Ultrasound-guided periadventitial administration of rapamycin-fibrin glue attenuates neointimal hyperplasia in the rat carotid artery injury model.

INTRODUCTION: Arterial restenosis caused by intimal hyperplasia (IH) is a serious complication after vascular interventions. In the rat carotid balloon injury model, we injected phosphate buffer saline (PBS), rapamycin-phosphate buffer saline suspension (RPM-PBS), blank fibrin glue (FG) and rapamycin-fibrin glue (RPM-FG) around the injured carotid artery under ultrasound guidance and observed the inhibitory effect on IH. METHODS: The properties of RPM-FG in vitro were verified by scanning electron microscopy (SEM) and determination of the drug release rate. FG metabolism in vivo was observed by fluorescence imaging. The rat carotid balloon injury models were randomly classified into 4 groups: PBS group (control group), RPM-PBS group, FG group, and RPM-FG group. Periadventitial administration was performed by ultrasound-guided percutaneous puncture on the first day after angioplasty. Carotid artery specimens were analyzed by immunostaining, Evans blue staining and hematoxylin-eosin staining. RESULTS: The RPM particles showed clustered distributions in the FG block. The glue was maintained for a longer time in vivo (> 14 days) than in vitro (approximately 7 days). Two-component liquid FG administered by ultrasound-guided injection completely encapsulated the injured artery before coagulation. The RPM-FG inhibited IH after carotid angioplasty vs. control and other groups. The proliferation of vascular smooth muscle cells (VSMCs) was significantly inhibited during neointima formation, whereas endothelial cell (EC) repair was not affected. CONCLUSION: Periadventitial delivery of RPM-FG contributed to inhibiting IH in the rat carotid artery injury model without compromising re-endothelialization. Additionally, FG provided a promising platform for the future development of a safe, effective, and minimally invasive perivascular drug delivery method to treat vascular disease.

Omentin-1 enhances the inhibitory effect of endothelial progenitor cells on neointimal hyperplasia by inhibiting the p38 MAPK/CREB pathway.

AIMS: Endothelial progenitor cells (EPCs) play an important role in vascular repair. However, they are dysfunctional in the inflammatory microenvironment during restenosis. In this study, we investigated whether omentin-1, an anti-inflammatory factor, could reduce neointima formation after carotid artery injury (CAI) in rats by improving EPC functions that were damaged by inflammation and the underlying mechanisms. MAIN METHODS: EPCs were transfected with adenoviral vectors expressing human omentin-1 or green fluorescent protein (GFP). Then, the rats received 2 × 106 EPCs expressing omentin-1 or GFP by tail vein injection directly after CAI and again 24 h later. Hematoxylin-eosin staining and immunohistochemistry were used for analyzing neointimal hyperplasia. Besides, EPCs were treated with omentin-1 and TNF-α to examine the underlying mechanism. KEY FINDINGS: Our results showed that omentin-1 could significantly improve EPC functions, including proliferation, apoptosis and tube formation. Meanwhile, EPCs overexpressed with omentin-1 could significantly reduce neointimal hyperplasia and tumor necrosis factor-α (TNF-α) expression after CAI in rats. TNF-α could notably induce EPC dysfunction, which could be markedly reversed by omentin-1 through the inhibition of the p38 MAPK/CREB pathway. Furthermore, a p38 MAPK agonist (anisomycin) significantly abrogated the protective effects of omentin-1 on EPCs damaged by TNF-α. SIGNIFICANCE: Our results indicated that genetically modifying EPC with omentin-1 could be an alternative strategy for the treatment of restenosis.

Ketogenic diet inhibits neointimal hyperplasia by suppressing oxidative stress and inflammation.

OBJECTIVE: Neointimal hyperplasia is the primary mechanism underlying atherosclerosis and restenosis after percutaneous coronary intervention. Ketogenic diet (KD) exerts beneficial effects in various diseases, but whether it could serve as non-drug therapy for neointimal hyperplasia remains unknown. This study aimed to investigate the effect of KD on neointimal hyperplasia and the potential mechanisms. METHODS AND RESULTS: Carotid artery balloon-injury model was employed in adult Sprague-Dawley rats to induce neointimal hyperplasia. Then, animals were subjected to either standard rodent chow or KD. For in-vitro experiment, impacts of ß-hydroxybutyrate (ß-HB), the main mediator of KD effects, on platelet-derived growth factor BB (PDGF-BB) induced vascular smooth muscle cell (VSMC) migration and proliferation were determined. Balloon injury induced event intimal hyperplasia and upregulation of protein expression of proliferating cell nuclear antigen (PCNA) and α-smooth muscle actin (α-SMA), and these changes were significantly ameliorated by KD. In addition, ß-HB could markedly inhibit PDGF-BB induced VMSC migration and proliferation, as well as inhibiting expressions of PCNA and α-SMC. Furthermore, KD inhibited balloon-injury induced oxidative stress in carotid artery, indicated by reduced ROS level, malondialdehyde (MDA) and myeloperoxidase (MPO) activities, and increased superoxide dismutase (SOD) activity. We also found balloon-injury induced inflammation in carotid artery was suppressed by KD, indicated by decreased expressions of proinflammatory cytokines IL-1ß and TNF-α, and increased expression of anti-inflammatory cytokine IL-10. CONCLUSION: KD attenuates neointimal hyperplasia through suppressing oxidative stress and inflammation to inhibit VSMC proliferation and migration. KD may represent a promising non-drug therapy for neointimal hyperplasia associated diseases.

Paeonol Promotes Reendothelialization After Vascular Injury Through Activation of c-Myc/VEGFR2 Signaling Pathway.

Purpose: Dysfunction of endothelium is associated with multiple pathological vascular diseases. However, how to regulate reendothelialization after vascular injury is not well defined. This study aims to determine whether and how Paeonol controls reendothelialization following artery injury. Methods: The endothelium of murine carotid artery was denuded by catheter guide wires injury. H&E staining and IF staining were performed to determine whether Paeonol is critical for reendothelialization. BRDU Incorporation Assay, Boyden Chamber Migration Assay, Tube Formation Assay, and Spheroid Sprouting Assay were used to investigate whether Paeonol is involved in regulating proliferation and migration of endothelial cells. The underlying mechanism of how Paeonol regulates reendothelialization was determined by Molecular docking simulation and CO-IP Assay. Results: Paeonol treatment significantly inhibits neointima formation in carotid artery ligation model by promoting proliferation and migration of endothelial cells. Mechanistically, Paeonol enhances c-Myc expression, consequently interacts with VEGFR2 results in activating VEGF signaling pathway, and eventually promotes reendothelialization after vascular injury. Conclusion: Our data demonstrated that Paeonol plays a critical role in regulating vascular reendothelialization, which may be therapeutically used for treatment of pathological vascular diseases.

Galangin inhibits neointima formation induced by vascular injury via regulating the PI3K/AKT/mTOR pathway.

Aims: The proliferation and migration of vascular smooth muscle cells (VSMCs) play vital roles in the pathological process of neointima formation after vascular injury. Galangin, an extract of the ginger plant galangal, is involved in numerous biological activities, including inhibiting the proliferation and migration of tumor cells, but its effect on VSMCs is unknown. This study focused on the role and mechanism of galangin in the neointima formation induced by vascular injury. Methods and results: In this study, we found that galangin restrained the PDGF-BB-induced proliferation, migration and phenotypic switching of VSMCs in a concentration-dependent manner. In vivo, we established a model of carotid artery balloon injury in rats, followed by intragastric administration of galangin (40 mg kg-1 day-1 or 80 mg kg-1 day-1) for 14 or 28 consecutive days. Then, the degree of neointima hyperplasia was evaluated by H&E staining, and the level of relevant protein expression was assessed by immunofluorescence and western blotting. In vitro, we isolated and grew primary rat aortic smooth muscle cells, which were treated with PDGF-BB and different doses of galangin, and then CCK-8 assay, wound healing assay, transwell assay, western blotting and immunofluorescence assays were performed. We found that galangin significantly inhibited PDGF-BB-induced proliferation, migration, and phenotypic switching of VSMCs and promoted autophagy in VSMCs in vitro, and galangin significantly inhibited neointimal hyperplasia after the common carotid artery balloon injury in rats. In terms of mechanisms, galangin inhibited the PI3K/AKT/mTOR pathway, thereby suppressing VSMC's switch from a contractile to a synthetic phenotype, inhibiting VSMC proliferation, migration and phenotypic switching and upregulating the Beclin1 protein expression levels and the ratio of LC3BII/I, promoting VSMC autophagy, and thereby inhibiting neointimal hyperplasia after vascular injury. Conclusion: Our study suggests that galangin inhibits neointimal hyperplasia after vascular injury by inhibiting smooth muscle cell proliferation, migration and phenotypic switching and by promoting autophagy, and that galangin may be a promising drug for the prevention and treatment of vascular restenosis after PCI.

Icariside II Attenuates Vascular Remodeling Via Wnt7b/CCND1 Axis.

ABSTRACT: Angioplasty often fails due to the abnormal proliferation of vascular smooth muscle cells (VSMCs). Success rates of angioplasty may increase following the administration of an agent that effectively ameliorates aberrant vascular remodeling. Icariside II (ICS-II) is a natural flavonol glycoside extract from the Chinese herbal medicine Epimedii that possesses several medicinal qualities that are beneficial in humans. Nevertheless, the role of ICS-II in addressing aberrant vascular remodeling have yet to be clarified. The current investigation studies the molecular effects of ICS-Ⅱ on balloon-inflicted neointimal hyperplasia in rats in vivo and on platelet-derived growth factor-induced vascular proliferation in primary rat aortic smooth muscle cells (VSMCs) in vitro. ICS-II was found to be as effective as rapamycin, the positive control used in this study. ICS-II inhibited neointimal formation in injured rat carotid arteries and notably reduced the expression of Wnt7b. ICS-Ⅱ significantly counteracted platelet-derived growth factor-induced VSMCs proliferation. Cell cycle analysis showed that ICS-II triggered cell cycle arrest during the G1/S transition. Western blot analysis further indicated that this cell cycle arrest was likely through Wnt7b suppression that led to CCND1 inhibition. In conclusion, our findings demonstrate that ICS-II possesses significant antiproliferative qualities that counteracts aberrant vascular neointimal hyperplasia. This phenomenon most likely occurs due to the suppression of the Wnt7b/CCND1 axis.

Paeonol Suppresses Vasculogenesis Through Regulating Vascular Smooth Muscle Phenotypic Switching.

OBJECTIVE: Smooth muscle cell (SMC) phenotypic switching is associated with development of a variety of occlusive vascular diseases. Paeonol has been reported to be involved in suppressing SMC proliferation. However, it is still unknown whether paeonol can regulate SMC phenotypic switching, and which eventually result in suppressing vasculogenesis. METHODS: Murine left common carotid artery was injured by completely ligation, and paeonol was administrated by intraperitoneal injection. Hematoxylin and eosin (H&E) staining was performed to visualize vascular neointima formation. Rat aortic SMCs were used to determine whether paeonol suppresses cell proliferation and migration. And murine hind limb ischemia model was performed to confirm the function role of paeonol in suppressing vasculogenesis. RESULTS: Complete ligation of murine common carotid artery successfully induced neointima formation. Paeonol treatment dramatically reduced the size of injury-induced neointima. Using rat aortic primary SMC, we identified that paeonol strongly suppressed cell proliferation, migration, and decreased extracellular matrix deposition. And paeonol treatment dramatically suppressed vasculogenesis after hind limb ischemia injury. CONCLUSION: Paeonol could regulate SMC phenotypic switching through inhibiting proliferation and migration of SMC, which results in inhibiting ischemia-induced vasculogenesis.

A neutralizing IL-11 antibody reduces vessel hyperplasia in a mouse carotid artery wire injury model.

Vascular restenosis remains a major problem in patients with coronary artery disease (CAD) and peripheral artery disease (PAD). Neointimal hyperplasia, defined by post-procedure proliferation and migration of vascular smooth muscle cells (VSMCs) is a key underlying pathology. Here we investigated the role of Interleukin 11 (IL-11) in a mouse model of injury-related plaque development. Apoe-/- mice were fed a hyperlipidaemic diet and subjected to carotid wire injury of the right carotid. Mice were injected with an anti-IL11 antibody (X203), IgG control antibody or buffer. We performed ultrasound analysis to assess vessel wall thickness and blood velocity. Using histology and immunofluorescence approaches, we determined the effects of IL-11 inhibition on VSMC and macrophages phenotypes and fibrosis. Treatment of mice with carotid wire injury using X203 significantly reduced post-endothelial injury vessel wall thickness, and injury-related plaque, when compared to control. Immunofluorescence staining of the injury-related plaque showed that X203 treatment did not reduce macrophage numbers, but reduced the number of VSMCs and lowered matrix metalloproteinase 2 (MMP2) levels and collagen content in comparison to control. X203 treatment was associated with a significant increase in smooth muscle protein 22α (SM22α) positive cells in injury-related plaque compared to control, suggesting preservation of the contractile VSMC phenotype. Interestingly, X203 also reduced the collagen content of uninjured carotid arteries as compared to IgG, showing an additional effect on hyperlipidemia-induced arterial remodeling in the absence of mechanical injury. Therapeutic inhibition of IL-11 reduced vessel wall thickness, attenuated neointimal hyperplasia, and has favorable effects on vascular remodeling following wire-induced endothelial injury. This suggests IL-11 inhibition as a potential novel therapeutic approach to reduce arterial stenosis following revascularization in CAD and PAD patients.

Physalin B inhibits PDGF-BB-induced VSMC proliferation, migration and phenotypic transformation by activating the Nrf2 pathway.

Vascular intimal hyperplasia is a hallmark event in vascular restenosis. The excessive proliferation, migration and phenotypic transformation of vascular smooth muscle cells (VSMCs) play important roles in the pathological mechanism of vascular intimal hyperplasia. Physalin B is an alcoholate isolated from Physalis (Solanaceae) that has a wide range of biological activities. However, the effect of physalin B on VSMCs is currently unclear. In this study, we demonstrated that physalin B significantly inhibited the proliferation, migration and phenotypic transformation of VSMCs induced by PDGF-BB. Physalin B also reduced inflammation and oxidative stress in VSMCs induced by PDGF-BB. Mechanistic studies showed that physalin B plays a role mainly by activating Nrf2. After Nrf2 activation, physalin B mitigates oxidative stress by enhancing the expression of the antioxidant gene HO-1; on the other hand, physalin B inhibits the NF-κB pathway to alleviate the inflammatory response. These two effects ultimately reduce the proliferation, migration and phenotypic transformation of VSMCs induced by PDGF-BB. In addition, in the mouse carotid artery ligation model, physalin B prevented intimal hyperplasia and inhibited the proliferation, migration and phenotypic transformation of cells in the hyperplastic intima. In conclusion, we provided significant evidence that physalin B abrogates PDGF-BB-induced VSMC proliferation, migration, phenotypic transformation and intimal hyperplasia by activating Nrf2-mediated signal transduction. Therefore, physalin B may be a potential therapeutic agent for preventing or treating restenosis.

Human urinary kininogenase reduces the endothelial injury by inhibiting Pyk2/MCU pathway.

The injury of endothelial cells is one of the initiating factors in restenosis after endovascular treatment. Human urinary kallidinogenase (HUK) is a tissue kallikrein which is used for ischemia-reperfusion injury treatment. Studies have shown that HUK may be a potential therapeutic agent to prevent stenosis after vascular injury, however, the precise mechanisms have not been fully established. This study is to investigate whether HUK can protect endothelial cells after balloon injury or H2O2-induced endothelial cell damage through the proline-rich tyrosine kinase 2 (Pyk2)/mitochondrial calcium uniporter (MCU) pathway. Intimal hyperplasia, a decrease of pinocytotic vesicles and cell apoptosis were found in the common carotid artery balloon injury and H2O2-induced endothelial cell damage, Pyk2/MCU was also up-regulated in such pathological process. HUK could prevent these injuries partially via the bradykinin B2 receptor by inhibiting Pyk2/MCU pathway, which prevented the mitochondrial damage, maintained calcium balance, and eventually inhibited cell apoptosis. Furthermore, MCU expression was not markedly increased if Pyk2 was suppressed by shRNA technique in the H2O2 treatment group, and cell viability was significantly better than H2O2-treated only. In short, our results indicate that the Pyk2/MCU pathway is involved in endothelial injury induced by balloon injury or H2O2-induced endothelial cell damage. HUK plays an protective role by inhibiting the Pyk2/MCU pathway in the endothelial injury.

Rapid Hemostatic Biomaterial from a Natural Bath Sponge Skeleton.

Uncontrolled bleeding is the main cause of mortality from trauma. Collagen has been developed as an important hemostatic material due to its platelet affinity function. A bath sponge skeleton is rich in collagen, also known as spongin. To understand the hemostatic effect of spongin, spongin materials, SX, SFM and SR were prepared from the bath sponge Spongia officinalis, and hemostatic experiments were performed. The SX, SFM and SR were significantly better than the positive control, type I collagen, in shortening the whole blood clotting time in vitro and hemostasis upon rat tail amputation. In a hemostatic experiment of rabbit common carotid artery injury, the hemostatic time and 3 h survival rate of the SFM group were 3.00 ± 1.53 min and 100%, respectively, which are significantly better than those of the commercial hemostat CELOX-A (10.33 ± 1.37 min and 67%, respectively). Additionally, the SFM showed good coagulation effects in platelet-deficient blood and defibrinated blood, while also showing good biocompatibility. Through a variety of tests, we speculated that the hemostatic activity of the SFM is mainly caused by its hyperabsorbency, high affinity to platelets and high effective concentration. Overall, the SFM and spongin derivates could be potential hemostatic agents for uncontrolled bleeding and hemorrhagic diseases caused by deficiency or dysfunction of coagulation factors.

Pinellia ternata attenuates carotid artery intimal hyperplasia and increases endothelial progenitor cell activity via the PI3K/Akt signalling pathway in wire-injured rats.

CONTEXT: Clinically, Pinellia ternata (Thunb.) Breit. (Araceae) (PT) has been widely used in the treatment of atherosclerosis and hyperlipidaemia, but the underlying mechanisms are still not clearly understood. OBJECTIVE: This research was conducted to confirm the mechanism by which PT affects carotid artery intimal hyperplasia. MATERIALS AND METHODS: An intestinal hyperplasia Sprague-Dawley rat model was established by carotid artery injury. The rats were randomly divided into five groups (n = 8): sham, model, PT (with daily intragastric administration of 10 g/mL/kg PT tubers water extract), PT+LY294002 (with intraperitoneal injection of 50 mg/kg LY294002 + 10 g/mL/kg PT) and endothelial progenitor cells (EPCs) (with injection of 5 × 105/cells), and treated for 4 or 8 weeks. RESULTS: HE staining showed that PT attenuated intimal hyperplasia. RT-PCR, Western blotting and immunohistochemistry showed that PT increased the expression of vascular endothelial growth factor (VEGF) and eNOS in the atherosclerotic carotid artery. PT increased the Dil-acLDL+/FITC-UEA-1+ population (from 0.41 ± 0.085% to 0.60 ± 0.092%) in the blood, decreased TCHO, TG, LDL-C, IL-6 and TNF-α levels, and increased HDL-C and IL-10 levels in the blood. However, these changes were reversed by the PI3K/Akt pathway inhibitor LY294002. DISCUSSION AND CONCLUSIONS: PT can be developed as an atherosclerosis and carotid intimal hyperplasia treatment drug. Therefore, further study will focus on the effects of PT on intimal hyperplasia in wire-injured atherosclerosis patients and explore in depth some other relevant molecular mechanisms.

Tamoxifen Accelerates Endothelial Healing by Targeting ERα in Smooth Muscle Cells.

RATIONALE: Tamoxifen prevents the recurrence of breast cancer and is also beneficial against bone demineralization and arterial diseases. It acts as an ER (estrogen receptor) α antagonist in ER-positive breast cancers, whereas it mimics the protective action of 17ß-estradiol in other tissues such as arteries. However, the mechanisms of these tissue-specific actions remain unclear. OBJECTIVE: Here, we tested whether tamoxifen is able to accelerate endothelial healing and analyzed the underlying mechanisms. METHODS AND RESULTS: Using 3 complementary mouse models of carotid artery injury, we demonstrated that both tamoxifen and estradiol accelerated endothelial healing, but only tamoxifen required the presence of the underlying medial smooth muscle cells. Chronic treatment with 17ß-estradiol and tamoxifen elicited differential gene expression profiles in the carotid artery. The use of transgenic mouse models targeting either whole ERα in a cell-specific manner or ERα subfunctions (membrane/extranuclear versus genomic/transcriptional) demonstrated that 17ß-estradiol-induced acceleration of endothelial healing is mediated by membrane ERα in endothelial cells, while the effect of tamoxifen is mediated by the nuclear actions of ERα in smooth muscle cells. CONCLUSIONS: Whereas tamoxifen acts as an antiestrogen and ERα antagonist in breast cancer but also on the membrane ERα of endothelial cells, it accelerates endothelial healing through activation of nuclear ERα in smooth muscle cells, inviting to revisit the mechanisms of action of selective modulation of ERα.

AMD3100 and SDF­1 regulate cellular functions of endothelial progenitor cells and accelerate endothelial regeneration in a rat carotid artery injury model.

The present study was conducted to assess the effects of AMD3100 and stromal cell-derived factor 1 (SDF-1) on cellular functions and endothelial regeneration of endothelial progenitor cells (EPCs). The cell proliferation and adhesion capacity of EPCs were evaluated in vitro following treatment with AMD3100 and SDF­1 using a Cell Counting Kit­8 assay. Furthermore, the expression levels of C­X­C motif chemokine receptor 4 (CXCR4) and C­X­C motif chemokine receptor 7 (CXCR7) were detected before and after treatment with AMD3100 and SDF­1 to elucidate their possible role in regulating the cellular function of EPCs. A rat carotid artery injury model was established to assess the influences of AMD3100 and SDF­1 on endothelial regeneration. AMD3100 reduced the proliferation and adhesion capacity of EPCs to fibronectin (FN), whereas it increased the adhesion capacity of EPCs to human umbilical vein endothelial cells (HUVECs). However, SDF­1 stimulated the proliferation and cell adhesion capacity of EPCs to HUVECs and FN. Additionally, the expression levels of CXCR7 but not CXCR4 were upregulated following AMD3100 treatment, whereas the expression levels of both CXCR4 and CXCR7 were upregulated after SDF­1 treatment. In vivo results demonstrated that AMD3100 increased the number of EPCs in the peripheral blood and facilitated endothelial repair at 7 days after treatment. However, local administration of SDF­1 alone did not enhance reendothelialization 7 and 14 days after treatment. Importantly, the combination of AMD3100 with SDF­1 exhibited superior therapeutic effects compared with AMD3100 treatment alone, accelerated reendothelialization 7 days after treatment, and attenuated neointimal hyperplasia at day 7 and 14 by recruiting more EPCs to the injury site. In conclusion, AMD3100 could positively regulate the adhesion capacity of EPCs to HUVECs via elevation of the expression levels of CXCR7 but not CXCR4, whereas SDF­1 could stimulate the proliferation and adhesion capacity of EPCs to FN and HUVECs by elevating the expression levels of CXCR4 and CXCR7. AMD3100 combined with SDF­1 outperformed AMD3100 alone, promoted early reendothelialization and inhibited neointimal hyperplasia, indicating that early reendothelialization attenuated neointimal hypoplasia following endothelial injury.

Pseudoaneurysm after carotid stenting: A case report and review of the literature.

Carotid artery stenting has been a widely used interventional treatment method for the last 3 decades in the treatment of carotid artery stenosis. In the current literature, unlike major cardiovascular complications, less emphasis has been placed on carotid pseudoaneurysm (PA). A carotid artery PA can be caused by trauma, spontaneous infection, vasculitis, or it may be iatrogenic. However, the incidence of PA secondary to carotid stenting is extremely rare. Although it may be completely asymptomatic in rare instances, it usually progresses symptomatically (neck swelling, nerve compression, respiratory distress, hoarseness, dysphagia, and ischemic cerebrovascular events). Doppler ultrasound, contrast-enhanced computed tomography, and conventional angiography are the main diagnostic tools. Primary closure, including graft interposition, has been described as a surgical therapeutic option. An endovascular approach with placement of a covered or bare metal stent is an alternative treatment method to surgery.

Resveratrol Inhibits Neointimal Growth after Arterial Injury in High-Fat-Fed Rodents: The Roles of SIRT1 and AMPK.

We have shown that both insulin and resveratrol (RSV) decrease neointimal hyperplasia in chow-fed rodents via mechanisms that are in part overlapping and involve the activation of endothelial nitric oxide synthase (eNOS). However, this vasculoprotective effect of insulin is abolished in high-fat-fed insulin-resistant rats. Since RSV, in addition to increasing insulin sensitivity, can activate eNOS via pathways that are independent of insulin signaling, such as the activation of sirtuin 1 (SIRT1) and AMP-activated kinase (AMPK), we speculated that unlike insulin, the vasculoprotective effect of RSV would be retained in high-fat-fed rats. We found that high-fat feeding decreased insulin sensitivity and increased neointimal area and that RSV improved insulin sensitivity (p < 0.05) and decreased neointimal area in high-fat-fed rats (p < 0.05). We investigated the role of SIRT1 in the effect of RSV using two genetic mouse models. We found that RSV decreased neointimal area in high-fat-fed wild-type mice (p < 0.05), an effect that was retained in mice with catalytically inactive SIRT1 (p < 0.05) and in heterozygous SIRT1-null mice. In contrast, the effect of RSV was abolished in AMKPα2-null mice. Thus, RSV decreased neointimal hyperplasia after arterial injury in both high-fat-fed rats and mice, an effect likely not mediated by SIRT1 but by AMPKα2.

Mutation of Arginine 264 on ERα (Estrogen Receptor Alpha) Selectively Abrogates the Rapid Signaling of Estradiol in the Endothelium Without Altering Fertility.

OBJECTIVE: ERα (estrogen receptor alpha) exerts nuclear genomic actions and also rapid membrane-initiated steroid signaling. The mutation of the cysteine 451 into alanine in vivo has recently revealed the key role of this ERα palmitoylation site on some vasculoprotective actions of 17ß-estradiol (E2) and fertility. Here, we studied the in vivo role of the arginine 260 of ERα which has also been described to be involved in its E2-induced rapid signaling with PI-3K (phosphoinositide 3-kinase) as well as G protein in cultured cell lines. Approach and Results: We generated a mouse model harboring a point mutation of the murine counterpart of this arginine into alanine (R264A-ERα). In contrast to the C451A-ERα, the R264A-ERα females are fertile with standard hormonal serum levels and normal control of hypothalamus-pituitary ovarian axis. Although R264A-ERα protein abundance was normal, the well-described membrane ERα-dependent actions of estradiol, such as the rapid dilation of mesenteric arteries and the acceleration of endothelial repair of carotid, were abrogated in R264A-ERα mice. In striking contrast, E2-regulated gene expression was highly preserved in the uterus and the aorta, revealing intact nuclear/genomic actions in response to E2. Consistently, 2 recognized nuclear ERα-dependent actions of E2, namely atheroma prevention and flow-mediated arterial remodeling were totally preserved. CONCLUSIONS: These data underline the exquisite role of arginine 264 of ERα for endothelial membrane-initiated steroid signaling effects of E2 but not for nuclear/genomic actions. This provides the first model of fertile mouse with no overt endocrine abnormalities with specific loss-of-function of rapid ERα signaling in vascular functions.

Apatinib attenuates phenotypic switching of arterial smooth muscle cells in vascular remodelling by targeting the PDGF Receptor-ß.

Apatinib (YN968D1) is a small-molecule tyrosine kinase inhibitor（TKI）which can inhibit the activity of vascular endothelial growth factor receptor-2 (VEGFR-2). It has been reported that apatinib has anti-tumour effect of inhibiting proliferation and inducing apoptosis of a variety of solid tumour cells, whereas its effect on vascular smooth muscle cells (VSMC) remains unclear. This study investigated the effect of apatinib on phenotypic switching of arterial smooth muscle cells in vascular remodelling. Compared to the vehicle groups, mice that were performed carotid artery ligation injury and treated with apatinib produced a reduction in abnormal neointimal area. For in vitro experiment, apatinib administration inhibited VSMC proliferation, migration and reversed VSMC dedifferentiation with the stimulation of platelet-derived growth factor type BB (PDGF-BB).In terms of mechanism, with the preincubation of apatinib, the activations of PDGF receptor-ß (PDGFR-ß) and phosphoinositide-specific phospholipase C-γ1 (PLC-γ1) induced by PDGF-BB were inhibited in VSMCs. With the preincubation of apatinib, the phosphorylation of PDGFR-ß, extracellular signal-related kinases (ERK1/2) and Jun amino-terminal kinases (JNK) induced by PDGF-BB were also inhibited in rat vascular smooth muscle cell line A7r5. Herein, we found that apatinib attenuates phenotypic switching of arterial smooth muscle cells induced by PDGF-BB in vitro and vascular remodelling in vivo. Therefore, apatinib is a potential candidate to treat vascular proliferative diseases.

Glycoprotein VI (GPVI)-functionalized nanoparticles targeting arterial injury sites under physiological flow.

Thrombus formation at athero-thrombotic sites is initiated by the exposure of collagen followed by platelet adhesion mediated by the platelet-specific collagen receptor glycoprotein VI (GPVI). Here, dimeric GPVI was used as a targeting motif to functionalize polymeric nanoparticle-based drug carriers and to show that with proper design, such GPVI-coated nanoparticles (GPNs) can efficiently and specifically target arterial injury sites while withstanding physiological flow. In a microfluidic model, under physiological shear levels (1-40 dyne/cm2), 200 nm and 2 µm GPNs exhibited a >60 and >10-fold increase in binding to collagen compared to control particles, respectively. In vitro experiments in an arterial stenosis injury model, subjected to physiological pulsatile flow, showed shear-enhanced adhesion of 200 nm GPNs at the stenosis region which was confirmed in vivo in a mice ligation carotid injury model using intravital microscopy. Altogether, our results illustrate how engineering tools can be harnessed to design nano-carriers that efficiently target cardiovascular disease sites.

Involvement of Angiotensin II Type 1 Receptor and Calcium Channel in Vascular Remodeling and Endothelial Dysfunction in Rats with Pressure Overload.

Vascular remodeling is an adaptive response to various stimuli, including mechanical forces, inflammatory cytokines and hormones. In the present study, we investigated the role of angiotensin II type 1 receptor (AT1R) and calcium channel in carotid artery remodeling in response to increased biomechanical forces by using the transverse aortic constriction (TAC) rat model. TAC was induced on ten-week-old male Sprague-Dawley rats and these models were treated with AT1R blocker olmesartan (1 mg/kg/day) or/and calcium channel blocker (CCB) amlodipine (0.5 mg/kg/day) for 14 days. After the treatment, the right common carotid artery proximal to the band (RCCA-B) was collected for further assay. Results showed that olmesartan, but not amlodipine, significantly prevented TAC-induced adventitial hyperplasia. Similarly, olmesartan, but not amlodipine, signifcantly prevented vascular infammation, as indicated by increased tumor necrosis factor α (TNF-α) and increased p65 phosphorylation, an indicator of nuclear factor κ-light-chain-enhancer of activated B cells (NFκB) activation in RCCA-B. In contrast, both olmesartan and amlodipine reversed the decreased expression of endothelial nitric oxidase synthase (eNOS) and improved endothelium-dependent vasodilation, whereas combination of olmesartan and amlodipine showed no further synergistic protective effects. These results suggest that AT1R was involved in vascular remodeling and inflammation in response to pressure overload, whereas AT1R and subsequent calcium channel were involved in endothelial dysfunction.