsequent complications. Although statin drugs effectively prevent atherosclerosis, they fail to reduce plaque size and vascular stenosis. Bare metal stents (BMS) have shown promise in acute coronary disease treatment but are associated with restenosis in the stent. Drug-eluting stents (DES) have improved restenosis rates but present long-term complications. To overcome these limitations, nanomaterial-based modifications of the stent surfaces have been explored. This study focuses on the incorporation of detonation nanodiamonds (NDs) into a plasma electrolytic oxidation (PEO) coating on nitinol stents to enhance their performance. The functionalized ND showed a high surface-to-volume ratio and was incorporated into the oxide layer to mimic high-density lipoproteins (HDL) for reverse cholesterol transport (RCT). We provide substantial characterization of DND, including stability in two media (acetone and water), Fourier transmission infrared spectroscopy, and nanoparticle tracking analysis. The characterization of the modified ND revealed successful functionalization and adequate suspension stability. Scanning electron microscopy with EDX demonstrated successful incorporation of DND into the ceramic layer, but the formation of a porous surface is possible only in the high-voltage PEO. The biological assessment demonstrated the biocompatibility of the decorated nitinol surface with enhanced cell adhesion and proliferation. This study presents a novel approach to improving the performance of nitinol stents using ND-based surface modifications, providing a promising avenue for cardiovascular disease.

Updated recommendations on the therapeutic role of extracorporeal shock wave therapy for peyronie’s disease: systematic review and meta-analysis

BackgroundThe therapeutic role of extracorporeal shockwave therapy (ESWT) for Peyronie’s disease (PD) has been controversial in a long term. We aimed to further evaluate the therapeutic effect of ESWT for PD on the basis of available high-quality studies.MethodsThe PubMed, CENTRAL and Embase databases were searched for articles published from January 1st, 2000 to December 31, 2022. Only randomized controlled trials (RCTs) using ESWT to treat PD were included. Meta-analysis and forest plots were carried out using Review Manager 5.4.1 software, and outcomes were reviewed by 2 authors independently. Using the Risk of Bias assessment form (ROB-2) by Cochrane Collaboration for quality assessment. PRISMA 2020 guidelines were used in this article to achieve the quantitative and qualitative synthesis of data.ResultsA total of four RCTs were included. 151 patients in the ESWT group and 150 patients in the control group. The meta-analysis results showed that ESWT could significantly reduce plaque size (OR 2.59, 95%CI 1.15 to 5.85, P = 0.02) and relieve pain (MD -1.55, 95%CI -2.46 to -0.64, P = 0.0008); but it has no significant effect on reducing the penile curvature (OR 1.93, 95%CI 0.87–4.26, P = 0.11) and improving sexual function (MD 2.6, 95%CI -1.63 to 6.83, P = 0.23), there is also no significant difference in complication rates between groups (OR 2.94, 95%CI 0.66 to 13.03, P = 0.16). The risk of bias of results is low. The limitations of this study are that the number of included studies is too small, some experimental outcomes are missing, and the expression of outcomes is not unified.ConclusionsFor PD, ESWT can be considered as a safe short-term treatment, which can reduce plaque size and relieve pain, but cannot improve penile curvature and sexual function. Its long-term efficacy remains to be discussed.Registration numberPROSPERO (ID: CRD42023436744).

Long-term, in vivo therapeutic effects of a single dose of miR-145 micelles for atherosclerosis.

Atherosclerosis is a chronic inflammatory disease that is characterized by the build-up of lipid-rich plaques in the arterial walls. The standard treatment for patients with atherosclerosis is statin therapy aimed to lower serum lipid levels. Despite its widespread use, many patients taking statins continue to experience acute events. Thus, to develop improved and alternative therapies, we previously reported on microRNA-145 (miR-145 micelles) and its ability to inhibit atherosclerosis by targeting vascular smooth muscle cells (VSMCs). Importantly, one dose of miR-145 micelles significantly abrogated disease progression when evaluated two weeks post-administration. Thus, in this study, to evaluate how long the sustained effects of miR-145 micelles can be maintained and towards identifying a dosing regimen that is practical for patients with chronic disease, the therapeutic effects of a single dose of miR-145 micelles were evaluated for up to two months in vivo. After one and two months post-treatment, miR-145 micelles were found to reduce plaque size and overall lesion area compared to all other controls including statins without causing adverse effects. Furthermore, a single dose of miR-145 micelle treatment inhibited VSMC transdifferentiation into pathogenic macrophage-like and osteogenic cells in plaques. Together, our data shows the long-term efficacy and sustained effects of miR-145 micelles that is amenable using a dosing frequency relevant to chronic disease patients.