Common Carotid Artery Injury Research Articles

Expression profile of circular RNA in rat intimal hyperplasia and target gene prediction.

Intimal hyperplasia is an important cause of stenosis or occlusion after vascular injury. Circular RNAs (circRNAs) are known to be related to various cardiovascular diseases. However, the expression profile of circRNAs in the neointima has not been reported in detail. In this study, we established a rat common carotid artery (CCA) injury model. A microarray detection showed significant differences in circRNA expression between the normal and injured CCA. Real-time quantitative polymerase chain reaction verified the differences. We used bioinformatics to predict the microRNAs that possibly interact with the differentially expressed (DE) circRNAs and linked the potential functions of circRNAs to the target genes of the microRNAs. We believe that the DE circRNA in neointima may affect the differentiation, proliferation, and migration of vascular cells through a variety of target genes. The intervention or utilization of certain circRNAs should be a new method for preventing and treating intimal hyperplasia.

A case of common carotid artery injury with hemorrhagic shock

A case of common carotid artery injury with hemorrhagic shock

Abstract 18340: Alternatively Spliced Tissue Factor Promotes Atherosclerosis by Increasing Foam Cell Formation via LOX-1 and SR-A1 up-regulation

Introduction: Alternatively Spliced Tissue Factor (asTF) is an isoform of tissue factor that is expressed in human atherosclerotic plaques and promotes plaque progression in experimental atherosclerosis (Giannarelli C, Circulation 2014). Hypothesis: asTF is the isoform of tissue factor that most strongly promotes atherosclerosis by increasing foam cell formation. Methods: ApoE-/- mice (8 weeks old) were fed a Western-type diet starting 2 weeks before surgery. Immediately after transluminal wire injury of the left common carotid artery (LCCA), LCCA was incubated with lentivirus encoding asTF-GFP (asTF+;n=10), fl-TF-GFP (fl-TF+, n=10) or GFP (controls; n=5). Four weeks after, LCCA was removed and processed for the quantification of plaque size (H&E) and lipid accumulation (Oil-Red O). The effect of asTF on foam cell formation was tested in vitro by treating THP-1 derived macrophages with oxLDL (75μg/ml), with asTF (10nM) or vehicle. Total cholesterol (TC) and cholesterol esters (CE) were measured in lipid cell extracts. The mRNA levels of the oxLDL scavenger receptors LOX-1, SR-A1 and CD36 in macrophages and foam cells were assessed using qRT-PCR. Results: Plaque size and lipid accumulation were significantly greater in asTF+ vs. fl-TF+ and control mice (Fig.1, A-D). In vitro results showed that asTF promotes TC and CE accumulation in foam cells (Fig.1, E,F). Gene expression studies showed that asTF significantly increased the mRNA expression of scavenger receptors LOX-1, SR-A1 in both macrophages and foam cells (Fig.1, G-I). An increase in mRNA levels of CD36 (1.4-fold) was only detected in asTF-treated foam cells. Conclusions: In vivo results suggest that asTF promote plaque progression and lipid accumulation. In vitro studies imply that asTF promotes foam cell formation by increasing the expression of oxLDL scavenger receptors implicated in lipoprotein uptake by macrophages. These studies suggest a functional role for asTF in atherosclerotic plaque progression.

Preexisting smooth muscle cells contribute to neointimal cell repopulation at an incidence varying widely among individual lesions

With the diverse origin of neointimal cells, previous studies have documented differences of neointimal cell lineage composition across models, but the animal-to-animal difference has not attracted much attention, although the cellular heterogeneity may impact neointimal growth and its response to therapeutic interventions. R26R(+);Myh11-CreER(+), and R26R(+);Scl-CreER(+) mice were used to attach LacZ tags to the preexisting smooth muscle cells (SMCs) and endothelial cells (ECs), respectively. Neointimal lesions were created via complete ligation of the common carotid artery (CCA) and transluminal injury to the femoral artery (FA). LacZ-tagged SMCs were physically relocated from media to neointima and changed to a dedifferentiated phenotype in both CCA and FA lesions. The content of SMCs in the neointimal tissue, however, varied widely among specimens, ranging from 5 to 70% and 0 to 85%, with an average at low levels of 27% and 29% in CCA (n=15) and FA (n=15) lesions, respectively. Bone marrow cells, although able to home to the injured arteries, did not differentiate fully into SMCs after either type of injury. Preexisting ECs were located in the subendothelial region and produced mesenchymal marker α-actin, indicating endothelial-mesenchymal transition (EndoMT); however, EC-derived cells represented only 7% and 3% of the total neointimal cell pool of CCA (n=7) and FA (n=7) lesions, respectively. ECs located on the luminal surface exhibited little evidence of EndoMT. Neointimal hyperplasia proceeds with a wide range of variation in its cellular composition between individual lesions. Relative to ECs, SMCs are major contributors to the lesion-to-lesion heterogeneity in neointimal cell lineage composition.

Abstract 476: Glucagon Like Peptide -1 Receptor Activation Does Not Affect Re-endothelialization But Reduces Intimal Hyperplasia via Direct Effects on Smooth Muscle Cells in a Non-diabetic Model of Arterial Injury

Background/Aims: Diabetic patients have an increased risk of restenosis and late stent thrombosis after angioplasty, complications that are related to a defective re-endothelialization. Exendin-4, a stable GLP-1 receptor agonist has been suggested to influence the formation of intimal hyperplasia and to increase endothelial cell proliferation in vitro. Thus, the aim of the study was to investigate the mechanisms by which treatment with exendin-4 could influence re-endothelialization and intimal hyperplasia after vascular injury. Methods: 40 male Sprague-Dawley rats were subjected to balloon injury of the left common carotid artery (LCCA) and treated for four weeks with exendin-4 (1 nmol/day) or vehicle. Intimal hyperplasia and vessel wall elasticity were monitored non-invasively by high frequency ultrasound and re-endothelialization was evaluated upon sacrifice using Evans blue dye. The LCCA was then sectioned for subsequent morphometric and immunohistochemical analyses. To further investigate if and how smooth muscle cells (SMCs) are directly affected by exendin-4 treatment, we studied proliferation and apoptosis of SMCs in vitro. Results and Conclusion: Exendin-4 selectively reduced proliferation of SMCs and intimal hyperplasia in vivo without affecting the re-endothelialization process, however treatment with exendin-4 improved arterial wall elasticity. Our data also show that exendin-4 significantly decreased the proliferation and increased the apoptosis of SMCs in vitro, effects that appear to be mediated through cAMP signaling and endothelial nitric oxide synthase following GLP-1 receptor activation. Together, these effects of exendin-4 are highly desirable and may lead to improved outcome for patients undergoing vascular interventions.

Abstract 369: Protein Kinase C Epsilon-dependent Nox Activation Mediates Resistin-induced Cardiovascular Disease

Background: Adipokine resistin induces vascular smooth muscle cell (VSMC) dysfunction, implying a vital role in cardiovascular disease. In this study, we examined the in vivo and in vitro effects of resisin and investigated the underlying mechanisms involving protein kinase C epsilon (PKCε) and reactive oxygen species (ROS). Methods: After guide-wire injury of distal left common carotid artery, ApoE-/- mice fed with a western diet were treated with or without subcutaneous infusion of resistin in the presence or absence of PKCε inhibitor for 4 weeks. The progress of intimal hyperplasia was monitored using an ultrasound micro-image system. NADPH oxidase (Nox) activity in the mouse coronary arteries was measured using a chemiluminescence assay. Additionally, we treated human coronary artery smooth muscle with resistin in the presence or absence of PKCε and Nox-specific inhibitors to determine mechanistic and functional effects of resistin in VSMCs functions. Reactive oxygen species (ROS) production was analyzed using confocal microscopy. Results: Resistin significantly enhanced injury-induced intimal hyperplasia in our murine model, which was mitigated by the PKCε inhibitor. Nox activity was also substantially augmented in the injured artery in a resistin-PKCε dependent manner. On the cellular level, resistin-induced ROS production was time-dependent and mitochondrial ROS seemed to be the trigger for cytosolic Nox-induced ROS. Inhibition of Nox completely abolished resistin-exaggerated VSMC proliferation, migration and dedifferentiation. Consistently, inhibition of PKCε significantly reduced resistin-induced cytosolic ROS production, Nox activity, and VSMC dysfunction. Conclusion: This is the first study showing that resistin-induced VSMC dysfunction and intimal hyperplasia is mediated through PKCε-dependent Nox activation pathway. These findings suggest potential molecular targets for cardiovascular disease.

Glucagon-Like Peptide-1 Receptor Activation Does Not Affect Re-Endothelialization but Reduces Intimal Hyperplasia via Direct Effects on Smooth Muscle Cells in a Nondiabetic Model of Arterial Injury

Diabetic patients have an increased risk of restenosis and late stent thrombosis after angioplasty, i.e. complications that are related to a defective re-endothelialization. Exendin-4, a stable glucagon-like peptide (GLP)-1 receptor agonist, has been suggested to influence the formation of intimal hyperplasia and to increase endothelial cell proliferation in vitro. Thus, the aim of this study was to investigate the mechanisms by which treatment with exendin-4 could influence re-endothelialization and intimal hyperplasia after vascular injury. Methods: Sprague-Dawley rats were subjected to balloon injury of the left common carotid artery and treated for 4 weeks with exendin-4 or vehicle. Intimal hyperplasia and vessel wall elasticity were monitored noninvasively by high-frequency ultrasound, and re-endothelialization was evaluated upon sacrifice using Evans blue dye. Results and Conclusion: Exendin-4 selectively reduced the proliferation of smooth muscle cells (SMCs) and intimal hyperplasia in vivo without affecting the re-endothelialization process, but treatment with exendin-4 improved arterial wall elasticity. Our data also show that exendin-4 significantly decreased the proliferation and increased the apoptosis of SMCs in vitro, effects that appear to be mediated through cAMP signaling and endothelial nitric oxide synthase following GLP-1 receptor activation. Together, these effects of exendin-4 are highly desirable and may lead to an improved outcome for patients undergoing vascular interventions.

Abstract 16942: Alternatively Spliced Tissue Factor promotes plaque progression, inflammation and angiogenesis in experimental atherosclerosis

Introduction: Alternatively Spliced Tissue Factor (asTF) is novel isoform of tissue factor with angiogenic activity mediated via HIF-1α signaling (Giannarelli, AHA 13). asTF is highly expressed in human complicated atherosclerotic plaques (Giannarelli, ACC 13); however, it is unknown whether asTF has a functional role in atherogenesis. Hypothesis: asTF promotes atherosclerotic plaque progression, inflammation and angiogenesis. Methods: ApoE-/- mice (8-weeks old; n=15) were fed a Western-type diet from 2 weeks before surgery continuing through the experiment. Immediately after transluminal wire injury of the left common carotid artery (LCCA), LCCA was incubated with lentivirus encoding asTF-GFP (asTF+ group; n=10) or GFP (asTF- control group; n=5). Four weeks after, blood and spleen were collected for flow cytometry analysis of neutrophils and monocytes. LCCA was removed and processed for H&E, Oil-Red O staining and immunostaining for macrophages (MOMA-2 and MAC-3), vascular smooth muscle cells (VSMC, α-actin), endothelial cells (CD31) and HIF-1α. Results: Neointimal thickness and plaque lipid accumulation were significantly greater in asTF+ vs asTF- mice (Fig 1, A-C). An increase in plaque macrophages, neovessels and HIF-1α was observed in asTF+ vs asTF- (Fig 1, D-F). Medial thickness and VSMC density were similar between groups. Increased circulating neutrophils and Ly6C high (classical/inflammatory) monocytes were observed in asTF+ vs asTF- mice (Fig 1, G,H). In contrast, circulating Ly6C low (patrolling) monocytes were significantly reduced (Fig 1, I). Similar findings were observed in the spleen (Fig 1, J-L). Conclusions: Our results demonstrate that asTF expressed within atherosclerotic lesions promotes plaque progression towards a more advanced phenotype and is associated with systemic proinflammatory status. These data makes asTF an attractive marker of plaque vulnerability and a potential therapeutic target for plaque stabilization.

Abstract 515: Effects of the Glucagon-Like Peptide-1 Analog Exendin-4 on Reendothelialization and Intimal Hyperplasia Formation in an Animal Model of Vascular Injury

Background and aims: Diabetic patients have an increased risk of late stent thrombosis formation and restenosis, after angioplasty. It is hypothesized that delayed re-endothelization is a major underlying problem for late-stent thrombosis and also that rapid re-endothelization is essential for preventing restenosis. Exendin-4 is a stable GLP-1 receptor agonist, which is applied in clinical treatment of diabetes. Beside its insulinotropic action, it may also exert direct beneficial effects on endothelial function. We previously reported that exendin-4 is able to activate the GLP-1 receptor on human coronary artery endothelial cells, leading to increased cell proliferation and decreased apoptosis. Therefore, our aim was to study if exendin-4 can influence endothelialization and decrease neointima formation after vascular injury. Materials and Methods: Balloon injury of the left common carotid artery (LCCA) was performed on Sprague-Dawley rats. Rats were randomized into two groups and treated for four weeks with exendin-4 (1 nmol/day) or saline. Intimal hyperplasia and endothelialization was monitored non-invasively by high frequency ultrasound and upon sacrifice with Evans blue, respectively. The LCCA was then sectioned for subsequent morphometric and immunohistochemical analyses. To further investigate if and how smooth muscle cells (SMCs) are directly affected by exendin-4 treatment, we studied proliferation and apoptosis of SMCs in vitro . Results and conclusions: Our findings show that exendin-4 selectively reduces SMC proliferation and intimal hyperplasia through activation of cAMP signaling and endothelial nitric oxide synthase without influencing re-endothelialization. Treatment with exendin-4 did however improve arterial wall elasticity. Together, these effects of exendin-4 are highly desirable and may lead to improved outcome for patients undergoing vascular interventions

Early Changes of Gene Expression Profiles in the Rat Model of Arterial Injury

PurposeRestenosis caused by intimal hyperplasia (IH) remains a significant drawback for vascular interventions. It is crucial to understand the molecular mechanisms that control activation of smooth muscle cells (SMCs) after the injury in order to develop strategies to prevent IH. The purpose of the present study was to investigate the early alterations in arterial-wall gene expression after balloon injury in the rat carotid artery with focus on the induction of an inflammatory response. Materials and MethodsTwenty-four male Sprague–Dawley rats were subjected to injury of the left common carotid artery by using a 2-F Fogarty catheter. The arteries were harvested 5, 10, and 20 hours after injury. Uninjured arteries from an additional eight rats were used as controls. RNA was isolated and used for genome-wide microarray expression analysis, followed by validation of selected genes with quantitative real-time polymerase chain reaction (qRT-PCR). Immunohistochemistry was performed on the cross-sectioned vessels. ResultsAnalysis of gene expression by microarrays showed that the most differentially expressed genes were primarily associated with inflammation, cell proliferation, migration, and adhesion. As confirmed by qRT-PCR, microarray data showed a significant (P < .005) upregulation of cytokines and chemokines (IL-6, CCL2, CXCL1, AIMP1, and CD44) just 5 hours after injury. Immunohistochemistry demonstrated that CCL2 and the adhesion receptor CD44 were expressed by SMCs in the early response to injury and in the absence of leukocyte infiltration. ConclusionsArterial injury is followed by an early induction of inflammatory genes in the vessel wall that appears to be confined to SMCs.

Correlation of restenosis after rabbit carotid endarterectomy and inflammatory cytokines

ObjectiveTo establish rabbit model of restenosis after carotid endarterectomy surgery, and to study tissue inflammatory cytokines (TNF-α, IL-6) involved in restenosis. MethodsA total of 32 rabbits were randomly divided into two groups: model group and control group. The right common carotid artery in rabbits was damaged by carotid endar terectomy in model group. The tissues were harvested at different time points respectively, the pathological changes of the vascular wall after operation were observed at different time points. The changes of expression of tissue vascular wall inflammatory cytokines (TNF-α, IL-6) at different time points after the surgery was observed by RT-PCR, and the changes of serum inflammatory cytokines (TNF-α, IL −6) were detected by ELISA. ResultsThe new intima appeared after 7 days of the injury and reached the peak on 28 d which is uneven and significantly thicker than the control group (P<0.01). The tissue inflammatory cytokines (TNF-α, IL-6) were significantly increased after the rabbit common carotid artery injury, which was significant difference compared with normal control group (P<0.05). ConclusionsThe tissue inflammatory factors significantly increase after the rabbit carotid artery injury, which suggests the mutual concurrent effects of inflammatory cytokines can result in the proliferation of vascular restenosis.

Common carotid artery injury caused by a camel bite: case report and systematic review of the literature

A 25-year-old man was bitten in the neck by an aggressive camel, causing three small puncture wounds. The left carotid pulse of the patient was weakly palpated. Angiography showed irregular dissection of the distal part of the left common carotid artery. Neck exploration confirmed the findings. An interposition autogenous saphenous vein graft was performed successfully. The patient was discharged home in good condition. We have systematically reviewed the literature on this topic, and only four other similar cases were reported previously. Although camel bite wounds are small, they may penetrate deeply, causing serious injuries to the neck structures including the major vessels. Care should be taken when approaching male camels during the rutting season.

GW24-e2536 Effects of Captopril on Plaque Inflammation and Atherosclerosis Progression Evaluated by FDG-PET-CT and IVUS Imagines

ObjectivesMacrophages play an important role in the pathogenesis of atherosclerosis. Captopril is one kind of the effictive drugs to the atherosclerosis besides statins. However, how does macrophage change in the...

GW24-e3141 To Evaluation The Progression of Atherosclerosis by Contrast Enhanced Ultrasound and Intravascular Ultrasound Imaging

ObjectivesPlaque neovascularisation and the extent of adventitial vasa vasorum (VV) is closely associated with vulnerability of atherosclerotic plaque. To rigorously investigate the relationship between vasa vasorum (VV) proliferation and plaque...

The Effect of Short-term Intra-arterial Delivery of Paclitaxel on Neointimal Hyperplasia and the Local Thrombotic Environment after Angioplasty

To evaluate the effects of short-term intra-arterial delivery of paclitaxel on neointimal hyperplasia and the local thrombotic environment after angioplasty. An experimental common carotid artery injury model was established in 60 rats, which were divided into experimental groups (40 rats) and controls (20 rats). Local intra-arterial administration of paclitaxel was applied at 2 doses (90 and 180μg/30μl), and the effects of short-term delivery of paclitaxel on neointimal hyperplasia and the expression of tissue factor (TF), plasminogen activator inhibitor-1 (PAI-1) and tissue-type plasminogen activator (t-PA) were evaluated at days 15 and 30 by hematoxylin and eosin staining and immunohistochemistry. At 15 and 30days after injury, neointimal thickness and area, the ratio of intimal area to medial area and the stenotic rate were all significantly decreased in the group provided the high concentrations (180μg/30μl) of paclitaxel for 2min or 10min and in the group provided the low concentration (90μg/30μl) of paclitaxel for 10min (p<0.05). At 30days after injury, there were no significant changes in TF expression among all experimental groups. PAI-1 expression increased in the neointima of the high concentration 10min group (p<0.05), while t-PA expression decreased in the neointima of the high concentration 2min group (p<0.05). In the rat common carotid artery injury model, the short-term delivery of paclitaxel could effectively inhibit neointimal hyperplasia in the long term, with very little influence on the local expression of TF and PAI-1.

Ultrasound analysis of the relationship between right internal jugular vein and common carotid artery in the left head-rotation and head-flexion position

Common carotid artery (CCA) injury is a serious complication of internal jugular vein (IJV) cannulation. To minimize unintentional CCA puncture, the anatomic relationship between the IJV and the CCA and the size of IJV were compared under different head positions. Ultrasound analyses of the IJV and the CCA were performed in 103 consecutive patients. Overlapping angle (OA), real puncture angle (RPA) and diameter of IJV (D IJV) were evaluated with 30° and 60° left rotation and with 30° left flexion. When the head position was changed from 30° left rotation to 60° left rotation, OA increased significantly from 6.5° ± 7.7° to 14.5° ± 7.4° at the cricoid cartilage level (Cricoid-level) and from 14.4° ± 8.4° to 20.6° ± 6.9° at the middle triangle level (Triangle-level), whereas RPA decreased significantly at these levels (from 49.7° ± 11.9° to 43.5° ± 13.1° and from 51.1° ± 14.4° to 44.3° ± 13.9°, respectively; P < 0.01 for both). When the head position was changed from 30° left rotation to 30° left flexion, neither OA nor RPA significantly changed (OA: 6.3° ± 6.1° and 15.0° ± 7.2°, RPA: 48.5° ± 12.4° and 51.8° ± 13.6°, P not significant vs 30° left rotation). There was no difference in D IJV when comparing 30° left rotation and 30° left flexion, although D IJV was largest at 60° left rotation. RPA positively correlated with age, and D IJV positively correlated with body mass index. In conclusion, excessive left rotation should be avoided to minimize the probability of unintentional CCA puncture during IJV cannulation. When 30° left rotation is not feasible, the head-flexion position should be utilized.

Hybrid Stent-Graft Repair of an Iatrogenic Complex Proximal Right Common Carotid Artery Injury

Iatrogenic carotid trauma requires early diagnosis and adequate treatment. Classic open repair may be technically challenging if trauma is in base of the neck. We present a case of an iatrogenic carotid pseudoaneurysm treated with endovascular repair. An 87-year-old woman presented with a pulsatile neck mass 10 days after coronary artery bypass graft surgery. A computed tomographic angiogram showed a 1.6 × 1.0 × 2.0-cm pseudoaneurysm arising from the posterior wall of the proximal right common carotid artery. Endovascular management was considered, and a percutaneous angiogram demonstrated an arteriovenous fistula in addition to the pseudoaneurysm. Through a cervical cut-down, retrograde percutaneous access was obtained through the common carotid artery, which allowed easier access to the area of trauma owing to vessel tortuosity. Subsequently, a 5 mm × 2-cm Viabahn was deployed. The postdilation angiogram showed a significant endoleak that kept filling the pseudoaneurysm. A second 6mm × 5-cm Viabahn was placed and successfully postdilated with a 6 mm × 4-cm balloon. No endoleaks or fistulas were noted on the completion angiogram. The patient remains asymptomatic after 15 months. Follow-up images showed thrombosis of pseudoaneurysm. Endovascular treatment with self-expanding stent-grafts and open cut-down access are excellent options to treat major vessel injuries at the base of the neck, where anatomy and cumbersome access make open surgery a more difficult option.

Cinaciguat prevents neointima formation after arterial injury by decreasing vascular smooth muscle cell migration and proliferation

AimsVascular smooth muscle cell (VSMC) migration, proliferation and remodeling of the extracellular matrix contribute to lumen loss after arterial injury leading to restenosis. Several studies indicated the role of the cyclic guanosine monophosphate signaling in neointimal formation. Cinaciguat, the novel soluble guanylate cyclase activator, currently being in phase IIb clinical trial, has been shown to exert antiplatelet and anti-remodeling effects in animal models of vascular pathology. In this study we investigated the effects of cinaciguat on post-injury arterial stenosis. Methods and ResultsMale Sprague-Dawley rats (n=100) underwent endothelial denudation by wire injury of the right common carotid artery. Cinaciguat (10mg/kg/day orally) were administered to 50 rats (1-, 2-, 3-day and 1-, 3-week treatment time), while 50 rats received placebo. A 3-week treatment resulted in a significantly reduced vascular stenosis (17.53±10.84% in the treatment group vs. 43.25±30.83% in the control wire injury group) and neointima/media area ratio (0.45±0.32 in the treatment group vs. 1.09±0.69 in the control wire injury group). By using quantitative real-time PCR, Western blot and immunohistochemistry, matrix-metallopreoteinase-9 (MMP-9) was found to be upregulated in the control-injured carotids over the whole follow-up, and cinaciguat significantly decreased MMP-9 expression by 3weeks. As assessed by protein immunoblot, injury-induced local decrease of soluble guanylate cyclase β1 subunit could be recovered by cinaciguat. In vitro wound healing assay with VSMCs revealed dose-dependent antimigratory and antiproliferative effects of cinaciguat. Plasma level of cyclic guanosine monophosphate was significantly elevated after 3weeks of treatment. ConclusionOur results show that cinaciguat prevents injury-induced neointimal hyperplasia by decreasing VSMC migration and proliferation through the regulation of MMP-9.

Effects of SDF-1α/CXCR4 on vascular smooth muscle cells and bone marrow mesenchymal cells in a rat carotid artery balloon injury model

Summary Bone marrow mensenchyme cells (BMSCs) can differentiate into endothelial progenitor cells which then migrate to injured sites for the repair of neointima, and stromal cell-derived factor-1α (SDF-1α) can mediate the migration of CXCR4 expressing stem/progenitor cells to injured sites for pair. Protein and mRNA expression of SDF-1α and CXCR4 were determined by RT-PCR, Western blot and ELISA. Immediately after common carotid artery balloon injury, the mRNA expression of SDF-1α in vascular smooth muscle cells (VSMCs) first increased and then decreased 7 days later. VSMCs transfected with SDF-1α siRNA did not express SDF-1α mRNA, but after transfection with SDF-1α siRNA, the SDF-1α content in injured VSMCs gradually returned to the baseline level. Normal BMSCs rarely expressed CXCR4 mRNA, but the CXCR4 mRNA expression on BMSCs increased significantly 4 days after common carotid artery injury and was maintained. The migration of BMSCs after artery injury was enhanced when compared with normal BMSCs, but SDF-1α siRNA transfection of VSMCs and AMD3100 treatment remarkably decreased the chemotaxis of BMSCs to VSMCs and SDF-1α, respectively. We conclude that the SDF-1α/CXCR4 axis plays an important role in the migration of BMSCs after balloon injury and can ultimately cause abnormal proliferation of the intima.

Upregulation of Nox4 by TGFβ1 Oxidizes SERCA and Inhibits NO in Arterial Smooth Muscle of the Prediabetic Zucker Rat

Vascular smooth muscle cell (SMC) migration is an important pathological process in several vascular occlusive diseases, including atherosclerosis and restenosis, both of which are accelerated by diabetes mellitus. To determine the mechanisms of abnormal vascular SMC migration in type 2 diabetes, the obese Zucker rat (ZO), a model of obesity and insulin resistance, was studied. In culture, ZO aortic SMCs showed a significant increase in Nox4 mRNA and protein levels compared with the control lean Zucker rat (ZL). The sarco-/endoplasmic reticulum Ca(2+) ATPase (SERCA) nitrotyrosine-294,295 and cysteine-674 (C674)-SO(3)H were increased in ZO SMCs, indicating oxidant stress. Unlike ZL SMC, nitric oxide (NO) failed to inhibit serum-induced SMC migration in ZO. Transfection of Nox4 small interference RNA or overexpression of SERCA2b wild type, but not C674S mutant SERCA, restored the response to NO. Knockdown of Nox4 also decreased SERCA oxidation in ZO SMCs. In addition, transforming growth factor-β1 via Smad2 was necessary and sufficient to upregulate Nox4, oxidize SERCA, and block the antimigratory action of NO in ZO SMCs. Corresponding to the results in cultured SMCs, immunohistochemistry confirmed that Nox4 and SERCA C674-SO(3)H were significantly increased in ZO aorta. After common carotid artery injury, knockdown of Nox4 by adenoviral Nox4 short hairpin RNA decreased Nox4 and SERCA C674-SO(3)H staining and significantly decreased injury-induced neointima. These studies indicate that the upregulation of Nox4 by transforming growth factor-β1 in ZO SMCs is responsible for the impaired response to NO by a mechanism involving the oxidation of SERCA C674. Knockdown of Nox4 inhibits oxidation of SERCA, as well as neointima formation, after ZO common carotid artery injury.