RESUMEN

Common arterial grafts used in coronary artery bypass grafting include internal thoracic artery (ITA), radial artery (RA) and right gastroepiploic artery (RGA) grafts; of these, the ITA has the best clinical outcome. Here, by analyzing the single-cell transcriptome of different arterial grafts, we suggest optimization strategies for the RA and RGA based on the ITA as a reference. Compared with the ITA, the RA had more lipid-handling-related CD36+ endothelial cells. Vascular smooth muscle cells from the RGA were more susceptible to spasm, followed by those from the RA; comparison with the ITA suggested that potassium channel openers may counteract vasospasm. Fibroblasts from the RA and RGA highly expressed GDF10 and CREB5, respectively; both GDF10 and CREB5 are associated with extracellular matrix deposition. Cell-cell communication analysis revealed high levels of macrophage migration inhibitory factor signaling in the RA. Administration of macrophage migration inhibitory factor inhibitor to mice with partial carotid artery ligation blocked neointimal hyperplasia induced by disturbed flow. Modulation of identified targets may have protective effects on arterial grafts.

Asunto(s)

Arterias Mamarias , Animales , Humanos , Arterias Mamarias/trasplante , Arterias Mamarias/metabolismo , Análisis de la Célula Individual , Arteria Radial/trasplante , Arteria Radial/metabolismo , Arteria Gastroepiploica/metabolismo , Arteria Gastroepiploica/trasplante , Miocitos del Músculo Liso/metabolismo , Masculino , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/citología , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Neointima/patología , Neointima/metabolismo , Puente de Arteria Coronaria/métodos , Comunicación Celular , Fibroblastos/metabolismo , Células Endoteliales/metabolismo , Ratones , Transducción de Señal , Transcriptoma , Vasoconstricción/efectos de los fármacos , Células Cultivadas , Hiperplasia/metabolismo , Hiperplasia/patología , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo

RESUMEN

Smooth muscle cell (SMC) phenotypic modulation, primarily driven by PDGFRß signaling, is implicated in occlusive cardiovascular diseases. However, the promotive and restrictive regulation mechanism of PDGFRß and the role of protein tyrosine phosphatase non-receptor type 14 (PTPN14) in neointimal hyperplasia remain unclear. Our study observes a marked upregulation of PTPN14 in SMCs during neointimal hyperplasia. PTPN14 overexpression exacerbates neointimal hyperplasia in a phosphatase activity-dependent manner, while SMC-specific deficiency of PTPN14 mitigates this process in mice. RNA-seq indicates that PTPN14 deficiency inhibits PDGFRß signaling-induced SMC phenotypic modulation. Moreover, PTPN14 interacts with intracellular region of PDGFRß and mediates its dephosphorylation on Y692 site. Phosphorylation of PDGFRßY692 negatively regulates PDGFRß signaling activation. The levels of both PTPN14 and phospho-PDGFRßY692 are correlated with the degree of stenosis in human coronary arteries. Our findings suggest that PTPN14 serves as a critical modulator of SMCs, promoting neointimal hyperplasia. PDGFRßY692, dephosphorylated by PTPN14, acts as a self-inhibitory site for controlling PDGFRß activation.

Asunto(s)

Hiperplasia , Miocitos del Músculo Liso , Neointima , Receptor beta de Factor de Crecimiento Derivado de Plaquetas , Transducción de Señal , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genética , Animales , Hiperplasia/metabolismo , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Humanos , Neointima/metabolismo , Neointima/patología , Ratones , Fosforilación , Masculino , Proteínas Tirosina Fosfatasas no Receptoras/metabolismo , Proteínas Tirosina Fosfatasas no Receptoras/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Vasos Coronarios/patología , Vasos Coronarios/metabolismo , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología

RESUMEN

Restenosis following percutaneous revascularization is a major challenge in patients with insulin resistance and diabetes. Currently, the vascular effects of insulin are not fully understood. In vitro, insulin's effects on endothelial cells (ECs) are beneficial, whereas on vascular smooth muscle cells (SMCs), they are mitogenic. We previously demonstrated a suppressive effect of insulin on neointimal growth under insulin-sensitive conditions that was abolished in insulin-resistant conditions. Here, we aimed to determine the cell-specific effects of insulin on neointimal growth in a model of restenosis under insulin-sensitive and insulin-resistant conditions. Vascular cell-specific insulin receptor (IR)-deficient mice were fed a low-fat diet (LFD) or a high-fat, high-sucrose diet (HFSD) and implanted with an insulin pellet or vehicle prior to femoral artery wire injury. In insulin-sensitive conditions, insulin decreased neointimal growth only in controls. However, under insulin-resistant conditions, insulin had no effect in either control, EC-specific or SMC-specific IR-deficient mice. These data demonstrate that EC and SMC IRs are required for the anti-restenotic effect of insulin in insulin-sensitive conditions and that, in insulin resistance, insulin has no adverse effect on vascular SMCs in vivo.

Asunto(s)

Modelos Animales de Enfermedad , Células Endoteliales , Resistencia a la Insulina , Insulina , Receptor de Insulina , Animales , Insulina/metabolismo , Insulina/farmacología , Ratones , Receptor de Insulina/metabolismo , Células Endoteliales/metabolismo , Células Endoteliales/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , Neointima/patología , Neointima/metabolismo , Masculino , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/patología , Ratones Endogámicos C57BL

RESUMEN

The proliferative and migratory abilities of vascular smooth muscle cells (VSMCs) play a crucial role in neointima formation following vascular injury. Skp2 facilitates proliferation and migration in cells through cell cycle regulation, presenting an important therapeutic target for atherosclerosis, pulmonary hypertension, and vascular restenosis. This study aimed to identify a natural product capable of inhibiting neointima formation post vascular injury. Here, we demonstrate that troxerutin, a flavonoid, significantly reduced viability and downregulated Skp2 in VSMCs. Moreover, troxerutin exhibited anti-proliferative effects on VSMCs and mitigated neointima formation. These findings collectively elucidate the intrinsic mechanism of troxerutin in treating atherosclerosis, pulmonary hypertension, and vascular restenosis by targeting the E3-linked enzyme Skp2.

Asunto(s)

Proliferación Celular , Hidroxietilrutósido , Músculo Liso Vascular , Neointima , Proteínas Quinasas Asociadas a Fase-S , Hidroxietilrutósido/análogos & derivados , Hidroxietilrutósido/farmacología , Proteínas Quinasas Asociadas a Fase-S/metabolismo , Proteínas Quinasas Asociadas a Fase-S/antagonistas & inhibidores , Neointima/tratamiento farmacológico , Neointima/patología , Neointima/metabolismo , Animales , Proliferación Celular/efectos de los fármacos , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/citología , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Proteolisis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Movimiento Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Ratas

RESUMEN

Serum response factor (SRF) controls gene transcription in vascular smooth muscle cells (VSMCs) and regulates VSMC phenotypic switch from a contractile to a synthetic state, which plays a key role in the pathogenesis of cardiovascular diseases (CVD). It is not known how post-translational SUMOylation regulates the SRF activity in CVD. Here we show that Senp1 deficiency in VSMCs increased SUMOylated SRF and the SRF-ELK complex, leading to augmented vascular remodeling and neointimal formation in mice. Mechanistically, SENP1 deficiency in VSMCs increases SRF SUMOylation at lysine 143, reducing SRF lysosomal localization concomitant with increased nuclear accumulation and switching a contractile phenotype-responsive SRF-myocardin complex to a synthetic phenotype-responsive SRF-ELK1 complex. SUMOylated SRF and phospho-ELK1 are increased in VSMCs from coronary arteries of CVD patients. Importantly, ELK inhibitor AZD6244 prevents the shift from SRF-myocardin to SRF-ELK complex, attenuating VSMC synthetic phenotypes and neointimal formation in Senp1-deficient mice. Therefore, targeting the SRF complex may have a therapeutic potential for the treatment of CVD.

Asunto(s)

Músculo Liso Vascular , Miocitos del Músculo Liso , Proteínas Nucleares , Fenotipo , Factor de Respuesta Sérica , Sumoilación , Remodelación Vascular , Animales , Humanos , Masculino , Ratones , Enfermedades Cardiovasculares/metabolismo , Enfermedades Cardiovasculares/patología , Enfermedades Cardiovasculares/genética , Cisteína Endopeptidasas/metabolismo , Cisteína Endopeptidasas/genética , Proteína Elk-1 con Dominio ets/metabolismo , Proteína Elk-1 con Dominio ets/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismo , Neointima/metabolismo , Neointima/patología , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Factor de Respuesta Sérica/metabolismo , Factor de Respuesta Sérica/genética , Transactivadores/metabolismo , Transactivadores/genética

RESUMEN

The roles and mechanisms of A-kinase anchoring protein 1 (AKAP1) in vascular smooth muscle cell (VSMC) phenotypic modulation and neointima formation are currently unknown. AKAP1 is a mitochondrial PKA-anchored protein and maintains mitochondrial homeostasis. This study aimed to investigate how AKAP1/PKA signaling plays a protective role in inhibiting VSMC phenotypic transformation and neointima formation by regulating mitochondrial fission. The results showed that both PDGF-BB treatment and balloon injury reduced the transcription, expression, and mitochondrial anchoring of AKAP1. In vitro, the overexpression of AKAP1 significantly inhibited PDGF-BB mediated VSMC proliferation and migration, whereas AKAP1 knockdown further aggravated VSMC phenotypic transformation. Additionally, in the balloon injury model in vivo, AKAP1 overexpression reduced neointima formation, the muscle fiber area ratio, and rat VSMC proliferation and migration. Furthermore, PDGF-BB and balloon injury inhibited Drp1 phosphorylation at Ser637 and promoted Drp1 activity and mitochondrial midzone fission; AKAP1 overexpression reversed these effects. AKAP1 overexpression also inhibited the distribution of mitochondria at the plasma membrane and the reduction of PKARIIß expression induced by PDGF-BB, as evidenced by an increase in mitochondria-plasma membrane distance as well as PKARIIß protein levels. Moreover, the PKA agonist promoted Drp1 phosphorylation (Ser637) and inhibited PDGF-BB-mediated mitochondrial fission, cell proliferation, and migration. The PKA antagonist reversed the increase in Drp1 phosphorylation (Ser637) and the decline in mitochondrial midzone fission and VSMC phenotypic transformation caused by AKAP1 overexpression. The results of this study reveal that AKAP1 protects VSMCs against phenotypic modulation by improving Drp1 phosphorylation at Ser637 through PKA and inhibiting mitochondrial fission, thereby preventing neointima formation.

Asunto(s)

Proteínas de Anclaje a la Quinasa A , Dinaminas , Músculo Liso Vascular , Neointima , Animales , Masculino , Ratas , Proteínas de Anclaje a la Quinasa A/metabolismo , Proteínas de Anclaje a la Quinasa A/genética , Becaplermina/farmacología , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Dinaminas/metabolismo , Dinámicas Mitocondriales/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Músculo Liso Vascular/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/patología , Neointima/metabolismo , Neointima/patología , Fenotipo , Fosforilación , Ratas Sprague-Dawley , Transducción de Señal

RESUMEN

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.

Asunto(s)

Antiinflamatorios , Quimiocina CCL2 , Ergocalciferoles , Hiperplasia , Animales , Ratas , Ergocalciferoles/farmacología , Masculino , Quimiocina CCL2/metabolismo , Quimiocina CCL2/genética , Antiinflamatorios/farmacología , Neointima/metabolismo , Neointima/patología , Neointima/tratamiento farmacológico , Factor 15 de Diferenciación de Crecimiento/metabolismo , Factor 15 de Diferenciación de Crecimiento/genética , Túnica Íntima/patología , Túnica Íntima/efectos de los fármacos , Túnica Íntima/metabolismo , Antígenos de Diferenciación de Linfocitos B/metabolismo , Antígenos de Diferenciación de Linfocitos B/genética , Células Endoteliales/metabolismo , Células Endoteliales/efectos de los fármacos , Antígenos de Histocompatibilidad Clase II

RESUMEN

BACKGROUND: Neointimal hyperplasia (NIH) is the pathological basis of vascular injury disease. Vascular cells are the dominant cells in the process of NIH, but the extent of heterogeneity amongst them is still unclear. METHODS: A mouse model of NIH was constructed by inducing carotid artery ligation. Single-cell sequencing was then used to analyze the transcriptional profile of vascular cells. Cluster features were determined by functional enrichment analysis, gene set scoring, pseudo-time analysis, and cell-cell communication analysis. Additionally, immunofluorescence staining was conducted on vascular tissues from fibroblast lineage-traced (PdgfraDreER-tdTomato) mice to validate the presence of Pecam1+Pdgfra+tdTomato+ cells. RESULTS: The left carotid arteries (ligation) were compared to right carotid arteries (sham) from ligation-induced NIH C57BL/6 mice. Integrative analyses revealed a high level of heterogeneity amongst vascular cells, including fourteen clusters and seven cell types. We focused on three dominant cell types: endothelial cells (ECs), vascular smooth muscle cells (vSMCs), and fibroblasts. The major findings were: (1) four subpopulations of ECs, including ECs4, mesenchymal-like ECs (ECs1 and ECs2), and fibro-like ECs (ECs3); (2) four subpopulations of fibroblasts, including pro-inflammatory Fibs-1, Sca1+ Fibs-2, collagen-producing Fibs-3, and mesenchymal-like Fibs-4; (3) four subpopulations of vSMCs, including vSMCs-1, vSMCs-2, vSMCs-3, and vSMCs-3-derived vSMCs; (4) ECs3 express genes related to extracellular matrix (ECM) remodeling and cell migration, and fibro-like vSMCs showed strong chemokine secretion and relatively high levels of proteases; (5) fibro-like vSMCs that secrete Vegfa interact with ECs mainly through vascular endothelial growth factor receptor 2 (Vegfr2). CONCLUSIONS: This study presents the dynamic cellular landscape within NIH arteries and reveals potential relationships between several clusters, with a specific focus on ECs3 and fibro-like vSMCs. These two subpopulations may represent potential target cells for the treatment of NIH.

Asunto(s)

Perfilación de la Expresión Génica , Hiperplasia , Ratones Endogámicos C57BL , Músculo Liso Vascular , Neointima , Análisis de la Célula Individual , Animales , Neointima/patología , Neointima/metabolismo , Neointima/genética , Análisis de la Célula Individual/métodos , Hiperplasia/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Músculo Liso Vascular/citología , Ratones , Células Endoteliales/metabolismo , Células Endoteliales/patología , Arterias Carótidas/patología , Arterias Carótidas/metabolismo , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Masculino , Fibroblastos/metabolismo , Fibroblastos/patología , Modelos Animales de Enfermedad , Análisis de Expresión Génica de una Sola Célula

RESUMEN

Arteriovenous fistulas (AVFs) are the most common vascular access points for hemodialysis (HD), but they have a high incidence of postoperative dysfunction, mainly due to excessive neointimal hyperplasia (NIH). Our previous studies have revealed a highly conserved LncRNA-LncDACH1 as an important regulator of cardiomyocyte and fibroblast proliferation. Herein, we find that LncDACH1 regulates NIH in AVF in male mice with conditional knockout of smooth muscle cell-specific LncDACH1 and in male mice model of AVF with LncDACH1 overexpression by adeno-associated virus. Mechanistically, silence of LncDACH1 activates p-AKT through promoting the expression of heat shock protein 90 (HSP90) and serine/arginine-rich splicing factor protein kinase 1 (SRPK1). Moreover, LncDACH1 is transcriptionally activated by transcription factor KLF9 that binds directly to the promoter region of the LncDACH1 gene. In this work, during AVF NIH, LncDACH1 is downregulated by KLF9 and promotes NIH through the HSP90/ SRPK1/ AKT signaling axis.

Asunto(s)

Proteínas HSP90 de Choque Térmico , Hiperplasia , Factores de Transcripción de Tipo Kruppel , Miocitos del Músculo Liso , Neointima , Proteínas Proto-Oncogénicas c-akt , ARN Largo no Codificante , Animales , Humanos , Masculino , Ratones , Fístula Arteriovenosa/metabolismo , Fístula Arteriovenosa/genética , Fístula Arteriovenosa/patología , Proliferación Celular , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Neointima/patología , Neointima/metabolismo , Fenotipo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Transducción de Señal

RESUMEN

Intimal hyperplasia (IH) is a common pathological feature of vascular proliferative diseases, such as atherosclerosis and restenosis after angioplasty. Urotensin II (UII) and its receptor (UTR) are widely expressed in cardiovascular tissues. However, it remains unclear whether the UII/UTR system is involved in IH. Right unilateral common carotid artery ligation was performed and maintained for 21 days to induce IH in UTR knockout (UTR-/-) and wild-type (WT) mice. Histological analysis revealed that compared with WT mice, UTR-deficient mice exhibited a decreased neointimal area, angiostenosis and intima-media ratio. Immunostaining revealed fewer smooth muscle cells (SMCs), endothelial cells and macrophages in the lesions of UTR-/- mice than in those of WT mice. Protein interaction analysis suggested that the UTR may affect cell proliferation by regulating YAP and its downstream target genes. In vitro experiments revealed that UII can promote the proliferation and migration of SMCs, and western blotting also revealed that UII increased the protein expression of RhoA, CTGF, Cyclin D1 and PCNA and downregulated p-YAP protein expression, while these effects could be partly reversed by urantide. To evaluate the translational value of UTRs in IH management, WT mice were also treated with two doses of urantide, a UTR antagonist, to confirm the benefit of UTR blockade in IH progression. A high dose of urantide (600 µg/kg/day), rather than a low dose (60 µg/kg/day), successfully improved ligation-induced IH compared with that in mice receiving vehicle. The results of the present study suggested that the UII/UTR system may regulate IH partly through the RhoA-YAP signaling pathway.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales , Proliferación Celular , Hiperplasia , Ratones Noqueados , Receptores Acoplados a Proteínas G , Transducción de Señal , Proteínas Señalizadoras YAP , Proteína de Unión al GTP rhoA , Animales , Masculino , Ratones , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Movimiento Celular , Hiperplasia/metabolismo , Hiperplasia/patología , Ligadura , Ratones Endogámicos C57BL , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Neointima/metabolismo , Neointima/patología , Neointima/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Proteína de Unión al GTP rhoA/metabolismo , Proteína de Unión al GTP rhoA/genética , Túnica Íntima/patología , Túnica Íntima/metabolismo , Urotensinas/metabolismo , Urotensinas/genética , Urotensinas/farmacología , Proteínas Señalizadoras YAP/metabolismo

RESUMEN

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.

Asunto(s)

Hiperplasia , Macrófagos , Ratones Endogámicos C57BL , Ratones Noqueados , PPAR gamma , Factor 5 Asociado a Receptor de TNF , Animales , Macrófagos/metabolismo , Factor 5 Asociado a Receptor de TNF/genética , Factor 5 Asociado a Receptor de TNF/metabolismo , PPAR gamma/metabolismo , PPAR gamma/genética , Masculino , Ratones , Humanos , Arterias Carótidas/patología , Neointima/patología , Neointima/metabolismo , Interleucina-4/genética , Células Cultivadas , Túnica Íntima/patología , Lipopolisacáridos/farmacología

RESUMEN

Krüppel-like factor 13 (KLF13), a zinc finger transcription factor, is considered as a potential regulator of cardiomyocyte differentiation and proliferation during heart morphogenesis. However, its precise role in the dedifferentiation of vascular smooth muscle cells (VSMCs) during atherosclerosis and neointimal formation after injury remains poorly understood. In this study, we investigated the relationship between KLF13 and SM22α expression in normal and atherosclerotic plaques by bioanalysis, and observed a significant increase in KLF13 levels in the atherosclerotic plaques of both human patients and ApoE-/- mice. Knockdown of KLF13 was found to ameliorate intimal hyperplasia following carotid artery injury. Furthermore, we discovered that KLF13 directly binds to the SM22α promoter, leading to the phenotypic dedifferentiation of VSMCs. Remarkably, we observed a significant inhibition of platelet-derived growth factor BB-induced VSMCs dedifferentiation, proliferation, and migration when knocked down KLF13 in VSMCs. This inhibitory effect of KLF13 knockdown on VCMC function was, at least in part, mediated by the inactivation of p-AKT signaling in VSMCs. Overall, our findings shed light on a potential therapeutic target for treating atherosclerotic lesions and restenosis after vascular injury.

Asunto(s)

Desdiferenciación Celular , Proliferación Celular , Factores de Transcripción de Tipo Kruppel , Proteínas de Microfilamentos , Proteínas Musculares , Músculo Liso Vascular , Miocitos del Músculo Liso , Proteínas Represoras , Animales , Humanos , Masculino , Ratones , Aterosclerosis/genética , Aterosclerosis/patología , Aterosclerosis/metabolismo , Traumatismos de las Arterias Carótidas/patología , Traumatismos de las Arterias Carótidas/genética , Traumatismos de las Arterias Carótidas/metabolismo , Movimiento Celular/genética , Proliferación Celular/genética , Células Cultivadas , Factores de Transcripción de Tipo Kruppel/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Ratones Endogámicos C57BL , Proteínas Musculares/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Neointima/metabolismo , Neointima/patología , Neointima/genética , Fenotipo , Placa Aterosclerótica/patología , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/genética , Regiones Promotoras Genéticas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Transducción de Señal , Proteínas de Ciclo Celular , Proteínas de Microfilamentos/genética

RESUMEN

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.

Asunto(s)

Proteína HMGB1 , Hiperplasia , Ratones Noqueados , Músculo Liso Vascular , Miocitos del Músculo Liso , Proteínas del Factor Nuclear 90 , Estabilidad del ARN , Factor de Transcripción STAT3 , Túnica Íntima , Animales , Humanos , Masculino , Ratones , Movimiento Celular , Proliferación Celular , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Proteína HMGB1/metabolismo , Proteína HMGB1/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/metabolismo , Neointima/metabolismo , Neointima/patología , Proteínas del Factor Nuclear 90/metabolismo , Proteínas del Factor Nuclear 90/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal , Factor de Transcripción STAT3/metabolismo , Túnica Íntima/metabolismo , Túnica Íntima/patología

RESUMEN

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) plays an important role in vascular smooth muscle cell (VSMC) phenotypic switching, which is an early pathogenic event in various vascular remodeling diseases (VRDs). However, the underlying mechanism is not fully understood. METHODS: An IP‒LC‒MS/MS assay was conducted to identify new binding partners of G6PD involved in the regulation of VSMC phenotypic switching under platelet-derived growth factor-BB (PDGF-BB) stimulation. Co-IP, GST pull-down, and immunofluorescence colocalization were employed to clarify the interaction between G6PD and voltage-dependent anion-selective channel protein 1 (VDAC1). The molecular mechanisms involved were elucidated by examining the interaction between VDAC1 and apoptosis-related biomarkers, as well as the oligomerization state of VDAC1. RESULTS: The G6PD level was significantly elevated and positively correlated with the synthetic characteristics of VSMCs induced by PDGF-BB. We identified VDAC1 as a novel G6PD-interacting molecule essential for apoptosis. Specifically, the G6PD-NTD region was found to predominantly contribute to this interaction. G6PD promotes VSMC survival and accelerates vascular neointimal hyperplasia by inhibiting VSMC apoptosis. Mechanistically, G6PD interacts with VDAC1 upon stimulation with PDGF-BB. By competing with Bax for VDAC1 binding, G6PD reduces VDAC1 oligomerization and counteracts VDAC1-Bax-mediated apoptosis, thereby accelerating neointimal hyperplasia. CONCLUSION: Our study showed that the G6PD-VDAC1-Bax axis is a vital switch in VSMC apoptosis and is essential for VSMC phenotypic switching and neointimal hyperplasia, providing mechanistic insight into early VRDs.

Asunto(s)

Glucosafosfato Deshidrogenasa , Músculo Liso Vascular , Canal Aniónico 1 Dependiente del Voltaje , Humanos , Hiperplasia/metabolismo , Hiperplasia/patología , Becaplermina/genética , Becaplermina/metabolismo , Proliferación Celular , Proteína X Asociada a bcl-2/metabolismo , Glucosafosfato Deshidrogenasa/metabolismo , Músculo Liso Vascular/metabolismo , Cromatografía Liquida , Espectrometría de Masas en Tándem , Neointima/genética , Neointima/metabolismo , Neointima/patología , Apoptosis , Miocitos del Músculo Liso/metabolismo , Movimiento Celular/genética , Células Cultivadas , Fenotipo

RESUMEN

The abnormal proliferation, migration, and inflammation of vascular smooth muscle cells (VSMCs) play crucial roles in the development of neointimal hyperplasia and restenosis. Exposure to inflammatory cytokines such as platelet-derived growth factor (PDGF)-BB and tumour necrosis factor-alpha (TNF-α) induces the transformation of contractile VSMCs into abnormal synthetic VSMCs. Isoxanthohumol (IXN) has significant anti-inflammatory, antiproliferative, and antimigratory effects. This study aimed to explore the therapeutic impact and regulatory mechanism of IXN in treating neointimal hyperplasia. The present findings indicate that IXN effectively hinders the abnormal proliferation, migration, and inflammation of VSMCs triggered by PDGF or TNF-α. This inhibition is primarily achieved through the modulation of the apelin/AKT or AKT pathway, respectively. In an in vivo model, IXN effectively reduced neointimal hyperplasia in denuded femoral arteries. These results suggest that IXN holds promise as a potential and innovative therapeutic candidate for the treatment of restenosis.

Asunto(s)

Proteínas Proto-Oncogénicas c-akt , Factor de Necrosis Tumoral alfa , Xantonas , Humanos , Hiperplasia/tratamiento farmacológico , Proliferación Celular , Proteínas Proto-Oncogénicas c-akt/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Apelina , Movimiento Celular , Becaplermina/farmacología , Neointima/tratamiento farmacológico , Neointima/metabolismo , Inflamación

RESUMEN

BACKGROUND AND AIMS: Vascular injury-induced endothelium-denudation and profound vascular smooth muscle cells (VSMCs) proliferation and dis-regulated apoptosis lead to post-angioplasty restenosis. Coptisine (CTS), an isoquinoline alkaloid, has multiple beneficial effects on the cardiovascular system. Recent studies identified it selectively inhibits VSMCs proliferation. However, its effects on neointimal hyperplasia, re-endothelialization, and the underlying mechanisms are still unclear. METHODS: Cell viability was assayed by 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and cell counting kit-8 (CCK-8). Cell proliferation and apoptosis were measured by flow cytometry and immunofluorescence of Ki67 and TUNEL. Quantitative phosphoproteomics (QPP) was employed to screen CTS-responsive phosphor-sites in the key regulators of cell proliferation and apoptosis. Neointimal hyperplasia was induced by balloon injury of rat left carotid artery (LCA). Adenoviral gene transfer was conducted in both cultured cells and LCA. Re-endothelialization was evaluated by Evan's blue staining of LCA. RESULTS: 1) CTS had strong anti-proliferative and pro-apoptotic effects in cultured rat VSMCs, with the EC50 4∼10-folds lower than that in endothelial cells (ECs). 2) Rats administered with CTS, either locally to LCA's periadventitial space or orally, demonstrated a potently inhibited balloon injury-induced neointimal hyperplasia, but had no delaying effect on re-endothelialization. 3) The QPP results revealed that the phosphorylation levels of Pak1S144/S203, Pak2S20/S197, Erk1T202/Y204, Erk2T185/Y187, and BadS136 were significantly decreased in VSMCs by CTS. 4) Adenoviral expression of phosphomimetic mutants Pak1D144/D203/Pak2D20/D197 enhanced Pak1/2 activities, stimulated the downstream pErk1T202/Y204/pErk2T185/Y187/pErk3S189/pBadS136, attenuated CTS-mediated inhibition of VSMCs proliferation and promotion of apoptosis in vitro, and potentiated neointimal hyperplasia in vivo. 5) Adenoviral expression of phosphoresistant mutants Pak1A144/A203/Pak2A20/A197 inactivated Pak1/2 and totally simulated the inhibitory effects of CTS on platelet-derived growth factor (PDGF)-stimulated VSMCs proliferation and PDGF-inhibited apoptosis in vitro and neointimal hyperplasia in vivo. 6) LCA injury significantly enhanced the endogenous phosphorylation levels of all but pBadS136. CTS markedly attenuated all the enhanced levels. CONCLUSIONS: These results indicate that CTS is a promising medicine for prevention of post-angioplasty restenosis without adverse impact on re-endothelialization. CTS-directed suppression of pPak1S144/S203/pPak2S20/S197 and the subsequent effects on downstream pErk1T202/Y204/pErk2T185/Y187/pErk3S189 and pBadS136 underline its mechanisms of inhibition of VSMCs proliferation and stimulation of apoptosis. Therefore, the phosphor-sites of Pak1S144/S203/Pak2S20/S197 constitute a potential drug-screening target for fighting neointimal hyperplasia restenosis.

Asunto(s)

Berberina/análogos & derivados , Traumatismos de las Arterias Carótidas , Músculo Liso Vascular , Ratas , Animales , Hiperplasia/patología , Músculo Liso Vascular/patología , Células Endoteliales/metabolismo , Proliferación Celular , Neointima/metabolismo , Traumatismos de las Arterias Carótidas/patología , Células Cultivadas , Miocitos del Músculo Liso/patología , Movimiento Celular

RESUMEN

BACKGROUND AND AIMS: Myotubularin-related protein 7 (MTMR7) suppresses proliferation in various cell types and is associated with cardiovascular and cerebrovascular diseases. However, whether MTMR7 regulates vascular smooth muscle cell (VSMC) and vascular intimal hyperplasia remains unclear. We explored the role of MTMR7 in phenotypic switching of VSMC and vascular intimal hyperplasia after injury. METHODS AND RESULTS: MTMR7 expression was significantly downregulated in injured arteries. Compared to wild type (WT) mice, Mtmr7-transgenic (Mtmr7-Tg) mice showed reduced intima/media ratio, decreased percentage of Ki-67-positive cells within neointima, and increased Calponin expression in injured artery. In vitro, upregulating MTMR7 by Len-Mtmr7 transfection inhibited platelet derived growth factor (PDGF)-BB-induced proliferation, migration of VSMC and reversed PDGF-BB-induced decrease in expression of Calponin and SM-MHC. Microarray, single cell sequence, and other bioinformatics analysis revealed that MTMR7 is highly related to glucose metabolism and mammalian target of rapamycin complex 1 (mTORC1). Further experiments confirmed that MTMR7 markedly repressed glycolysis and mTORC1 activity in PDGF-BB-challenged VSMC in vitro. Restoring mTORC1 activity abolished MTMR7-mediated suppression of glycolysis, phenotypic shift in VSMC in vitro and protection against vascular intimal hyperplasia in vivo. Furthermore, upregulating MTMR7 in vitro led to dephosphorylation and dissociation of p62 from mTORC1 in VSMC. External expression of p62 in vitro also abrogated the inhibitory effects of MTMR7 on glycolysis and phenotypic switching in PDGF-BB-stimulated VSMC. CONCLUSIONS: Our study demonstrates that MTMR7 inhibits injury-induced vascular intimal hyperplasia and phenotypic switching of VSMC. Mechanistically, the beneficial effects of MTMR7 are conducted via suppressing p62/mTORC1-mediated glycolysis.

Asunto(s)

Músculo Liso Vascular , Neointima , Ratones , Animales , Becaplermina/farmacología , Becaplermina/metabolismo , Proliferación Celular , Músculo Liso Vascular/patología , Hiperplasia/patología , Neointima/metabolismo , Ratones Transgénicos , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/farmacología , Glucosa/metabolismo , Miocitos del Músculo Liso/patología , Movimiento Celular , Células Cultivadas , Mamíferos

RESUMEN

BACKGROUND: Vascular smooth muscle cell (VSMC) proliferation is the leading cause of vascular stenosis or restenosis. Therefore, investigating the molecular mechanisms and pivotal regulators of the proliferative VSMC phenotype is imperative for precisely preventing neointimal hyperplasia in vascular disease. METHODS: Wire-induced vascular injury and aortic culture models were used to detect the expression of staphylococcal nuclease domain-containing protein 1 (SND1). SMC-specific Snd1 knockout mice were used to assess the potential roles of SND1 after vascular injury. Primary VSMCs were cultured to evaluate SND1 function on VSMC phenotype switching, as well as to investigate the mechanism by which SND1 regulates the VSMC proliferative phenotype. RESULTS: Phenotype-switched proliferative VSMCs exhibited higher SND1 protein expression compared to the differentiated VSMCs. This result was replicated in primary VSMCs treated with platelet-derived growth factor (PDGF). In the injury model, specific knockout of Snd1 in mouse VSMCs reduced neointimal hyperplasia. We then revealed that ETS transcription factor ELK1 (ELK1) exhibited upregulation and activation in proliferative VSMCs, and acted as a novel transcription factor to induce the gene transcriptional activation of Snd1. Subsequently, the upregulated SND1 is associated with serum response factor (SRF) by competing with myocardin (MYOCD). As a co-activator of SRF, SND1 recruited the lysine acetyltransferase 2B (KAT2B) to the promoter regions leading to the histone acetylation, consequently promoted SRF to recognize the specific CArG motif, and enhanced the proliferation- and migration-related gene transcriptional activation. CONCLUSIONS: The present study identifies ELK1/SND1/SRF as a novel pathway in promoting the proliferative VSMC phenotype and neointimal hyperplasia in vascular injury, predisposing the vessels to pathological remodeling. This provides a potential therapeutic target for vascular stenosis.

Asunto(s)

Músculo Liso Vascular , Lesiones del Sistema Vascular , Ratones , Animales , Hiperplasia/metabolismo , Lesiones del Sistema Vascular/genética , Lesiones del Sistema Vascular/metabolismo , Lesiones del Sistema Vascular/patología , Proliferación Celular , Factor de Respuesta Sérica/genética , Factor de Respuesta Sérica/metabolismo , Constricción Patológica/metabolismo , Constricción Patológica/patología , Factores de Transcripción/metabolismo , Fenotipo , Neointima/genética , Neointima/metabolismo , Neointima/patología , Miocitos del Músculo Liso/metabolismo , Células Cultivadas , Movimiento Celular

RESUMEN

Neointimal hyperplasia is a pathological vascular remodeling caused by abnormal proliferation and migration of subintimal vascular smooth muscle cells (VSMCs) following intimal injury. There is increasing evidence that tRNA-derived small RNA (tsRNA) plays an important role in vascular remodeling. The purpose of this study is to search for tsRNAs signature of neointima formation and to explore their potential functions. The balloon injury model of rat common carotid artery was replicated to induce intimal hyperplasia, and the differentially expressed tsRNAs (DE-tsRNAs) in arteries with intimal hyperplasia were screened by small RNA sequencing and tsRNA library. A total of 24 DE-tsRNAs were found in the vessels with intimal hyperplasia by small RNA sequencing. In vitro, tRF-Glu-CTC inhibited the expression of fibromodulin (FMOD) in VSMCs, which is a negative modulator of TGF-ß1 activity. tRF-Glu-CTC also increased VSMC proliferation and migration. In vivo experiments showed that inhibition of tRF-Glu-CTC expression after balloon injury of rat carotid artery can reduce the neointimal area. In conclusion, tRF-Glu-CTC expression is increased after vascular injury and inhibits FMOD expression in VSMCs, which influences neointima formation. On the other hand, reducing the expression of tRF-Glu-CTC after vascular injury may be a potential approach to prevent vascular stenosis.

Asunto(s)

Traumatismos de las Arterias Carótidas , Lesiones del Sistema Vascular , Animales , Ratas , Traumatismos de las Arterias Carótidas/genética , Traumatismos de las Arterias Carótidas/metabolismo , Movimiento Celular , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Fibromodulina/metabolismo , Hiperplasia/complicaciones , Hiperplasia/metabolismo , Hiperplasia/patología , Miocitos del Músculo Liso/metabolismo , Neointima/metabolismo , Neointima/patología , Neointima/prevención & control , Ratas Sprague-Dawley , ARN/metabolismo , ARN de Transferencia/metabolismo , Remodelación Vascular , Lesiones del Sistema Vascular/metabolismo

RESUMEN

BACKGROUND AND AIMS: New treatments are needed to prevent neointimal hyperplasia that contributes to post-angioplasty and stent restenosis in patients with coronary artery disease (CAD) and peripheral arterial disease (PAD). We investigated whether modulating mitochondrial function using mitochondrial division inhibitor-1 (Mdivi-1) could reduce post-vascular injury neointimal hyperplasia by metabolic reprogramming of macrophages from a pro-inflammatory to anti-inflammatory phenotype. METHODS AND RESULTS: In vivo Mdivi-1 treatment of Apoe-/- mice fed a high-fat diet and subjected to carotid-wire injury decreased neointimal hyperplasia by 68%, reduced numbers of plaque vascular smooth muscle cells and pro-inflammatory M1-like macrophages, and decreased plaque inflammation, endothelial activation, and apoptosis, when compared to control. Mdivi-1 treatment of human THP-1 macrophages shifted polarization from a pro-inflammatory M1-like to an anti-inflammatory M2-like phenotype, reduced monocyte chemotaxis and migration to CCL2 and macrophage colony stimulating factor (M-CSF) and decreased secretion of pro-inflammatory mediators. Finally, treatment of pro-inflammatory M1-type-macrophages with Mdivi-1 metabolically reprogrammed them to an anti-inflammatory M2-like phenotype by inhibiting oxidative phosphorylation and attenuating the increase in succinate levels and correcting the decreased levels of arginine and citrulline. CONCLUSIONS: We report that treatment with Mdivi-1 inhibits post-vascular injury neointimal hyperplasia by metabolic reprogramming macrophages towards an anti-inflammatory phenotype thereby highlighting the therapeutic potential of Mdivi-1 for preventing neointimal hyperplasia and restenosis following angioplasty and stenting in CAD and PAD patients.

Asunto(s)

Quinazolinonas , Lesiones del Sistema Vascular , Humanos , Ratones , Animales , Hiperplasia/patología , Lesiones del Sistema Vascular/genética , Reprogramación Metabólica , Movimiento Celular , Músculo Liso Vascular/patología , Neointima/metabolismo , Antiinflamatorios/farmacología , Modelos Animales de Enfermedad , Proliferación Celular