Deficiency of smooth muscle cell ILF3 alleviates intimal hyperplasia via HMGB1 mRNA degradation-mediated regulation of the STAT3/DUSP16 axis.

Intimal hyperplasia is a complicated pathophysiological phenomenon attributable to in-stent restenosis, and the underlying mechanism remains unclear. Interleukin enhancer-binding factor 3 (ILF3), a double-stranded RNA-binding protein involved in regulating mRNA stability, has been recently demonstrated to assume a crucial role in cardiovascular disease; nevertheless, its impact on intimal hyperplasia remains unknown. In current study, we used samples of human restenotic arteries and rodent models of intimal hyperplasia, we found that vascular smooth muscle cell (VSMC) ILF3 expression was markedly elevated in human restenotic arteries and murine ligated carotid arteries. SMC-specific ILF3 knockout mice significantly suppressed injury induced neointimal formation. In vitro, platelet-derived growth factor type BB (PDGF-BB) treatment elevated the level of VSMC ILF3 in a dose- and time-dependent manner. ILF3 silencing markedly inhibited PDGF-BB-induced phenotype switching, proliferation, and migration in VSMCs. Transcriptome sequencing and RNA immunoprecipitation sequencing depicted that ILF3 maintained its stability upon binding to the mRNA of the high-mobility group box 1 protein (HMGB1), thereby exerting an inhibitory effect on the transcription of dual specificity phosphatase 16 (DUSP16) through enhanced phosphorylation of signal transducer and activator of transcription 3 (STAT3). Therefore, the results both in vitro and in vivo indicated that the loss of ILF3 in VSMC ameliorated neointimal hyperplasia by regulating the STAT3/DUSP16 axis through the degradation of HMGB1 mRNA. Our findings revealed that vascular injury activates VSMC ILF3, which in turn promotes intima formation. Consequently, targeting specific VSMC ILF3 may present a potential therapeutic strategy for ameliorating cardiovascular restenosis.

Improved therapeutic effects on vascular intimal hyperplasia by mesenchymal stem cells expressing MIR155HG that function as a ceRNA for microRNA-205.

Coronary artery bypass grafting (CABG) is an effective treatment for coronary heart disease, with vascular transplantation as the key procedure. Intimal hyperplasia (IH) gradually leads to vascular stenosis, seriously affecting the curative effect of CABG. Mesenchymal stem cells (MSCs) were used to alleviate IH, but the effect was not satisfactory. This work aimed to investigate whether lncRNA MIR155HG could improve the efficacy of MSCs in the treatment of IH and to elucidate the role of the competing endogenous RNA (ceRNA). The effect of MIR155HG on MSCs function was investigated, while the proteins involved were assessed. IH was detected by HE and Van Gieson staining. miRNAs as the target of lncRNA were selected by bioinformatics analysis. qRT-PCR and dual-luciferase reporter assay were performed to verify the binding sites of lncRNA-miRNA. The apoptosis, Elisa and tube formation assay revealed the effect of ceRNA on the endothelial protection of MIR155HG-MSCs. We observed that MIR155HG improved the effect of MSCs on IH by promoting viability and migration. MIR155HG worked as a sponge for miR-205. MIR155HG/miR-205 significantly improved the function of MSCs, avoiding apoptosis and inducing angiogenesis. The improved therapeutic effects of MSCs on IH might be due to the ceRNA role of MIR155HG/miR-205.

Prevention of neointimal hyperplasia after coronary artery bypass graft via local delivery of sirolimus and rosuvastatin: network pharmacology and in vivo validation.

BACKGROUND: Coronary artery bypass graft (CABG) is generally used to treat complex coronary artery disease. Treatment success is affected by neointimal hyperplasia (NIH) of graft and anastomotic sites. Although sirolimus and rosuvastatin individually inhibit NIH progression, the efficacy of combination treatment remains unknown. METHODS: We identified cross-targets associated with CABG, sirolimus, and rosuvastatin by using databases including DisGeNET and GeneCards. GO and KEGG pathway enrichment analyses were conducted using R studio, and target proteins were mapped in PPI networks using Metascape and Cytoscape. For in vivo validation, we established a balloon-injured rabbit model by inducing NIH and applied a localized perivascular drug delivery device containing sirolimus and rosuvastatin. The outcomes were evaluated at 1, 2, and 4 weeks post-surgery. RESULTS: We identified 115 shared targets between sirolimus and CABG among databases, 23 between rosuvastatin and CABG, and 96 among all three. TNF, AKT1, and MMP9 were identified as shared targets. Network pharmacology predicted the stages of NIH progression and the corresponding signaling pathways linked to sirolimus (acute stage, IL6/STAT3 signaling) and rosuvastatin (chronic stage, Akt/MMP9 signaling). In vivo experiments demonstrated that the combination of sirolimus and rosuvastatin significantly suppressed NIH progression. This combination treatment also markedly decreased the expression of inflammation and Akt signaling pathway-related proteins, which was consistent with the predictions from network pharmacology analysis. CONCLUSIONS: Sirolimus and rosuvastatin inhibited pro-inflammatory cytokine production during the acute stage and regulated Akt/mTOR/NF-κB/STAT3 signaling in the chronic stage of NIH progression. These potential synergistic mechanisms may optimize treatment strategies to improve long-term patency after CABG.

Deficiency of thrombospondin-2 alleviates intimal hyperplasia in mice by modulating vascular smooth muscle cell proliferation and migration.

Thrombospondin-2 (Tsp2), a glycoprotein in the extracellular matrix, plays a critical role in the maintenance of vascular homeostasis. However, its role in the pathogenesis of cardiovascular disorders such as intimal hyperplasia is not fully elucidated. This study, therefore, aims to explore the effect of Tsp2 on intimal hyperplasia and its associated underlying mechanisms. Intimal hyperplasia (IH) was established using a modified wire-mediated femoral artery injury model. Immunofluorescence and qPCR identified upregulated Tsp2 expression in the injured femoral artery compared with the uninjured femoral artery. Similarly, TSP2 expression was also increased in human samples from the atherosclerotic femoral artery and colocalized with vascular smooth muscle cells (VSMCs). Compared with the wild-type littermates, Tsp2 knockout mice displayed a mitigated IH in the injured femoral artery, as demonstrated by a decreased neointimal area and intimal/median ratio. Primary mouse VSMCs were cultured to explore the mechanism by which Tsp2 influenced IH in vitro. PDGF-stimulated VSMCs presented an elevated Tsp2 expression and enhanced migration and proliferation. However, Tsp2 knockdown by siRNA blocked the increased migration and proliferation of VSMCs. Further analysis identified an association between Notch3 and IH when the intracellular domain of Notch3 (Nicd3) was upregulated in PDGF-stimulated VSMCs and femoral arteries with IH in human tissues. Along with the overexpression and downregulation of Tsp2, the Nicd3 expression was also up and downregulated accordingly. Tsp2 was associated with IH and may serve as a therapeutic target for IH. Downregulation of Tsp2 could mitigate the progression of IH by modulating the proliferation and migration of VSMCs.

Dihydroartemisinin inhibits restenosis after balloon angioplasty via circHSPA4/miR-19a-5p axis.

Dihydroartemisinin (DHA) inhibits restenosis following balloon angioplasty. However, data on the mechanisms of DHA activity in restenosis remains scant. Here, we investigated the role of circRNAs in mediating the inhibitory activity of DHA in neointimal formation. We used total RNA sequencing data to profile the expression of mRNA, circRNA and small RNA in sham, vascular balloon injury (VBI) and DHA-treated groups. CCK8 and EdU assays were employed to analyze cell proliferation, while qRT-PCR and Western blot were used to analyze the RNA or protein expression. In addition, we used RNA immunoprecipitation and luciferase reporter assay to assess the binding of circHSPA4 with miR-19a-5p. RNA sequencing demonstrated that circHSPA4 was upregulated in VBI. Treatment with DHA effectively suppressed the upregulation of the circHSPA4. In addition, analysis of platelet-derived growth family factor bb (PDGFbb)-induced HA-VSMCs showed upregulation of circHSPA4, which was associated with cell proliferation and differentiation. CircHSPA4 was shown to induce dedifferentiation and proliferation of smooth muscle cells. PDGFBB-induced overexpression of CircHSPA4 in HA-VSMCs led to suppression of miR-19a-5p, a phenomenon that was reversed by DHA, in concentration-dependent fashion. In addition, miR-19a-5p reduced the dedifferentiation and proliferation of the smooth muscle cells. Our data demonstrated that CircHSPA4 regulates proliferation and differentiation of smooth muscle cells. DHA and miR-19a-5p modulates CircHSPA4 and can be used as coated drugs on balloon catheter to improve the success rate of vascular remodeling.

Tumor necrosis factor receptor-associated factor 5 protects against intimal hyperplasia by regulation of macrophage polarization via directly targeting PPARγ.

OBJECTIVES: Intimal hyperplasia is a serious clinical problem associated with the failure of therapeutic methods in multiple atherosclerosis-related coronary heart diseases, which are initiated and aggravated by the polarization of infiltrating macrophages. The present study aimed to determine the effect and underlying mechanism by which tumor necrosis factor receptor-associated factor 5 (TRAF5) regulates macrophage polarization during intimal hyperplasia. METHODS: TRAF5 expression was detected in mouse carotid arteries subjected to wire injury. Bone marrow-derived macrophages, mouse peritoneal macrophages and human myeloid leukemia mononuclear cells were also used to test the expression of TRAF5 in vitro. Bone marrow-derived macrophages upon to LPS or IL-4 stimulation were performed to examine the effect of TRAF5 on macrophage polarization. TRAF5-knockout mice were used to evaluate the effect of TRAF5 on intimal hyperplasia. RESULTS: TRAF5 expression gradually decreased during neointima formation in carotid arteries in a time-dependent manner. In addition, the results showed that TRAF5 expression was reduced in classically polarized macrophages (M1) subjected to LPS stimulation but was increased in alternatively polarized macrophages (M2) in response to IL-4 administration, and these changes were demonstrated in three different types of macrophages. An in vitro loss-of-function study with TRAF5 knockdown plasmids or TRAF5-knockout mice revealed high expression of markers associated with M1 macrophages and reduced expression of genes related to M2 macrophages. Subsequently, we incubated vascular smooth muscle cells with conditioned medium of polarized macrophages in which TRAF5 expression had been downregulated or ablated, which promoted the proliferation, migration and dedifferentiation of VSMCs. Mechanistically, TRAF5 knockdown inhibited the activation of anti-inflammatory M2 macrophages by directly inhibiting PPARγ expression. More importantly, TRAF5-deficient mice showed significantly aggressive intimal hyperplasia. CONCLUSIONS: Collectively, this evidence reveals an important role of TRAF5 in the development of intimal hyperplasia through the regulation of macrophage polarization, which provides a promising target for arterial restenosis-related disease management.

Spatial Transcriptomic Profiling of Human Saphenous Vein Exposed to Ex Vivo Arterial Haemodynamics-Implications for Coronary Artery Bypass Graft Patency and Vein Graft Disease.

Vein graft disease is the process by which saphenous vein grafts, utilised for revascularisation during coronary artery bypass graft surgery, undergo an inflammation-driven intimal hyperplasia and accelerated atherosclerosis process in subsequent years after implantation. The role of the arterial circulation, particularly the haemodynamic properties' impact on graft patency, have been investigated but have not to date been explored in depth at the transcriptomic level. We have undertaken the first-in-man spatial transcriptomic analysis of the long saphenous vein in response to ex vivo acute arterial haemodynamic stimulation, utilising a combination of a custom 3D-printed perfusion bioreactor and the 10X Genomics Visium Spatial Gene Expression technology. We identify a total of 413 significant genes (372 upregulated and 41 downregulated) differentially expressed in response to arterial haemodynamic conditions. These genes were associated with pathways including NFkB, TNF, MAPK, and PI3K/Akt, among others. These are established pathways involved in the initiation of an early pro-inflammatory response, leukocyte activation and adhesion signalling, tissue remodelling, and cellular differentiation. Utilising unsupervised clustering analysis, we have been able to classify subsets of the expression based on cell type and with spatial resolution. These findings allow for further characterisation of the early saphenous vein graft transcriptional landscape during the earliest stage of implantation that contributes to vein graft disease, in particular validation of pathways and druggable targets that could contribute towards the therapeutic inhibition of processes underpinning vein graft disease.

Paricalcitol Has a Potent Anti-Inflammatory Effect in Rat Endothelial Denudation-Induced Intimal Hyperplasia.

Neointimal hyperplasia is the main cause of vascular graft failure in the medium term. Vitamin D receptor activation modulates the biology of vascular smooth muscle cells and has been reported to protect from neointimal hyperplasia following endothelial injury. However, the molecular mechanisms are poorly understood. We have now explored the impact of the selective vitamin D receptor activator, paricalcitol, on neointimal hyperplasia, following guidewire-induced endothelial cell injury in rats, and we have assessed the impact of paricalcitol or vehicle on the expression of key cell stress factors. Guidewire-induced endothelial cell injury caused neointimal hyperplasia and luminal stenosis and upregulated the expression of the growth factor growth/differentiation factor-15 (GDF-15), the cytokine receptor CD74, NFκB-inducing kinase (NIK, an upstream regulator of the proinflammatory transcription factor NFκB) and the chemokine monocyte chemoattractant protein-1 (MCP-1/CCL2). Immunohistochemistry confirmed the increased expression of the cellular proteins CD74 and NIK. Paricalcitol (administered in doses of 750 ng/kg of body weight, every other day) had a non-significant impact on neointimal hyperplasia and luminal stenosis. However, it significantly decreased GDF-15, CD74, NIK and MCP-1/CCL2 mRNA expression, which in paricalcitol-injured arteries remained within the levels found in control vehicle sham arteries. In conclusion, paricalcitol had a dramatic effect, suppressing the stress response to guidewire-induced endothelial cell injury, despite a limited impact on neointimal hyperplasia and luminal stenosis. This observation identifies novel molecular targets of paricalcitol in the vascular system, whose differential expression cannot be justified as a consequence of improved tissue injury.

CDKN2B-AS1 mediates proliferation and migration of vascular smooth muscle cells induced by insulin.

Excessive proliferation and migration of vascular smooth muscle cells (VSMCs) contribute to the intimal hyperplasia in type 2 diabetes mellitus (T2DM) patients after percutaneous coronary intervention. We aimed to investigate the role of lncRNA cyclin-dependent kinase inhibitor 2B antisense RNA 1 (CDKN2B-AS1) in VSMC proliferation and migration, as well as the underlying mechanism. T2DM model mice with carotid balloon injury were used in vivo and mouse aortic vascular smooth muscle cells (MOVAS) stimulated by insulin were used in vitro to assess the role of CDKN2B-AS1 in VSMC proliferation and migration following vascular injury in T2DM state. To investigate cell viability and migration, MTT assay and Transwell assay were conducted. To elucidate the underlying molecular mechanisms, the methylation-specific polymerase chain reaction, RNA immunoprecipitation, RNA-pull down, co-immunoprecipitation, and chromatin immunoprecipitation were performed. In vivo, CDKN2B-AS1 was up-regulated in common carotid artery tissues. In vitro, insulin treatment increased CDKN2B-AS1 level, enhanced MOVAS cell proliferation and migration, while the promoting effect was reversed by CDKN2B-AS1 knockdown. CDKN2B-AS1 forms a complex with enhancer of zeste homolog 2 (EZH2) and DNA methyltransferase (cytosine-5) 1 (DNMT1) to regulate smooth muscle 22 alpha (SM22α) methylation levels. In insulin-stimulated cells, SM22α knockdown abrogated the inhibitory effect of CDKN2B-AS1 knockdown on cell viability and migration. Injection of lentivirus-sh-CDKN2B-AS1 relieved intimal hyperplasia in T2DM mice with carotid balloon injury. Up-regulation of CDKN2B-AS1 induced by insulin promotes cell proliferation and migration by targeting SM22α through forming a complex with EZH2 and DNMT1, thereby aggravating the intimal hyperplasia after vascular injury in T2DM.

Exosomes from umbilical cord-derived mesenchymal stem cells combined with gelatin methacryloyl inhibit vein graft restenosis by enhancing endothelial functions.

BACKGROUND: The prevalence of coronary artery disease is increasing. As a common treatment method, coronary artery bypass transplantation surgery can improve heart problems while also causing corresponding complications. Venous graft restenosis is one of the most critical and intractable complications. Stem cell-derived exosomes could have therapeutic promise and value. However, as exosomes alone are prone to inactivation and easy removal, this therapeutic method has not been widely used in clinical practice. Methacrylated gelatin (GelMA) is a polymer with a loose porous structure that maintains the biological activity of the exosome and can control its slow release in vivo. In this study, we combined human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-Exos) and GelMA to explore their effects and underlying mechanisms in inhibiting venous graft restenosis. RESULTS: Human umbilical cord mesenchymal stem cells (hUCMSCs) were appraised using flow cytometry. hUCMSC-Exos were evaluated via transmission electron microscopy and western blotting. hUCMSC-Exos embedded in a photosensitive GelMA hydrogel (GelMA-Exos) were applied topically around venous grafts in a rat model of cervical arteriovenous transplantation, and their effects on graft reendothelialization and restenosis were evaluated through ultrasonic, histological, and immunofluorescence examinations. Additionally, we analyzed the material properties, cellular reactions, and biocompatibility of the hydrogels. We further demonstrated that the topical application of GelMA-Exos could accelerate reendothelialization after autologous vein transplantation and reduce restenosis in the rat model. Notably, GelMA-Exos caused neither damage to major organs in mice nor excessive immune rejection. The uptake of GelMA-Exos by endothelial cells stimulated cell proliferation and migration in vitro. A bioinformatic analysis of existing databases revealed that various cell proliferation and apoptosis pathways, including the mammalian target of rapamycin (mTOR)-phosphoinositide 3-kinase (PI3K)-AKT signaling pathways, might participate in the underlying regulatory mechanism. CONCLUSIONS: Compared with the tail vein injection of hUCMSC-Exos, the local application of a mixture of hUCMSC-Exos and GelMA was more effective in promoting endothelial repair of the vein graft and inhibiting restenosis. Therefore, the proposed biomaterial-based therapeutic approach is a promising treatment for venous graft restenosis.

Prevalence and risk factors of phlebosclerosis in the great saphenous vein.

OBJECTIVES: Phlebosclerosis is a fibrous degeneration of the vein wall, predominantly the intima, with or without calcification. The prevalence and etiology of phlebosclerosis of the great saphenous vein are not well documented. This study aimed to estimate the prevalence and define the risk factors of phlebosclerosis of the great saphenous vein. METHODS: The study was conducted on 300 volunteers who underwent duplex ultrasound. Volunteers with symptoms and signs of acute or chronic venous disease or known varicose veins, thrombosis, chronic vein insufficiency, and any operation in the lower extremities were excluded. The imaging hallmarks of phlebosclerosis include wall brightness, calcification, and increased wall thickness. Demographics of the volunteers (sex, age, weight, and height), Body Mass Index (BMI) and the presence of smoking, hypertension, diabetes mellitus, and dyslipidemia were recorded. Data obtained were consolidated and statistically evaluated using SPSS Version 16. RESULTS: Of the 300 volunteers who underwent duplex ultrasound, 60.3% were females, and 39.7% were males. The mean age was 60 ± 13, while the mean BMI was 26.01 ± 4.76. Moreover, 66.3% were non-smokers, and 62.3%, 81.3%, and 58.7% did not suffer from hypertension, diabetes mellitus, and dyslipidemia, respectively. The prevalence of phlebosclerosis was found to be 2.3%. Hypertension was a risk factor for the development of phlebosclerosis (p = 0.045). Moreover, there was a link between phlebosclerosis and age, as volunteers with phlebosclerosis were older than volunteers without phlebosclerosis (74.2 vs 59.11 years, p < 0.001). CONCLUSIONS: The prevalence of phlebosclerosis of the great saphenous vein is low, specifically 2.3%. Hypertension and increased age are risk factors for the development of phlebosclerosis. Both sexes are equally affected, while BMI, smoking, diabetes mellitus, and dyslipidemia do not contribute to the development of phlebosclerosis.

The effect of cilostazol on the platelet-derived growth factor-beta/beta isoform reduction on venous hyperplasia in an experimental balloon-induced injury model.

BACKGROUND: Intimal hyperplasia is the response to endothelial injury. Platelet-derived growth factor is released early and favors the formation of intimal hyperplasia. Although multiple treatments, from open surgery to endovascular techniques, have been used they remain controversial. There is currently interest in developing pharmacological strategies to address this pathology. Local vascular inflammation induced by vessel barotrauma generates intimal hyperplasia due to mechanical stress over the venous endothelium. Cilostazol is a selective phosphodiesterase type 3 (PDE3) selective inhibitor with a regulatory effect over intimal hyperplasia. The objective was to investigate cilostazol's role in inhibiting smooth muscle cell proliferation due to changes in the expression and release of PDGF-BB isoform and the effect on developing IH using an experimental model of vascular barotrauma (balloon-induced injury model). METHODS: We included 12 New Zealand rabbits. The balloon-induced injury model (BIIM) and experimental group cilostazol (20 mg/kg/day) included 6 rabbits each. Contralateral veins from 6 rabbits used in BIIM model has been taken as control group. We measured and compared the expression of PDGF-BB and the development of IH. A pathologist board chooses a PDGFRα antibody to localized its expression by immunohistochemistry analysis. Subsequently, using an automated immunohistochemical staining machine, the PDGFR expression was evaluated using a Zeiss Primo Star 4 light microscope. RESULTS: The measurement obtained in the intimal layer was: 126.12 µm2 in the CG, 232 µm2 in the BIIM group, and 178 µm2 in the EG. A statistically significant difference was observed. Baseline serum concentrations of PDGF-BB in the BIIM group were 0.22 pg/mL. At 12 h 0.42 pg/mL, and 0.17 pg/mL at seven days. In the experimental group, the basal levels were 0.33 pg/mL. With the use of cilostazol, a lower peak was obtained at 12 h (0.08 pg/mL). This difference was statistically significant. CONCLUSIONS: Cilostazol induced a significant reduction of IH caused by barotrauma in the venous endothelium, which correlates with decrease in the PDGF-BB in serum. This could be attributed to the pharmacologic effect on PDGFR expression.

Inhibition of mitochondrial fission alters neo-intimal hyperplasia via PI3K/Akt signaling in arteriovenous fistulas.

BACKGROUND/OBJECTIVE: Arteriovenous fistulas (AVFs) are the preferred vascular access for hemodialysis of patients with end-stage renal disease. However, there is a high incidence of AVF failures caused by insufficient outward remodeling or venous neo-intimal hyperplasia formation. Abnormal proliferation and migration of vascular smooth muscle cells (VSMCs) play an important role in many cardiovascular diseases. Abnormal VSMC proliferation and migration could be abolished by inhibition of mitochondrial division. METHOD: We found that abnormal proliferation and migration of VSMCs and increased mitochondrial fission were associated with AVF stenosis in patients. We also investigated the mechanisms, particularly the role of mitochondrial dynamics, underlying these VSMC behaviors. In vitro, we observed that inhibition of mitochondrial fission and Akt phosphorylation can diminish proliferation and migration of VSMCs induced by platelet-derived growth factor-BB (PDGF-BB). In vivo, daily intraperitoneal injections of mitochondrial division inhibitor 1 (Mdivi-1) decreased VSMC proliferation and reduced AVF wall thickness in a rat AVF model. CONCLUSION AND RESULT: Our results suggest that inhibition of mitochondrial fission improves AVF patency by reducing wall thickening through the PI3K/Akt signaling pathway. Therefore, inhibition of mitochondrial fission has the clinical potential to improve AVF patency.

Large animal model of vein grafts intimal hyperplasia: A systematic review.

Coronary artery bypass grafting remains the treatment of choice for a large cohort of patients with significant coronary disease. Despite the increased use of arterial grafts, the long saphenous vein remains the most commonly used conduit. Long-term graft patency continues to be the Achilles heel of saphenous vein grafts. This is due to the development of intimal hyperplasia, a chronic inflammatory disease that results in the narrowing and occlusion of a significant number of vein grafts. Research models for intimal hyperplasia are essential for a better understanding of pathophysiological processes of this condition. Large animal models resemble human anatomical structures and have been used as a surrogate to study disease development and prevention over the years. In this paper, we systematically review all published studies that utilized large animal models of vein graft disease with a focus on the type of model and any therapeutic intervention, specifically the use of external stents/mesh.

Clinical Potential of Hydrogen Sulfide in Peripheral Arterial Disease.

Peripheral artery disease (PAD) affects more than 230 million people worldwide. PAD patients suffer from reduced quality of life and are at increased risk of vascular complications and all-cause mortality. Despite its prevalence, impact on quality of life and poor long-term clinical outcomes, PAD remains underdiagnosed and undertreated compared to myocardial infarction and stroke. PAD is due to a combination of macrovascular atherosclerosis and calcification, combined with microvascular rarefaction, leading to chronic peripheral ischemia. Novel therapies are needed to address the increasing incidence of PAD and its difficult long-term pharmacological and surgical management. The cysteine-derived gasotransmitter hydrogen sulfide (H2S) has interesting vasorelaxant, cytoprotective, antioxidant and anti-inflammatory properties. In this review, we describe the current understanding of PAD pathophysiology and the remarkable benefits of H2S against atherosclerosis, inflammation, vascular calcification, and other vasculo-protective effects.

The PPAR-γ Agonist Pioglitazone Modulates Proliferation and Migration in HUVEC, HAOSMC and Human Arteriovenous Fistula-Derived Cells.

The failure of arteriovenous fistulas (AVFs) following intimal hyperplasia (IH) increases morbidity and mortality rates in patients undergoing hemodialysis for chronic kidney disease. The peroxisome-proliferator associated receptor (PPAR-γ) may be a therapeutic target in IH regulation. In the present study, we investigated PPAR-γ expression and tested the effect of pioglitazone, a PPAR-γ agonist, in different cell types involved in IH. As cell models, we used Human Endothelial Umbilical Vein Cells (HUVEC), Human Aortic Smooth Muscle Cells (HAOSMC), and AVF cells (AVFCs) isolated from (i) normal veins collected at the first AVF establishment (T0), and (ii) failed AVF with IH (T1). PPAR-γ was downregulated in AVF T1 tissues and cells, in comparison to T0 group. HUVEC, HAOSMC, and AVFC (T0 and T1) proliferation and migration were analyzed after pioglitazone administration, alone or in combination with the PPAR-γ inhibitor, GW9662. Pioglitazone negatively regulated HUVEC and HAOSMC proliferation and migration. The effect was antagonized by GW9662. These data were confirmed in AVFCs T1, where pioglitazone induced PPAR-γ expression and downregulated the invasive genes SLUG, MMP-9, and VIMENTIN. In summary, PPAR-γ modulation may represent a promising strategy to reduce the AVF failure risk by modulating cell proliferation and migration.

Transfection of Vein Grafts with Early Growth Response Factor-1 Oligodeoxynucleotide Decoy: Effects on Stem-Cell Genes and Toll-like Receptor-Mediated Inflammation.

The long-term patency of vein grafts is challenged by intimal hyperplasia. We sought to explore the intricate relationships between the transcription factor Egr-1, toll-like receptors (TLRs), and stem cell genes and also assessed oligodeoxynucleotide decoys (ODNs) as a strategy to prevent vein graft failures. A total of 42 New Zealand white rabbits were fed hyperlipidemic chow and classified into three groups. A double-stranded Egr-1 ODN was synthesized and infused in vein grafts prior to anastomosis with the common carotid artery. All vein grafts were retrieved at the conclusion of the predefined experimental period. Real-time quantitative polymerase chain reaction was performed to estimate expression patterns for several genes of interest. MYD88, TLR2-4, TLR8, NF-kB, TNF-α, IFNß, and IFNγ; chemokines CCL4, CCL20, CCR2; numerous interleukins; and stem cell genes KFL4, NANOG, HOXA5, and HIF1α were universally downregulated in the ODN arm compared with the controls. By understanding these multifaceted interactions, our study offers actionable insights that may pave the way for innovative interventions in vascular reconstructions.

Mechanistic exploration of Yiqi Liangxue Shengji prescription on restenosis after balloon injury by integrating metabolomics with network pharmacology.

CONTEXT: Yiqi Liangxue Shengji prescription (YQLXSJ) is a traditional Chinese medicine (TCM) formula that has long been used for treatment after percutaneous coronary intervention (PCI). OBJECTIVE: To investigate the putative pharmacological mechanism of YQLXSJ on restenosis through an integrated approach utilizing metabolomics and network pharmacology. MATERIALS AND METHODS: Forty male Sprague-Dawley rats were divided into sham, model, YQLXSJ, and positive groups. YQLXSJ group received the treatment of YQLXSJ (6 g/kg/d, i.g.) and the positive group was treated with atorvastatin (2 mg/kg/d, i.g.). After 4 weeks, the improvement in intimal hyperplasia was evaluated by ultrasound, H&E staining, and immunofluorescence. UPLC-MS/MS technology was utilized to screen the differential metabolites. Network pharmacology was conducted using TCMSP, GeneCards, and Metascape, etc., in combination with metabolomics. Eventually, the core targets were acquired and validated. RESULTS: Compared to models, YQLXSJ exhibited decreased intima-media thickness on ultrasound (0.23 ± 0.02 mm vs. 0.20 ± 0.01 mm, p < 0.01) and reduced intima thickness by H&E (30.12 ± 6.05 µm vs. 14.32 ± 1.37 µm, p < 0.01). We identified 18 differential metabolites and 5 core targets such as inducible nitric oxide synthase (NOS2), endothelial nitric oxide synthase (NOS3), vascular endothelial growth factor-A (VEGFA), ornithine decarboxylase-1 (ODC1) and group IIA secretory phospholipase A2 (PLA2G2A). These targets were further confirmed by molecular docking and ELISA. DISCUSSION AND CONCLUSIONS: This study confirms the effects of YQLXSJ on restenosis and reveals some biomarkers. TCM has great potential in the prevention and treatment of restenosis by improving metabolic disorders.

M6A methylation-mediated elevation of SM22α inhibits the proliferation and migration of vascular smooth muscle cells and ameliorates intimal hyperplasia in type 2 diabetes mellitus.

Abnormal proliferation of vascular smooth muscle cells (VSMCs) induced by insulin resistance facilitates intimal hyperplasia of type 2 diabetes mellitus (T2DM) and N6-methyladenosine (m6A) methylation modification mediates the VSMC proliferation. This study aimed to reveal the m6A methylation modification regulatory mechanism. In this study, m6A demethylase FTO was elevated in insulin-treated VSMCs and T2DM mice with intimal injury. Functionally, FTO knockdown elevated m6A methylation level and further restrained VSMC proliferation and migration induced by insulin. Mechanistically, FTO knockdown elevated Smooth muscle 22 alpha (SM22α) expression and m6A-binding protein IGF2BP2 enhanced SM22α mRNA stability by recognizing and binding to m6A methylation modified mRNA. In vivo studies confirmed that the elevated m6A modification level of SM22α mRNA mitigated intimal hyperplasia in T2DM mice. Conclusively, m6A methylation-mediated elevation of SM22α restrained VSMC proliferation and migration and ameliorated intimal hyperplasia in T2DM.

Activation of macrophage TBK1-HIF-1α-mediated IL-17/IL-10 signaling by hyperglycemia aggravates the complexity of coronary atherosclerosis: An in vivo and in vitro study.

Our purpose was to study the effect of hyperglycemia on macrophage TBK1-HIF-1α-mediated IL-17/IL-10 signaling and its correlation with coronary atherosclerosis. A total of 135 patients with coronary heart disease (CHD) were divided into a stable CHD (SCHD) group (n = 30) and an acute myocardial infarction (AMI) group (n = 105) [nondiabetes mellitus (non-DM)-AMI, n = 60; DM-AMI, n = 45] from January to September 2020. The SYNTAX score and metabolic and inflammatory markers were quantified and compared. THP-1 cell studies and an animal study of coronary intimal hyperplasia were also carried out. We found that the DM-AMI group showed a higher SYNTAX score than the non-DM-AMI group (P < .05). The DM-AMI group showed the highest expression levels of TANK-binding kinase 1 (TBK1), hypoxia-inducible factor 1α (HIF-1α), and interleukin (IL)-17 and the lowest expression level of IL-10, followed by the non-DM-AMI group and the SCHD group (P < .05). THP-1 cell studies showed that BAY87-2243 (a HIF-1α inhibitor) reversed the increase in IL-17 and decrease in IL-10 expression induced by hyperglycemia. Amlexanox (a TBK1 inhibitor) reversed the increase in HIF-1α expression induced by hyperglycemia. Amlexanox treatment resulted in lower coronary artery intimal hyperplasia and a larger lumen area in a diabetic swine model. We conclude that hyperglycemia might aggravate the complexity of coronary atherosclerosis through activation of TBK1-HIF-1α-mediated IL-17/IL-10 signaling. Thus, TBK1 may be a novel drug therapy target for CHD complicated with DM.