Clinical Relevance of Patent Foramen Ovale and Atrial Septum Aneurysm in Stroke: Findings of a Single-Center Cross-Sectional Study.

BACKGROUND: A significant proportion of ischemic strokes are cryptogenic. In this context, the clinical pertinence of patent foramen ovale (PFO) with and without atrial septum aneurysm (ASA) remains controversial. The aim of this study was to identify how PFO +/-ASA and cryptogenic stroke are associated in a representative sample of stroke patients. METHODS: We enrolled all patients (n = 909) with ischemic stroke or transient ischemic attack admitted to the certified stroke unit or neurological intensive care unit of our university medical center who underwent transesophageal echocardiography (TEE) between 2012 and 2014. The baseline characteristics, cardio-/neurovascular risk factors, clinical parameters and TEE findings were analyzed. RESULTS: PFO was present in 26.2%, and PFO was combined with an ASA in 9.9%. In cryptogenic stroke, the prevalence of PFO was higher compared to other etiologies (30.9 vs. 21.9%; p < 0.002). Patients with PFO had lower National Institute of Health Stroke Score (NIHSS) values at admission than those without (2 [0-5] vs. 3 [1-7]; p = 0.001; 95% CI [0.62-0.88]). No difference was found in NIHSS values of PFO patients with or without ASA (2 [0-5] vs. 2 [0-5]; p = 0.683; 95% CI 0.94 [0.68-1.28]). CONCLUSIONS: Our study indicates that a detected PFO +/-ASA could exhibit a stroke-relevant finding, if classical risk factors for the stroke were lacking.

In vitro and in vivo effects of human monocytes and their subsets on new vessel formation.

OBJECTIVE: Human monocytes can be divided into CD16(-) monocytes and CD16(+) monocytes. Studies in mice suggested differential effects of monocyte subsets during new vessel formation. METHODS: The functional role of human monocyte subsets in neovascularization processes was investigated. For in vivo experiments, nude mice underwent unilateral hindlimb ischemia surgery before being injected with either total monocytes, CD16(-) monocytes or CD16(+) monocytes isolated from healthy individuals. RESULTS: In vitro, cytokine array analysis demonstrated that monocytes release numerous angiogenic cytokines, some of which were differentially expressed in monocyte subsets. Sprout length was enhanced in EC spheroids being cultured in conditioned medium obtained from total monocytes and, to a lesser extent, also in supernatants of CD16(-) monocytes. Laser Doppler perfusion imaging up to day 28 after surgery revealed a trend toward improved revascularization in mice treated with monocytes, but no significant differences between monocyte subsets. Histological analyses four weeks after surgery showed an increased arteriole size in mice having received CD16(+) monocytes, whereas the number of capillaries did not significantly differ between groups. CONCLUSIONS: Our findings suggest additive and differential effects of monocyte subsets during neovascularization processes, possibly due to an altered secretion of angiogenic factors and their paracrine capacity to stimulate new vessel formation.

Leptin-dependent and leptin-independent paracrine effects of perivascular adipose tissue on neointima formation.

OBJECTIVE: Clinical and experimental evidence suggests that periadventitial adipose tissue may modulate vascular lesion formation. The aim of this study was to determine the role of perivascular leptin expression on neointima formation and to differentiate it from local inflammation and systemically elevated leptin levels. APPROACH AND RESULTS: Increased neointima formation after carotid artery injury was observed in hyperleptinemic, diet-induced obese wild-type mice, but not in leptin-deficient ob/ob mice. High-fat diet was associated with increased leptin expression in visceral adipose tissue (VAT) as well as in perivascular adipose tissue. Perivascular leptin overexpression achieved by adenoviral vectors enhanced intimal cell proliferation and neointima formation in wild-type mice, but not in leptin receptor-deficient mice. Perivascular transplantation of VAT from high-fat diet-induced obese wild-type mice around the carotid artery of immunodeficient mice also promoted neointima formation, without affecting body weight or systemic leptin levels, and this effect was absent, if VAT from ob/ob mice was used. On the contrary, perivascular transplantation of VAT from ob/ob mice fed high-fat diet, characterized by marked immune cell accumulation, promoted neointimal hyperplasia also in the absence of leptin. In vitro, recombinant leptin and VAT-conditioned medium increased human arterial smooth muscle cell proliferation in a (partly) leptin-dependent manner. CONCLUSIONS: Our findings suggest that locally elevated leptin levels may promote neointima formation, independent of obesity and systemic hyperleptinemia, but also underline the importance of perivascular inflammation in mediating the increased cardiovascular risk in obesity.

Fulminant Lyme myocarditis without any other signs of Lyme disease in a 37-year-old male patient with microscopic polyangiitis-a case report.

Background: Lyme disease is a tick-borne multisystem infection. The most common cardiac manifestation is an acute presentation of Lyme carditis, which often manifests as conduction disorder and rarely as myocarditis. Case summary: We report the case of a 37-year-old male with a history of microscopic polyangiitis receiving immunosuppressive therapy. He was admitted for severe dyspnoea secondary to acute heart failure. Echocardiography and cardiac magnetic resonance imaging indicated a severely reduced left ventricular ejection fraction (LVEF) with global hypokinesia. Coronary heart disease was excluded, and endomyocardial biopsies (EMB) were performed. The left ventricular EMB revealed a rare case of fulminant Lyme carditis with evidence of typical lymphocytic myocarditis. Borrelia afzelii-DNA was detected without any relevant atrioventricular blockage or systemic signs of Lyme disease. The patient had no clinically apparent tick-borne infection or self-reported history of a tick bite. Immunological testing revealed a positive ELISA and Immunoblot for anti-Borrelia immunoglobulin G antibodies. After specific intravenous antibiotic therapy and optimized medical therapy for heart failure, the LVEF recovered, and the patient could be discharged in an improved condition. Repeat EMB a few months later revealed a dramatic regression of the cardiac inflammation and absence of Borrelia DNA in the myocardium. Discussion: A severely reduced LVEF can be the primary manifestation of Lyme disease even without typical systemic findings and can have a favourable prognosis with antibiotic treatment. A thorough workup for Lyme carditis is required in patients with unexplained heart failure, particularly with EMB, especially in immunosuppressed patients.

Overexpression of integrin beta 5 enhances the paracrine properties of circulating angiogenic cells via Src kinase-mediated activation of STAT3.

OBJECTIVE: To determine the intracellular mechanisms mediating the angiogenic effects of integrin alpha v beta 5 overexpression in circulating angiogenic cells (CACs). METHODS AND RESULTS: Integrin alpha v beta 5 is expressed on angiogenic endothelial cells, and integrin alpha v beta 5 activation was shown to improve the reparative functions of endothelial progenitors within the cardiovascular system. CACs were transiently transfected with the full-length cDNA of human integrin beta 5 (CAC-ITGB5) or control-vector (CAC-vector). Integrin beta 5 overexpression was confirmed using flow cytometry, Western blot, and PCR analysis; it enhanced the angiogenic capacities of CACs in vitro (spheroid and Matrigel angiogenesis assay) and stimulated new vessel formation in vivo (murine hind limb ischemia model). Overexpression of ITGB5 resulted in integrin alpha v beta 5 phosphorylation and activation of Src kinase and signal transducer and activator of transcription (STAT) 3. Furthermore, elevated mRNA and protein expression of the CXC chemokine CXCL8 and the CC chemokine CCL2 was detected in CAC-ITGB5, and conditioned medium from CAC-ITGB5 enhanced the sprouting of coincubated human endothelial cells in a STAT3-, CXCL8-, and CCL2-dependent manner. CONCLUSIONS: Src kinase-mediated activation of STAT3 and subsequent angiogenic gene expression mediate the effects of integrin alpha v beta 5 and may be exploited to enhance the paracrine activities of CACs.

Leptin enhances the potency of circulating angiogenic cells via src kinase and integrin (alpha)vbeta5: implications for angiogenesis in human obesity.

OBJECTIVE: To investigate the capacity of the adipokine leptin to promote angiogenesis by modulating the function of circulating angiogenic cells (CACs). METHODS AND RESULTS: In vitro, leptin specifically promoted CAC adhesion to tubular endothelial structures and migration along outgrowing sprouts of endothelial cells. In vivo, stimulation of CACs with leptin increased their capacity to promote new vessel formation in the chorioallantoic membrane of chicken embryos and to improve neovascularization of ischemic murine hind limbs. These effects required the phosphorylation of alphavbeta5 integrins, which depended on the interaction of leptin with its receptor ObR, and on Janus kinase (JAK) 2- and phospholipase C (PLC) gamma-mediated activation of Src kinase. Protein tyrosine phosphatase 1B, a negative regulator of leptin signaling, was overexpressed in CACs from obese, hyperleptinemic individuals, and this was associated with insensitivity of CACs to the angiogenic effects of leptin. Weight loss (by 30+/-15 kg) normalized protein tyrosine phosphatase 1B expression in CACs and restored their responsiveness to leptin. A similar dose-dependent response was found after incubation of CACs from obese subjects with a protein tyrosine phosphatase 1B inhibitor ex vivo. CONCLUSIONS: Our results point to the ObR-Src kinase-alphavbeta5 cross talk as a distinct novel component of the network of specific interactions between integrins and cytokine receptors in angiogenesis.

Leptin enhances the recruitment of endothelial progenitor cells into neointimal lesions after vascular injury by promoting integrin-mediated adhesion.

The adipocytokine leptin modulates vascular remodeling and neointima formation. Because endothelial progenitor cells (EPCs) participate in vascular repair, we analyzed the effects of leptin on human EPC function in vitro and in vivo. After 7 days in culture, EPCs expressed the leptin receptor and responded to leptin stimulation with increased STAT3 phosphorylation. Incubation of EPCs with leptin (at concentrations between 1 and 100 ng/mL) increased the number of EPCs adhering to vitronectin and fibronectin in a receptor-specific manner. It also enhanced the capacity of EPCs to incorporate into a monolayer of human endothelial cells and the adherence of these cells to activated platelets. Leptin upregulated alphavbeta5 and alpha4 integrin expression in EPCs, and the effects of leptin on EPC function could be prevented, at least in part, by RGD peptides and function-blocking antibodies. Intravenous injection of fluorescently labeled human EPCs into athymic nude mice shortly after vascular injury revealed that preincubation of EPCs with leptin augmented their accumulation within intimal lesions, accelerating reendothelialization and decreasing neointima formation in an alphavbeta5 and alpha4 integrin-dependent manner. Our findings suggest that leptin specifically modulates the adhesive properties and the homing potential of EPCs and may thus enhance their capacity to promote vascular regeneration in vivo.

Heart Failure Is Not Associated with a Poor Outcome after Mechanical Thrombectomy in Large Vessel Occlusion of Cerebral Arteries.

The impact of heart failure on outcome in stroke patients is not fully understood. There is evidence for an increased mortality and morbidity, but it remains uncertain whether thrombectomy in patients with large vessel occlusion (LVO) in the anterior circulation is less effective in patients with heart failure compared to patients without. Retrospectively, we analyzed echocardiographic data of all patients in our stroke database, who underwent mechanical thrombectomy (n=668) for the presence of heart failure. Furthermore, we collected baseline characteristics and neurological and neuroradiological parameters. In the analysis, 373 of the 668 patients of our stroke database underwent echocardiography. Of these 373 patients, 90 patients (24%) suffered from heart failure with reduced left ventricular ejection fraction measured by echocardiography according to the current guidelines. After adjustment for age, the Alberta stroke program early CT score (ASPECTS), and time from symptom onset to recanalization, the analysis revealed that thrombectomy in patients with heart failure and LVO is not associated with less favorable outcome measured by the modified Rankin Scale after 90 days (3 (0-6) vs. 3 (1-5); p=0.380). Moreover, we could not find a significant difference in mortality compared to patients without heart failure (11.0% vs. 7.4%; p=0.313).

Low flow in the left atrial appendage assessed by transesophageal echocardiography is associated with increased stroke severity-Results of a single-center cross-sectional study.

BACKGROUND: About 25% of strokes are thought to be cryptogenic. There is growing evidence that most of these cryptogenic strokes are thromboembolic and caused by an undetected atrial fibrillation. Measured slow flow in the left atrial appendage has been proposed to be an indicator for the thromboembolic risk since it is possibly associated with undetected atrial fibrillation. METHODS: We evaluated all patients (n = 909; mean of 62 years, 39% female) admitted with an ischemic stroke (84%) or transient ischemic attack to our university medical center who underwent transesophageal echocardiography examination 2012-2014. Baseline characteristics, cardio-/neurovascular risk factors, electrocardiogram monitoring data, National Institute of Health Stroke Scale values, the localization of the stroke, as well as transesophageal echocardiography findings were analyzed. RESULTS: Stroke severity measured by the National Institute of Health Stroke Scale in patients with a low flow in the left atrial appendage was significantly higher than in patients without left atrial appendage low flow (p < 0.001). There was a threshold phenomenon in flow velocity at about 60 cm/s above which no significant change in National Institute of Health Stroke Scale values could be observed. These patients were significantly different concerning incidence of heart failure (10.6% vs. 2.3%; p < 0.001) and atrial fibrillation (32.2% vs. 8.8%; p < 0.001). However, threshold phenomenon of left atrial appendage flow was unchanged when excluding atrial fibrillation and heart failure patients. CONCLUSIONS: A low flow in the left atrial appendage was associated with clinically elevated stroke severity and could be indicated as an independent stroke risk factor. Further prospective studies are warranted to evaluate beneficial effects of therapeutic options in these patients also in the assumed absence of atrial fibrillation.

Cigarette smoke exposure promotes arterial thrombosis and vessel remodeling after vascular injury in apolipoprotein E-deficient mice.

BACKGROUND: Cigarette smoking is a major risk factor for the development of cardiovascular disease. However, in terms of the vessel wall, the underlying pathomechanisms of cigarette smoking are incompletely understood, partly due to a lack of adequate in vivo models. METHODS: Apolipoprotein E-deficient mice were exposed to filtered air (sham) or to cigarette mainstream smoke at a total particulate matter (TPM) concentration of 600 microg/l for 1, 2, 3, or 4 h, for 5 days/week. After exposure for 10 +/- 1 weeks, arterial thrombosis and neointima formation at the carotid artery were induced using 10% ferric chloride. RESULTS: Mice exposed to mainstream smoke exhibited shortened time to thrombotic occlusion (p < 0.01) and lower vascular patency rates (p < 0.001). Morphometric and immunohistochemical analysis of neointimal lesions demonstrated that mainstream smoke exposure increased the amount of alpha-actin-positive smooth muscle cells (p < 0.05) and dose-dependently increased the intima-to-media ratio (p < 0.05). Additional analysis of smooth muscle cells in vitro suggested that 10 microg TPM/ml increased cell proliferation without affecting viability or apoptosis, whereas higher concentrations (100 and 500 microg TPM/ml) appeared to be cytotoxic. CONCLUSIONS: Taken together, these findings suggest that cigarette smoking promotes arterial thrombosis and modulates the size and composition of neointimal lesions after arterial injury in apolipoprotein E-deficient mice.

Leptin receptor is elevated in carotid plaques from neurologically symptomatic patients and positively correlated with augmented macrophage density.

BACKGROUND: Carotid artery lesions from symptomatic patients are characterized by inflammation and neovascularization. The adipokine leptin promotes angiogenesis and activates inflammatory cells, and the leptin receptor (ob gene-encoded receptor), ObR, is expressed in advanced atherosclerotic lesions. The present study quantitatively analyzed ObR messenger RNA (mRNA) expression and immunoreactivity in carotid artery plaques from symptomatic and asymptomatic persons. Plaque angiogenesis, gene expression of vascular endothelial growth factor (VEGF), and macrophage density were also analyzed. METHODS: Carotid endarterectomy specimens were collected from 26 patients undergoing surgery for hemispheric cerebrovascular symptoms (n = 13) or progressive asymptomatic internal carotid stenosis (n = 13). A representative sample, including part of the most active site, was collected from each lesion and evaluated by real-time polymerase chain reaction analysis for ObR(long) and ObR(common) isoforms, VEGF(165), and macrophage adhesion molecule-1 (Mac-1) mRNA, and by immunohistochemistry for ObR, von Willebrand factor (vWF), and CD68 antigen expression. RESULTS: All plaques exhibited advanced atherosclerosis (American Heart Association class IV through VI). Transcript levels were preferentially elevated in symptomatic plaques for ObR(long) (P = .0006) and ObR(common) (P = .033), with a simultaneous upregulation of VEGF(165) (P = .001) and Mac-1 mRNA expression (P = .003). Immunohistochemical analysis confirmed a significant increase of ObR antigen levels (P = .011) and CD68-positive inflammatory cells (P = .049) in symptomatic plaques, whereas neovascularization, evident in all plaques, was similar in both groups (P = .7). CONCLUSION: The ObR(long) and ObR(common) genes are upregulated and their protein preferentially synthesized in clinically symptomatic carotid plaques. Moreover, ObR expression is positively correlated with augmentation of gene transcripts related to macrophage density and neovascularization. These data suggest that ObR(long) and ObR(common) may be linked with histologic features of carotid plaque instability, which are associated with cerebral ischemic symptoms.

Plasminogen activator inhibitor-1 from bone marrow-derived cells suppresses neointimal formation after vascular injury in mice.

OBJECTIVE: To investigate the ability of bone marrow (BM)-derived cells to modulate neointimal growth after injury by expressing plasminogen activator inhibitor-1 (PAI-1). METHODS AND RESULTS: We performed BM transplantation (BMT) in lethally irradiated wild-type (WT) and PAI-1(-/-) mice. Three weeks after carotid injury with ferric chloride, analysis of Y-chromosome DNA expression in the vessel wall of female hosts revealed that 20.8+/-6.0% of the cells in the neointima and 37.6+/-5.7% of those in the media were of BM origin. Lack of PAI-1 in either the host or the donor cells did not affect recruitment of BM-derived cells into sites of vascular injury. The neointima consisted predominantly of smooth muscle cells, and a proportion of these cells expressed PAI-1. Overall, lack of PAI-1 was associated with enhanced neointimal formation. However, importantly, BMT(WT-->PAI-1(-/-)) mice exhibited reduced neointimal area (P=0.05) and luminal stenosis (P=0.04) compared with BMT(PAI-1(-/-)-->PAI-1(-/-)) mice. Although PAI-1-expressing cells were shown to be present in BMT(WT-->PAI-1(-/-)) lesions, these mice did not exhibit detectable levels of the inhibitor in the circulation, suggesting that local production of PAI-1 by cells in the neointima and media was sufficient to reduce luminal stenosis. CONCLUSIONS: PAI-1 from BM-derived cells appears capable of suppressing neointimal growth after vascular injury.

Mid- to long-term outcome of patients treated with everolimus-eluting bioresorbable vascular scaffolds: Data of the BVS registry Göttingen predominantly from ACS patients.

BACKGROUND: Bioresorbable vascular scaffolds (BVS) are widely used in routine clinical practice. While previous studies reported acceptable short- to midterm outcome after BVS implantation, data on longer-term outcome are rare. METHODS: Patients treated with at least one Absorb®-BVS were consecutively enrolled. Follow-up data were assessed after 834.0 [769.0-1026.0] days. The primary device-oriented composite endpoint (DOCE) was defined as cardiovascular death, myocardial infarction (MI) and/or target lesion revascularization (TLR). RESULTS: Between 2012 and 2014, 195 patients were included into study analysis. Overall, 244 BVS were implanted. Mean patient age was 64.0[54.3-74.0] years. Three-quarter of patients had an ACS; of those 42.9% had ST-elevation-MI and 40.8% had non-ST-elevation-MI. DOCE occurred in 3.1%, 6.7%, 11.8% and 15.4% of patients during hospital stay, within 6-months, 18-months or during the complete follow-up period, respectively. In those patients, median time until DOCE was 211.5[43.25-567.25] days. In 11 (36.7%) patients DOCE occurred after >12months. Using univariable analysis, bifurcation stenting was associated with a hazard ratio (HR) of 11.8[2.38-58.57] for TLR (p=0.002) and 2.1[1.02-4.49] for DOCE (p=0.045). Similarly, in ACS patients, bifurcation stenting was associated with an increased risk for TLR (HR=10.4[2.01-53.56]; p=0.005) and for DOCE (HR=2.4[1.09-5.32]; p=0.029) and in multivariable analysis, it remained an independent predictor of DOCE (HR=3.0; p=0.018). CONCLUSIONS: Although, the rates of (potentially) device-related complications following BVS implantation are acceptable, they are nonetheless not negligible. Interestingly, they did not decline over time. Bifurcation stenting could be found as relevant procedure-related predictor of DOCE, especially in ACS patients. Randomized trials are warranted to confirm these findings.

Successful Left Atrial Appendage Occlusion with the New Generation Amulet® Device after Late-Occurring Embolization of an Amplatzer® Cardiac Plug in a Patient with Repetitive Strokes.

The Amplatzer Cardiac Plug (ACP) is one of the most commonly used devices for percutaneous left atrial appendage (LAA) closure in order to prevent a stroke in patients with atrial fibrillation and contraindication for long-term oral anticoagulation therapy. We have previously described a patient who had experienced an embolization of the ACP device about 12 months after implantation and the device could be percutaneously retrieved. A few years later, he suffered from a posterior stroke and a stroke located in the brainstem as well as a transischemic attack (TIA). In order to protect him from further cardioembolic events a reocclusion of the LAA with the new generation of ACP device, the Amplatzer Amulet, was performed. A stable position of the device within follow-up period could be confirmed and the patient was free of additional strokes/TIA or bleeding events. This case stresses the importance of proper LAA sizing in order to prevent device embolization and notes that LAA size is not static. Moreover, it demonstrates that repeated implantation of an LAA occlusion device was still possible; one should be aware of undersizing the LAA dimensions and that the modifications of new generation LAA occlusion devices may overcome limitations of first-generation devices in order to prevent a cardioembolic stroke.

Endothelial deletion of protein tyrosine phosphatase-1B protects against pressure overload-induced heart failure in mice.

AIMS: Cardiac angiogenesis is an important determinant of heart failure. We examined the hypothesis that protein tyrosine phosphatase (PTP)-1B, a negative regulator of vascular endothelial growth factor (VEGF) receptor-2 activation, is causally involved in the cardiac microvasculature rarefaction during hypertrophy and that deletion of PTP1B in endothelial cells prevents the development of heart failure. METHODS AND RESULTS: Cardiac hypertrophy was induced by transverse aortic constriction (TAC) in mice with endothelial-specific deletion of PTP1B (End.PTP1B-KO) and controls (End.PTP1B-WT). Survival up to 20 weeks after TAC was significantly improved in mice lacking endothelial PTP1B. Serial echocardiography revealed a better systolic pump function, less pronounced cardiac hypertrophy, and left ventricular dilation compared with End.PTP1B-WT controls. Histologically, banded hearts from End.PTP1B-KO mice exhibited increased numbers of PCNA-positive, proliferating endothelial cells resulting in preserved cardiac capillary density and improved perfusion as well as reduced hypoxia, apoptotic cell death, and fibrosis. Increased relative VEGFR2 and ERK1/2 phosphorylation and greater eNOS expression were present in the hearts of End.PTP1B-KO mice. The absence of PTP1B in endothelial cells also promoted neovascularization following peripheral ischaemia, and bone marrow transplantation excluded a major contribution of Tie2-positive haematopoietic cells to the improved angiogenesis in End.PTP1B-KO mice. Increased expression of caveolin-1 as well as reduced NADPH oxidase-4 expression, ROS generation and TGFß signalling were observed and may have mediated the cardioprotective effects of endothelial PTP1B deletion. CONCLUSIONS: Endothelial PTP1B deletion improves cardiac VEGF signalling and angiogenesis and protects against chronic afterload-induced heart failure. PTP1B may represent a useful target to preserve cardiac function during hypertrophy.

Uncommon delayed and late complications after percutaneous left atrial appendage closure with Amplatzer(®) Cardiac Plug.

AIMS: Percutaneous left atrial appendage closure with Amplatzer(®) Cardiac Plug (St. Jude Medical Inc.) for the prevention of stroke in patients with atrial fibrillation is rapidly propagating. We sought to provide additional safety data. METHODS AND RESULTS: We have screened our database of patients having been treated with Amplatzer(®) Cardiac Plug and found 3 cases with uncommon complications that have not been reported previously. One patient experienced an embolisation of the occluder about 12 months after implantation that potentially resulted from mismatch of occluder size and landing zone. Another patient developed cardiac tamponade 9 days after implantation. This case of delayed effusion was probably not a result of interventional trauma, but might have been provoked by scratching of the inner pericardial membrane. A third patient developed a large thrombus in the left atrium which was considered to be caused by injury of the endothelial wall during implantation. The first two cases could be treated by a percutaneous procedure, the last case by cardiac surgery without any sequelae. CONCLUSIONS: Complications after left atrial appendage closure not related to a device-related thrombus can occur later after implantation. With appropriate percutaneous or surgical management these complications can be handled without sequelae.

Leptin promotes neointima formation and smooth muscle cell proliferation via NADPH oxidase activation and signalling in caveolin-rich microdomains.

AIMS: Apolipoprotein E (apoE) may act as a vasculoprotective factor by promoting plasma lipid clearance and cholesterol efflux. Moreover, apoE accumulates at sites of vascular injury and modulates the effect of growth factors on smooth muscle cells (SMCs). Experimental data suggested that hypothalamic apoE expression is reduced in obesity and associated with leptin resistance. In this study, we examined the role of apoE in mediating the effects of leptin on vascular lesion formation. METHODS AND RESULTS: Leptin was administered to apoE knockout (apoE-/-) mice via osmotic pumps to increase its circulating levels. Morphometric analysis revealed that leptin did not alter neointima formation and failed to increase α-actin- or PCNA-immunopositive SMCs after vascular injury. Similar findings were obtained after analysis of atherosclerotic lesions. Comparison of apoE-/-, wild-type, or LDL receptor-/- mice and functional analyses in aortic SMCs from WT or apoE-/- mice or human arterial SMCs after treatment with small interfering (si)RNA or heparinase revealed that leptin requires the presence of apoE, expressed, secreted and bound to the cell surface, to fully activate leptin receptor signalling and to promote SMC proliferation and neointima formation. Mechanistically, leptin induced the phosphorylation and membrane translocation of caveolin (cav)-1, and apoE down-regulation or caveolae disruption inhibited the leptin-induced p47phox activation, ROS formation and SMC proliferation. Finally, leptin failed to increase neointima formation in mice lacking cav-1. CONCLUSION: Our findings suggest that apoE mediates the effects of leptin on vascular lesion formation by stabilizing cav-1-enriched cell membrane microdomains in SMCs, thus allowing NADPH oxidase assembly and ROS-mediated mitogenic signalling.

Leptin promotes the mobilization of vascular progenitor cells and neovascularization by NOX2-mediated activation of MMP9.

AIMS: Bone marrow (BM) progenitors participate in new vessel formation and endothelial repair. The leptin receptor (ObR) is expressed on hematopoietic cells; however, the effects of leptin on BM progenitor cells and their angiogenic potential are unknown. METHODS AND RESULTS: In the present study, we show that the short-term administration of leptin (over five consecutive days) into wild-type mice increased the number of circulating, BM-derived sca-1(+), flk-1(+) vascular progenitors, 95 ± 1.7% of which also expressed ObR. Ex vivo stimulation of BM cells with leptin enhanced the expression of NADPH oxidase isoform 2 (NOX2), and the leptin-induced increase in reactive oxygen species production, matrix metalloproteinase-9 (MMP9) expression and circulating soluble KitL levels was absent in mice lacking NOX2. Furthermore, intraperitoneal injections of leptin improved perfusion and increased the number of BM-derived, CD31-positive endothelial cells in ischaemic hindlimbs after femoral artery ligation. The effects of leptin on the mobilization of sca-1(+), flk-1(+) cells and neovascularization were abolished in mice transplanted with BM from ObR-deficient and in NOX2(-/-) mice. CONCLUSION: Our findings suggest that the angiogenic effects of leptin involve sca-1(+), flk-1(+) vascular progenitor cells mobilized from the BM in response to ObR-mediated activation of NOX2, increased MMP9 expression, and sKitL release.

Effect of smoking cessation on the number and adhesive properties of early outgrowth endothelial progenitor cells.

BACKGROUND: Endothelial progenitor cells participate in angiogenesis and vascular repair, and cardiovascular risk factors may reduce their numbers or impair their functional properties. Cigarette smoking is a leading cause of preventable cardiovascular death, however, the functional properties of these cells before and after discontinuation of tobacco use have not been systematically analyzed. METHODS: We examined changes in the number and function of early outgrowth endothelial progenitor cells (EPC), isolated from individuals (n=144; mean age, 47.8 ± 12.0 years; 43% males; more than 50% with additional cardiovascular risk factors or disease) who successfully completed a 5-week smoking cessation (SC) programme. RESULTS: SC significantly reduced total white blood cell count (WBC; P<0.0001), plasma LDL cholesterol (P=0.0002) and fibrinogen (P<0.0001) levels, but did not alter the number of circulating CD34(+), VEGFR2(+) or CD34(+), CD133(+) cells (P=0.14 and 0.57, respectively). Fewer acLDL(+), lectin(+) cells could be expanded from peripheral blood mononuclear cells in comparison to baseline (P<0.001). Furthermore, SC was associated with reduced EPC adhesion to fibronectin (P<0.001) or TNFα-activated endothelial cells (P=0.003), and a diminished incorporation of EPC into endothelial cell networks (P=0.035). Mechanistically, significantly reduced ß1- and ß2-integrin expression (P<0.001 and 0.007) and lower contents of intracellular reactive oxygen species (P<0.007) were detected in EPC following SC, in addition to reduced plasma asymmetric dimethyl-L-arginine (ADMA) levels (P=0.0003). CONCLUSIONS: Our findings suggest that the oxidative and inflammatory stress reduction associated with smoking cessation impair the adhesiveness of monocyte-derived EPC.