Smooth Muscle Cell Infiltration Research Articles

Enhanced atherosclerosis in apolipoprotein E knockout rabbits: role of apoB48-rich remnant lipoproteins.

Apolipoprotein E (apoE) acts as a binding molecule for both the low-density lipoprotein receptor and the lipoprotein receptor-related protein and this function is essential for facilitating the hepatocyte uptake of lipoproteins containing apoB. The absence of apoE leads to increased atherogenicity in both humans and mice, although the precise molecular mechanisms remain incompletely understood. This study aimed to investigate the susceptibility of apoE knockout (KO) rabbits, in comparison with wild-type (WT) rabbits, to diet-induced hyperlipidemia and atherosclerosis. ApoE KO rabbits and WT rabbits were fed a diet containing 0.3% cholesterol for 16 weeks. Plasma lipid levels, lipoproteins, and apolipoproteins were analyzed. Atherosclerosis was evaluated at the endpoint of experiments. In addition, we evaluated the oxidizability of those lipoproteins containing apoB to investigate the possible mechanisms of atherosclerosis. Male apoE KO rabbits showed significantly elevated levels of total cholesterol and triglycerides compared to WT rabbits, while female apoE KO rabbits displayed similar high total cholesterol levels, albeit with significantly higher triglycerides levels than WT controls. Notably, both male (2.1-fold increase) and female (1.6-fold increase) apoE KO rabbits exhibited a significantly augmented aortic lesion area compared to WT controls. Pathological examination showed that the increased intimal lesions in apoE KO rabbits were featured by heightened infiltration of macrophages (2.7-fold increase) and smooth muscle cells (2.5-fold increase). Furthermore, coronary atherosclerotic lesions were also increased by 1.3-fold in apoE KO rabbits. Lipoprotein analysis revealed that apoB48-rich beta-very-low-density lipoproteins were notably abundant in apoE KO rabbits, suggesting that these remnant lipoproteins of intestinal origin serve as the primary atherogenic lipoproteins. Moreover, apoB48-rich remnant lipoproteins isolated from apoE KO rabbits exhibited heightened susceptibility to copper-induced oxidation. The findings indicate that apoB48-rich remnant lipoproteins, resulting from apoE deficiency, possess greater atherogenic potential than apoB100-rich remnant lipoproteins, regardless of plasma TC levels.

The promotion of vascular reconstruction by hierarchical structures in biodegradable small-diameter vascular scaffolds

Tissue engineering of small-diameter vessels remains challenging due to the inadequate ability to promote endothelialization and infiltration of smooth muscle cells (SMCs). Ideal vascular graft is expected to provide the ability to support endothelial monolayer formation and SMCs infiltration. To achieve this, vascular scaffolds with both orientation and dimension hierarchies were prepared, including hierarchically random vascular scaffold (RVS) and aligned vascular scaffold (AVS), by utilizing degradable poly(ε-caprolactone)-co-poly(ethylene glycol) (PCE) and the blend of PCE/gelatin (PCEG) as raw materials. In addition to the orientation hierarchy, dimension hierarchy with small pores in the inner layer and large pores in the outer layer was also constructed in both RVS and AVS to further investigate the promotion of vascular reconstruction by hierarchical structures in vascular scaffolds. The results show that the AVS with an orientation hierarchy that consists with the natural vascular structure had better mechanical properties and promotion effect on the proliferation of vascular cells than RVS, and also exhibited excellent contact guidance effects on cells. While the dimension hierarchy in both RVS and AVS was favorable to the rapid infiltration of SMCs in a short culture time in vitro. Besides, the results of subcutaneous implantation further demonstrate that AVS achieved a fully infiltrated outer layer with wavy elastic fibers-mimic strips formation by day 14, ascribing to hierarchies of aligned orientation and porous dimension. The results further indicate that the scaffolds with both orientation and dimension hierarchical structures have great potential in the application of promoting the vascular reconstruction.

Effect of curcumin nanoparticles on proliferation and migration of mouse airway smooth muscle cells and airway inflammatory infiltration.

Curcumin (CUR) possesses the capability to inhibit various inflammatory factors, exert anti-inflammatory effects, and alleviate asthma attacks; however, its hydrophobicity and instability significantly impede its clinical application. In this study, we synthesized CUR-loaded nanoparticles (CUR-NPs) and evaluated their impact on the proliferation, migration, and inflammatory infiltration of mouse airway smooth muscle cells (ASMCs), while investigating their underlying mechanisms. To achieve this objective, ASMCs were isolated from BALB/c mice and subjected to TGF-β1-induced cell proliferation and migration. Our findings demonstrate that CUR-NPs effectively regulate the release of CUR within cells with superior intracellular uptake compared to free CUR. The CCK-8 assay results indicate that the blank carrier does not exhibit any cytotoxic effects on cells, thus rendering the impact of the carrier itself negligible. The TGF-β1 group exhibited a significant increase in cell proliferation, whereas treatment with CUR-NPs significantly suppressed TGF-β1-induced cell proliferation. The findings from both the cell scratch assay and transwell assay demonstrated that TGF-β1 substantially enhanced cell migration, while CUR-NPs treatment effectively attenuated TGF-β1-induced cell migration. The Western blot analysis demonstrated a substantial increase in the expression levels of TGF-β1, p-STAT3, and CTGF in ASMCs following treatment with TGF-β1 when compared to the control group. Nevertheless, this effect was effectively counteracted upon administration of CUR-NPs. Furthermore, an asthma mouse model was successfully established and CUR-NPs were administered through tail vein injection. The serum levels of TGF-β1 and the expression levels of TGF-β1, p-STAT3, and CTGF proteins in the lung tissue of mice in the model group exhibited significant increases compared to those in the control group. However, CUR-NPs treatment effectively attenuated this change. Our research findings suggest that CUR-NPs possess inhibitory effects on ASMC proliferation, migration, and inflammatory infiltration by suppressing activation of the TGF-β1/p-STAT3/CTGF signaling pathway, thereby facilitating inhibition of airway remodeling.

Modulation of Smooth Muscle Cell Phenotype for Translation of Tissue-Engineered Vascular Grafts.

Translation of small-diameter tissue-engineered vascular grafts (TEVGs) for the treatment of coronary artery disease (CAD) remains an unfulfilled promise. This is largely due to the limited integration of TEVGs into the native vascular wall-a process hampered by the insufficient smooth muscle cell (SMC) infiltration and extracellular matrix deposition, and low vasoactivity. These processes can be promoted through the judicious modulation of the SMC toward a synthetic phenotype to promote remodeling and vascular integration; however, the expression of synthetic markers is often accompanied by a decrease in the expression of contractile proteins. Therefore, techniques that can precisely modulate the SMC phenotypical behavior could have the potential to advance the translation of TEVGs. In this review, we describe the phenotypic diversity of SMCs and the different environmental cues that allow the modulation of SMC gene expression. Furthermore, we describe the emerging biomaterial approaches to modulate the SMC phenotype in TEVG design and discuss the limitations of current techniques. In addition, we found that current studies in tissue engineering limit the analysis of the SMC phenotype to a few markers, which are often the characteristic of early differentiation only. This limited scope has reduced the potential of tissue engineering to modulate the SMC toward specific behaviors and applications. Therefore, we recommend using the techniques presented in this review, in addition to modern single-cell proteomics analysis techniques to comprehensively characterize the phenotypic modulation of SMCs. Expanding the holistic potential of SMC modulation presents a great opportunity to advance the translation of living conduits for CAD therapeutics.

Early vascular responses to abluminal biodegradable polymer-coated versus circumferential durable polymer-coated newer-generation drug-eluting stents in humans: a pathological study.

Recent clinical studies are testing strategies for short (1-3 months) dual antiplatelet therapy following newer-generation drug-eluting stent (DES) placement. However, detailed biological responses to newer-generation DES remain unknown in humans. We sought to evaluate early pathologic responses to abluminal biodegradable polymer-coated(BP-) DES compared with circumferential durable polymer-coated (DP-) DES in human autopsy cases. The study included 38 coronary lesions with newer-generation DES implanted for<90 days(DP-DES=24, BP-DES=14) in 26 autopsy cases. The degree of strut coverage was defined as follows: grade 0 (bare), grade 1 (with fibrin or tissues/cells without endothelium), grade 2 (with single-layered endothelium), and grade 3 (with endothelium and underlying smooth muscle cell layers). The duration following implantation was similar in DP- and BP-DES (median=20 vs 17 days). A total of 2,022 struts (DP-DES=1,297, BP-DES=725) were pathologically analysed. Focal grade 2 coverage was observed as early as 5 days after the implantation in both stents. The multilevel mixed-effects ordered logistic regression model demonstrated that BP-DES exhibited greater strut coverage compared with DP-DES (odds ratio [OR]: 3.64, 95% confidence interval [CI]: 1.37-9.67; p=0.009), which remained significant after adjustment for the duration following implantation and underlying tissue characteristics (OR: 2.74, 95% CI: 1.10-6.80; p=0.030). The predictive probability of grade 2 and 3 coverage was comparably limited at 30 days (DP-DES=17.1%, BP-DES=28.7%) and increased at 90 days (DP-DES=76.5%, BP-DES=86.6%). Both stents showed low inflammation and a similar degree of fibrin deposition. Single-layered endothelial coverage begins in the days after newer-generation DES placement, and BP-DES potentially exhibit faster strut coverage with smooth muscle cell infiltration than DP-DES in humans. Nevertheless, vessel healing remains suboptimal in both stents at 30 days.

Preclinical in vivo assessment of a cell-free multi-layered scaffold prepared by 3D printing and electrospinning for small-diameter blood vessel tissue engineering in a canine model.

Tissue-engineered vascular grafts (TEVGs) are promising alternatives to existing prosthetic grafts. The objective of this study is to evaluate the clinical feasibility of a novel multi-layered small-diameter vascular graft that has a hierarchical structure. Vascular grafts with elaborately designed composition and architecture were prepared by 3D printing and electrospinning and were implanted into the femoral artery of 5 dogs. The patency of the grafts was assessed using Doppler ultrasonography. After 6 months, the grafts were retrieved and histological and SEM examinations were conducted. During implantation, the grafts exhibited resistance to kinking and no blood seepage thanks to the helical structure of the innermost and outermost layers. The grafts showed a high patency rate and remodelling ability. At 6 months post-implantation, the lumen was endothelialized and middle layers were regenerated by infiltration of smooth muscle cells (SMCs) and deposition of extracellular matrix (ECM). These results suggest that the multi-layered vascular graft may be a promising candidate for small-diameter blood vessel tissue engineering in clinical practice.

Early vascular responses to abluminal biodegradable polymer-coated versus circumferential durable polymer-coated newer-generation drug-eluting stents in humans: a pathologic study

Abstract Background Recent clinical trials are testing strategies for short (1–3 months) dual antiplatelet therapy (DAPT) following newer-generation drug-eluting stent (DES) placement. However, the safety of short DAPT regimens is not supported by biological evidence in humans. Purpose We sought to evaluate early pathologic responses to newer-generation DES by comparing abluminal biodegradable polymer-coated DES (BP-DES) with circumferential durable polymer-coated DES (DP-DES) in human autopsy cases. Methods The study included a total of 37 coronary lesions with thin strut newer-generation DES (DP-DES=23 [XIENCE=18, Resolute Integrity=5] and BP-DES=14 [SYNERGY=9, Ultimaster=5]) with duration of implantation &amp;lt;90 days in 25 autopsy cases. The process of stent healing was precisely evaluated for every single strut in association with underlying tissue characteristics. The degree of strut coverage was defined as follows: grade 0 (bare struts), grade 1 (struts covered with thrombus, fibrin, or other tissues or cells without endothelium), grade 2 (struts covered with single-layered endothelium without underlying smooth muscle cell layers), and grade 3 (struts covered with endothelium and underlying smooth muscle cell layers) (Figure 1). Results Duration of implantation was similar in lesions with DP-DES and those with BP-DES (median=20 vs. 17 days). A total of 1986 struts (DP-DES=1261, BP-DES=725) were pathologically analyzed. Focal grade 2 coverage was observed as early as 5 days after the implantation in both stents. Multilevel mixed-effects ordered logistic regression model demonstrated that BP-DES exhibited greater strut coverage compared with DP-DES (odds ratio; 3.50, 95% CI; 1.31–9.41, P=0.013), which remained significant after adjustment for duration of implantation and underlying tissue characteristics (odds ratio; 2.64, 95% CI; 1.04–6.68, P=0.040). The time course of vessel healing assessed as predictive probability of strut coverage (grade 0–3) stratified by duration of implantation is shown in Figure 2. Predictive probability of grade 2 and 3 coverage was comparably limited at 30 days (DP-DES=17.7% vs. BP-DES=29.0%) and increased at 90 days (DP-DES=76.1% vs. BP-DES=85.9%). Both stents showed few inflammation and similar degree of fibrin deposition. Conclusions The current first pathologic study on early biological responses to newer-generation DES in humans demonstrated that single-layered endothelial coverage begins in days following the stent placement, and abluminal BP-DES potentially exhibit faster strut coverage with smooth muscle cell infiltration than circumferential DP-DES. Nevertheless, vessel healing remains suboptimal at 30 days in both DP- and BP-DES, which progresses with time to become substantial at 90 days. Our results suggest that very short duration of DAPT for 1 month should be applied with caution, taking into account the trade-off between bleeding and thrombotic risks. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Grant-in-Aid for Scientific Research (C) from the Japan Society for the Promotion of Science

Chitosan/silk fibroin biomimic scaffolds reinforced by cellulose acetate nanofibers for smooth muscle tissue engineering

Reconstruction of multilayered and functional smooth muscle tissues remains a challenge nowadays. Cryogels possess great advantages in three-dimensional tissue regeneration owing to the interconnected macroporous structure, but their applications have been hindered because of limited mechanical properties. Inspired by the natural extracellular matrix, cellulose acetate electrospun nanofibers (NFs) were incorporated to chitosan/silk fibroin (CS/SF) cryogel scaffolds to address this problem in this work. Compared with pure CS/SF scaffolds, CS/SF/NFs composite scaffolds showed roughened surface and enlarged pore size (216.2 ± 65.3 μm vs. 263.1 ± 75.7 μm) allowing for cell adhesion and proliferation. Incorporation of nanofibers upgraded the mechanical properties of scaffolds with a six-fold increase in compressive modulus. The proliferation and infiltration of smooth muscle cells (SMCs) were remarkably improved with the existence of nanofibers. Besides, SMCs cultured on CS/SF/NFs scaffolds displayed higher expression of contraction-related genes, verifying their potential for smooth muscle tissue engineering.

Abstract 379: Smooth Muscle-specific Lncrna Carmn Plays A Potential Role In Aortic Aneurysm

Background: Aortic aneurysm is a life-threatening vascular disease characterized by vascular smooth muscle cell (VSMC) depletion, ECM degradation, and infiltration of immune cells. Previous studies have shown that Carmn , a SMC-enriched long non-coding RNA, is critical for maintaining VSMC contractile phenotype. However, its functional role in aortic aneurysm remains completely unknown. Methods and Results: We re-analyzed the publicly available aortic aneurysm bulk RNA-seq and scRNA-seq datasets in mouse and human. These unbiased analyses revealed that Carmn is downregulated in the SMCs of aortic aneurysm samples. To examine the potential role of Carmn in aortic aneurysm, we generated Carmn global knockout (gKO) mice or inducible SMC-specific KO (iKO) mice driven by Myh11 -Cre ER T2, respectively. Unexpectedly, both Carmn gKO and iKO mice exhibited premature lethality, due to a severe colonic pseudo-obstruction resulting from deletion of Carmn in visceral SMCs. Despite the dominant gastrointestinal phenotype observed in Carmn KO mice, morphological analysis of the Carmn KO thoracic aorta revealed thinner vascular wall compared to that of controls. Results from TEM showed that the VSMC in the Carmn KO thoracic aorta are disorganized and degradative with aneurysmal dissection. Data from bulk RNA-seq of Carmn -deficient aorta revealed that the down-regulated genes are associated with degradation of ECM while the up-regulated genes are associated with cell death. Moreover, scRNA-seq of Carmn -deficient aorta revealed that the percentage of the SMCs is decreased while the percentage of infiltrated immune cells is increased. These data indicated that Carmn deficiency may play a critical role in promoting the development of aortic aneurysm. To circumvent the lethal visceral SMC phenotype, we are in the progress of crossing Carmn flox mice with the novel vascular SMC-specific inducible Itga8 -Cre ER T2 mouse to generate VSMC-specific Carmn KO mice. Conclusions: Our data suggest that Carmn is indispensable for maintaining gastrointestinal contractile function, and implicate that Carmn deficiency plays a potential role in contributing the development of aortic aneurysm.

Physico-mechanical and biological evaluation of heparin/VEGF-loaded electrospun polycaprolactone/decellularized rat aorta extracellular matrix for small-diameter vascular grafts

Although the continuous development of small-diameter vascular grafts (SDVGs) (D < 5 mm) continues, most vascular grafts are made from synthetic polymers, which lead to serious complications from arteriosclerosis, thrombosis, and vascular ischemia. Here, to address these shortcomings, we combine synthetic polymers with natural decellularized small-diameter vessels and loaded with growth factor. We fabricated vascular grafts by electrospinning polycaprolactone (PCL) to decellularized rat aorta matrix (ECM) followed by heparin and vascular endothelial growth factor (VEGF) loading. In- vitro studies showed that PCL/ECM/VEGF vascular grafts, showed excellent hemocompatibility and biocompatibility properties. The vascular grafts implanted into the rat aorta revealed that the PCL/ECM/VEGF grafts promotes endothelial cells and smooth-muscle cells infiltration with a rate of FLK-1, ICAM1, and a-SMA distribution higher than that of the PCL and PCL/ECM vascular grafts at 2 weeks and 4 weeks after implantation. The PCL/ECM/VEGF vascular graft should be considered for potential small-diameter vascular grafts in clinical fields.

Omalizumab and modification of bronchial asthma natural course

Clinical and molecular heterogeneity of bronchial asthma has been documented in recent years. The search for novel solutions to enhance efficient patient support related first of all to understanding of asthma heterogenic nature and allows to personalize each patient treatment. Biological therapy application can influence to achieve better control at greater extent for patients with severe uncontrolled asthma. Nowadays 5 biological drugs are registered on Russian Federation territory and implemented according to severe asthma phenotypes: anti-IgE, anti-IL-4,13 and anti-IL-5 class therapies. Omalizumab become the first target drug for uncontrolled allergic asthma patients (monoclonal antibody against IgE). This medication is prescribed for uncontrolled moderate and severe allergic (atopic) asthma in patients on basic asthma therapy according GINA step 4 and 5 (Level of evidence A). Clinical trials confidently reported that anti- IgE-therapy reduces the rate of asthma exacerbations, severity of disease in patients with chronic severe asthma on high doses of inhaled steroids or systemic steroids and allows to reduce or withdraw systemic steroids doses in case of steroid-dependent asthma. For the last years special attention led to and demonstrated omalizumab positive effect on airways remodeling and modification of bronchial asthma natural course in adults and children. Antiinflammatory effect of omalizumab is documented. Omalizumab significantly reduces eosinophilic infiltration of submucosal bronchi layer among patients with atopic asthma, sputum eosinophilia, which correlates with reduction of FeNO during biologic treatment, reduces mast cells infiltration of smooth muscle cells in bronchi. Omalizumab significantly reduces the thickness of the bronchial wall, increases the lumen of the bronchi (positive dynamics of CT-scan parameters), which is clinically manifested by increased of FEV1.

Histopathological analysis of in vivo specimens of recurrent aneurysms after coil embolization

BackgroundAneurysm recurrence after coil embolization remains a challenging problem.ObjectiveTo determine the histopathological features of recurrent aneurysm specimens and explore the mechanism of aneurysm recurrence.MethodsNine aneurysm specimens were collected from eight...

Metformin Suppresses the Progress of Diabetes-Accelerated Atherosclerosis by Inhibition of Vascular Smooth Muscle Cell Migration Through AMPK-Pdlim5 Pathway.

Backgrounds: Our previous work revealed that AMP-activated protein kinase (AMPK) activation inhibits vascular smooth muscle cell migration in vitro by phosphorylating PDZ and LIM domain 5 (Pdlim5). As metformin is an AMPK activator, we used a mouse vascular smooth muscle cell (VSMC) line and a Myh11-cre-EGFP mice to investigate whether metformin could inhibit the migration of VSMCs in vitro and in a wire-injury model in vivo. It is recognized that VSMCs contribute to the major composition of atherosclerotic plaques. In order to investigate whether the AMPK–Pdlim5 pathway is involved in the protective function of metformin against atherosclerosis, we utilized ApoE−/− male mice to investigate whether metformin could suppress diabetes-accelerated atherosclerosis by inhibition of VSMC migration via the AMPK–Pdlim5 pathway.Methods: The mouse VSMC cell line was exogenously transfected wild type, phosphomimetic, or unphosphorylatable Pdlim5 mutant before metformin exposure. Myh11-cre-EGFP mice were treated with saline solution or metformin after these were subjected to wire injury in the carotid artery to study whether metformin could inhibit the migration of medial VSMCs into the neo-intima. In order to investigate whether the AMPK–Pdlim5 pathway is involved in the protective function of metformin against atherosclerosis, ApoE−/− male mice were divided randomly into control, streptozocin (STZ), and high-fat diet (HFD)-induced diabetes mellitus; STZ+HFD together with metformin or Pdlim5 mutant carried the adenovirus treatment groups.Results: It was found that metformin could induce the phosphorylation of Pdlim5 and inhibit cell migration as a result. The exogenous expression of phosphomimetic S177D-Pdlim5 inhibits lamellipodia formation and migration in VSMCs. It was also demonstrated that VSMCs contribute to the major composition of injury-induced neointimal lesions, while metformin could alleviate the occlusion of the carotid artery. The data of ApoE−/− mice showed that increased plasma lipids and aggravated vascular smooth muscle cell infiltration into the atherosclerotic lesion in diabetic mice were observed Metformin alleviated diabetes-induced metabolic disorders and atherosclerosis and also reduced VSMC infiltration in atherosclerotic plaques, while the Pdlim5 phospho-abolished mutant that carried adenovirus S177A-Pdlim5 undermines the protective function of metformin.Conclusions: The activation of the AMPK–Pdlim5 pathway by metformin could interrupt the migratory machine of VSMCs and inhibit cell migration in vitro and in vivo. The maintenance of AMPK activity by metformin is beneficial for suppressing diabetes-accelerated atherosclerosis.

Intracellular glutamine level determines vascular smooth muscle cell-derived thrombogenicity

Background and aimsThe everolimus-eluting stent (EES), one of the effective stents for in-stent restenosis (ISR), has a lower incidence of stent thrombosis; however, the underlying mechanism remains unknown. This study aimed to identify the effects of everolimus on vascular metabolism and thrombogenicity and examine their mechanistic link. MethodsEESs and bare-metal stents were implanted in rabbit iliac arteries with smooth muscle cell (SMC)-rich neointima induced by endothelial denudation. Four weeks after stent implantation, the stented arteries were examined for histological analysis and metabolomics. Additionally, everolimus effects in coronary artery SMCs metabolism, tissue factor (TF) expression, and procoagulant activity were assessed in vitro. ResultsEES-implanted arteries showed decreased neointima formation, less SMCs infiltration, and reduced TF expression. Concomitantly, they were metabolically characterized by increased levels of metabolites in amino acids, such as glutamine. Similarly, everolimus increased intracellular glutamine levels, decreased TF expression, and reduced procoagulant activity in SMCs in vitro. On the contrary, exogenous glutamine administration also increased intracellular glutamine level, decreased TF expression, and reduced procoagulant activity despite enhanced mammalian target of rapamycin (mTOR) activity. ConclusionsIntracellular glutamine level is likely to determine vascular SMC-related thrombogenicity regardless of mTOR pathway activity. Therefore, increased intracellular glutamine level might contribute partially to the beneficial effect of EES use on stent thrombosis.

Anti-Atherosclerotic Effect of Afrocyclamin A against Vascular Smooth Muscle Cells Is Mediated via p38 MAPK Signaling Pathway.

Objective Research suggests that fine particulate matter (PM2.5) contributes to the expansion and development of atherosclerosis. Infiltration and proliferation of vascular smooth muscle cells (VSMCs) from the blood vessel media into the intima, is an important step in the atherosclerosis pathophysiology. Afrocyclamin A, is an oleanane-type triterpene saponin, isolated from Androsace umbellate, which is commonly used in Chinese herbal medicine. In the study, we examined the effect of Afrocyclamin A on PM2.5-induced VSMCs proliferation and scrutinized possible mechanisms of action. Materials and Methods In the experimental study, counting Kit-8 (CCK-8) assay was used for estimation of VSMCs viability. BrdU immunofluorescence was used for estimation of VSMCs proliferation. The levels of antioxidant parameters such as malonaldehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH); proinflammatory cytokines such as interleukin-1β (IL-1β), IL-6, tumor necrosis factor-α (TNF-α), nitric oxide (NO), endothelin-1 (ET-1), and vascular cell adhesion molecule-1 (VCAM-1), were estimated. The expression of proliferating cell nuclear antigen (PCNA) and phospho-p38 MAPK (p-p38 MAPK) was assessed. Results Compared to PM2.5-treated cells, in addition to reducing PM2.5-induced VSMCs proliferation, Afrocyclamin A reduced the expression of PCNA and p-p38 MAPK, down-regulated the level of TNF-α, IL-1β, IL-6, VCAM-1, MDA and ET-1, and up-regulated SOD, GSH and NO level. Furthermore, the anti-proliferative effect of Afrocyclamin A was considerably increased following co-incubation of Afrocyclamin A with SB203580 (p38 MAPK inhibitor) in comparison with Afrocyclamin A-treated cells. ConclusionBased on the results, we can conclude that Afrocyclamin A might reduce PM2.5-induced VSMCs proliferation via reduction of p38 MAPK signaling pathway.

Single-Cell RNA Sequencing Reveals Heterogeneity of Vascular Cells in Early Stage Murine Abdominal Aortic Aneurysm-Brief Report.

Abdominal aortic aneurysm (AAA) is a life-threatening vascular disease characterized by smooth muscle cell depletion, ECM (extracellular matrix) degradation, and infiltration of immune cells. The cellular and molecular profiles that govern the heterogeneity of the AAA aorta are yet to be elucidated. Approach and Results: We performed single-cell RNA sequencing on mouse AAA tissues. AAA was induced in C57BL/6J mice by perivascular application of CaCl2. Unbiased clustering identified 12 distinct populations of 8 cell types. Percentages of each population and gene expression were compared between sham and AAA tissue. Furthermore, we characterized the transcriptional profiles and potential functional features of populations in smooth muscle cells, fibroblasts, and macrophages and revealed the unique regulons in each cell type. Together, these data provide high-resolution insight into the complexity and heterogeneity of mouse AAA and indicate that populations within major cell types such as smooth muscle cells, fibroblasts, and macrophages may contribute differently to AAA pathogenesis. Graphic Abstract: A graphic abstract is available for this article.

Wnt Signaling Cascades and Their Role in Coronary Artery Health and Disease

The Wnt signaling is classified as two distinct pathways of canonical Wnt/β-catenin signaling, and the non-canonical pathways of planar cell polarity and Wnt/Ca2+ pathways. However, the scientific discoveries in recent years have shown that canonical and non-canonical Wnts pathways are intertwined and have complex interaction with other major signaling pathways such as hedgehog, Hippo and TOR signaling. Wnt signaling plays important roles in cell proliferation, differentiation and migration during embryonic development. The impairment of these pathways during embryonic development often leads to major congenital defects. In adult organisms Wnt expression is more restricted to proliferating tissues, where it plays a key role in tissue regeneration. In addition, the disruption of homeostatic processes of multicellular organisms may give rise to reactivation and/or altered activation of Wnt signaling, leading to development of malignant tumors and chronic diseases such as type-2 diabetes and adult cardiovascular diseases.Coronary artery disease (CAD) is the leading cause of death in the world. The disease is the consequences of two distinct disease processes: Atherosclerosis, a primarily inflammatory disease and plaque erosion, a disease process associated with endothelial cell defect and smooth muscle proliferation with only modest contribution of inflammatory cells. The atherosclerosis is itself a multifactorial disease that is initiated by lipid deposition and endothelial dysfunction, triggering vascular inflammation via recruitment and aggregation of monocytes and their transformation to foam cell by the uptake of modified low-density lipoprotein (LDL), culminating in an atheromatous plaque core formation. Further accumulation of lipids, infiltration and proliferation of vascular smooth muscle cells (VSMCs) and extracellular matrix deposition result in intimal hyperplasia. Myocardial infarction is the ultimate consequence of these processes and is caused by plaque rupture and hypercoagulation. In vivo studies have established the role of the Wnt pathway in all phases of atherosclerosis development, though much remains unknown or controversial. Less is known about the mechanisms that induce plaque erosion. The limited evidence in mouse models of Wnt coreceptor LRP6 mutation and heterozygous TCF7L2 knock out mice implicate altered Wnt signaling also in the pathogenesis of plaque erosion. In this article we focus and review the role of the Wnt pathway in CAD pathophysiology from clinical and experimental standpoints.

The role of Wnt signalling in development of coronary artery disease and its risk factors.

The Wnt signalling pathways are composed of a highly conserved cascade of events that govern cell differentiation, apoptosis and cell orientation. Three major and distinct Wnt signalling pathways have been characterized: the canonical Wnt pathway (or Wnt/β-catenin pathway), the non-canonical planar cell polarity pathway and the non-canonical Wnt/Ca2+ pathway. Altered Wnt signalling pathway has been associated with diverse diseases such as disorders of bone density, different malignancies, cardiac malformations and heart failure. Coronary artery disease is the most common type of heart disease in the United States. Atherosclerosis is a multi-step pathological process, which starts with lipid deposition and endothelial cell dysfunction, triggering inflammatory reactions, followed by recruitment and aggregation of monocytes. Subsequently, monocytes differentiate into tissue-resident macrophages and transform into foam cells by the uptake of modified low-density lipoprotein. Meanwhile, further accumulations of lipids, infiltration and proliferation of vascular smooth muscle cells, and deposition of the extracellular matrix occur under the intima. An atheromatous plaque or hyperplasia of the intima and media is eventually formed, resulting in luminal narrowing and reduced blood flow to the myocardium, leading to chest pain, angina and even myocardial infarction. The Wnt pathway participates in all different stages of this process, from endothelial dysfunction to lipid deposit, and from initial inflammation to plaque formation. Here, we focus on the role of Wnt cascade in pathophysiological mechanisms that take part in coronary artery disease from both clinical and experimental perspectives.

Exploring the role of TREM-1 receptor in experimental Abdominal Aortic Aneurysm

Non-syndromic Abdominal Aortic Aneurysm (AAA) is one of the leading causes of cardiovascular death in elderly men. AAA is characterized by extracellular matrix degradation, smooth muscle cell apoptosis and infiltration of inflammatory cells in the aortic wall. TREM (Triggering Receptor Expressed on Myeloid cells)-1, a receptor expressed by myeloid cells, is a key regulator of innate immune cell activity, and we hypothesized that it might be involved in AAA development. We aim to decipher the role of TREM-1 in the immuno-inflammatory response associated with AAA development and rupture. To study the role of TREM-1 in AAA development, we used an experimental model of Angiotensin-II-induced AAA in ApoE-/-Trem-1-/- and ApoE-/-Trem-1+/+ mice. At day 0, 3, 7 and 28, we analyzed monocyte trafficking (flow cytometry), inflammatory cell infiltration within the aortic wall (immunostaining, qPCR), aortic wall remodeling (histology) and finally AAA development. Angiotensin II infusion increased TREM-1 expression on both classical and non-classical monocytes. TREM-1 deficiency limits Angiotensin II-induced monocytosis at day 3 and limits macrophage infiltration in the aorta at day 7. Trem-1 genetic deletion limits pro-inflammatory response and metalloprotease activity in the aortic wall and protects against elastin degradation. Finally, TREM-1 deficiency significantly reduced AAA rupture and severity. We showed that TREM-1 deficiency reduced AAA development and rupture in an Angiotensin II-induced experimental mouse model.

Extracellular Vesicles Enhance the Remodeling of Cell-Free Silk Vascular Scaffolds in Rat Aortae.

Vascular tissue engineering is aimed at developing regenerative vascular grafts to restore tissue function by bypassing or replacing defective arterial segments with tubular biodegradable scaffolds. Scaffolds are often combined with stem or progenitor cells to prevent acute thrombosis and initiate scaffold remodeling. However, there are limitations to cell-based technologies regarding safety and clinical translation. Extracellular vesicles (EVs) are nanosized particles released by most cell types, including stem and progenitor cells, that serve to transmit protein and RNA cargo to target cells throughout the body. EVs have been shown to replicate the therapeutic effect of their parent cells; therefore, EVs derived from stem or progenitor cells may serve as a more translatable, cell-free, therapeutic base for vascular scaffolds. Our study aims to determine if EV incorporation provides a positive effect on graft patency and remodeling in vivo. We first assessed the effect of human adipose-derived mesenchymal stem cell (hADMSC) EVs on vascular cells using in vitro bioassays. We then developed an EV-functionalized vascular graft by vacuum-seeding EVs into porous silk-based tubular scaffolds. These constructs were implanted as aortic interposition grafts in Lewis rats, and their remodeling capacity was compared to that observed for hADMSC-seeded and blank (non-seeded) controls. The EV group demonstrated improved patency (100%) compared to the hADMSC (56%) and blank controls (82%) following eight weeks in vivo. The EV group also produced significantly more elastin (126.46%) and collagen (44.59%) compared to the blank group, while the hADMSC group failed to produce significantly more elastin (57.64%) or collagen (11.21%) compared to the blank group. Qualitative staining of the explanted neo-tissue revealed improved endothelium formation, increased smooth muscle cell infiltration, and reduced macrophage numbers in the EV group compared to the controls, which aids in explaining this group's favorable pre-clinical outcomes.