Induction of inducible nitric oxide synthase (iNOS) expression by oxLDL inhibits macrophage derived foam cell migration.

OBJECTIVE: Deletion of inducible nitric oxide synthase (iNOS) in apolipoprotein E knockout mice was shown to mitigate the extent of arteriosclerosis. Oxidized low density lipoprotein (oxLDL) inhibits macrophage migration and traps foam cells, possibly through a mechanism involving oxidative stress. Here, we addressed whether a reduction of iNOS-mediated oxidative stress remobilizes macrophage-derived foam cells and may reverse plaque formation. METHODS: Migration of RAW264.7 cells and bone marrow cells was quantified using a modified Boyden chamber. iNOS expression, phalloidin staining, focal adhesion kinase phosphorylation, lipid peroxides, nitric oxide (NO) and reactive oxygen species (ROS) production were assessed. RESULTS: oxLDL treatment significantly reduced cell migration compared to unstimulated cells (p < 0.05). This migratory arrest was reversed by co-incubation with a pharmacologic iNOS inhibitor 1400 W (p < 0.05) and iNOS-siRNA (p > 0.05). Furthermore, apoE/iNOS double knockout macrophages do not show migratory arrest in response to oxLDL uptake, compared to apoE knockout controls (p > 0.05). We documented significantly increased iNOS expression following oxLDL treatment and downregulation using 1400 W and small inhibitory RNA (siRNA). iNOS inhibition was associated with a reduction in NO and peroxynitrite (ONOO-)- and increased superoxide generation. Trolox treatment of RAW264.7 cells restored migration indicating that peroxynitrite mediated lipid peroxide formation is involved in the signaling pathway mediating cell arrest.. CONCLUSIONS: Here, we provide pharmacologic and genetic evidence that oxLDL induced iNOS expression inhibits macrophage-derived foam cell migration. Therefore, reduction of peroxynitrite and possibly lipid hydroperoxide levels in plaques represents a valuable therapeutic approach to reverse migratory arrest of macrophage-derived foam cells and to impair plaque formation.

Sonographic and clinical pattern of extracranial and cranial giant cell arteritis.

OBJECTIVES: The aim of our study was to describe the sonographic pattern and clinical manifestations of extracranial (i.e. carotid and proximal arm arteries) and cranial arterial involvement in patients with giant cell arteritis (GCA). METHODS: One hundred and ten consecutive patients with an established diagnosis of GCA between January 2002 and June 2010 were identified retrospectively from a database. All patients underwent colour duplex sonography (CDS) of the superficial temporal, carotid, and proximal arm arteries at the time of diagnosis. Circumferential, homogeneous, hypoechogenic wall thickening was regarded as a typical sign for GCA. Sonographic and clinical characteristics of patients with and without extracranial vessel involvement were compared. RESULTS: Extracranial GCA was observed in 59 of 110 subjects (53.6%). The axillary artery (48.2%) was most frequently affected and bilateral vessel involvement was present in almost all patients (94.8%). Compared to patients with cranial GCA, patients with extracranial GCA were significantly younger, frequently did not meet the American College of Rheumatology (ACR) criteria for classification of cranial GCA, exhibited a lower rate of permanent visual impairment, and were diagnosed later after onset of clinical symptoms (all p < 0.01). With increasing age, a continuous shift from GCA with extracranial arterial involvement to cranial GCA was observed. CONCLUSION: Using CDS, extracranial GCA is a common finding, most frequently observed in the axillary arteries. The clinical pattern of GCA with extracranial arterial involvement differs from that of cranial GCA.

[Recent advances in the treatment of superficial vein thrombosis and extracranial carotid artery stenosis]. / Neues zur Therapie oberflächlicher Venenthrombosen und extrakranieller Karotisstenosen.

Superficial vein thrombosis (SVT) occurs at least as frequent as deep vein thrombosis (DVT), and shares common risk factors with venous thromboembolism. The CALISTO trial was the first to provide specific recommendations for the pharmacologic treatment of SVT. Before treatment is initiated, an accompanying DVT must be excluded and the proximal extension of the SVT assessed. If the proximal extension of the thrombus is closer than 3 cm towards the deep vein system, it should be treated like DVT. Under certain conditions treatment with fondaparinux is indicated in acute symptomatic SVT. Furthermore, compression treatment is recommended. Extracranial carotid artery stenosis can be treated by either surgical thrombarterectomy or catheter based endovascular stent implantation. Trials comparing the two methods have not provided conclusive results on whether the two strategies are equally safe and effective. Considering the latest data from RCTs, careful patient selection (symptoms, comorbidities, age, anatomy, re-stenosis) including individual interdisciplinary discussion appears of ample importance. To date no information is available on whether patients with asymptomatic high grade carotid stenosis receiving "best medical therapy" should be considered for revascularisation in general or only in selected circumstances.

[Cerebrovascular diseases]. / Erkrankungen der hirnversorgenden Gefässe.

In the majority of the cases cerebrovascular disease is caused by atherosclerosis. Duplexsonography is the diagnostic tool of first choice. Management of cardiovascular risk factors is of paramount importance in secondary prevention of atherosclerotic vascular complications. Patients with a symptomatic internal carotid artery stenosis >70% have a clear indication for revascularization. Asymptomatic patients with >60% stenosis benefit from revascularisation if the perioperative risk for death or stroke is below 3%. The optimal revascularization strategy highly depends on the expertise of the local surgeon or endovascular specialist. In younger patients with cerebrovascular disease rare causes such as dissection, large vessel arteritis, fibromuscular dysplasia or vasospasms have to be considered.

NO-synthase-/NO-independent regulation of human and murine platelet soluble guanylyl cyclase activity.

OBJECTIVES: Platelets, specialized adhesive cells, play key roles in normal and pathological hemostasis through their ability to rapidly adhere to subendothelial matrix proteins (adhesion) and to other activated platelets (aggregation), functions which are inhibited by nitric oxide (NO). Platelets have been reported to be regulated not only by exogenous endothelium-derived NO, but also by two isoforms of NO synthase, endothelial (eNOS) and inducible (iNOS), endogenously expressed in platelets. however, data concerning expression, regulation and function of eNOS AND iNOS in platelets remain controversial. METHODS AND RESULTS: Using important positive (endothelial cells, stimulated macrophages) and negative (eNOS/iNOS knock-out mouse) controls, as well as human platelets highly purified by a newly developed protocol, we now demonstrate that human and mouse platelets do not contain eNOS/iNOS proteins or mRNA. NOS substrate (L-arginine), NOS inhibitors (L-NAME, L-NMMA), and eNOS/iNOS deficiency did not produce detectable functional effects on human and mouse platelets. von Willebrand factor (VWF)/ristocetin treatment of platelets increased cGMP by NO-independent activation of soluble guanylyl cyclase (sGC) which correlated with Src kinase-dependent phosphorylation of sGC beta(1)-subunit-Tyr(192). CONCLUSIONS: Human and mouse platelets do not express eNOS/iNOS. VWF/ristocetin-mediated activation of the sGC/cGMP signaling pathway may contribute to feedback platelet inhibition.

Hypertension does not account for the accelerated atherosclerosis and development of aneurysms in male apolipoprotein e/endothelial nitric oxide synthase double knockout mice.

BACKGROUND: Apolipoprotein E (apoE)/endothelial nitric oxide synthase (eNOS) double knockout (DKO) mice demonstrate accelerated atherosclerosis and develop abdominal aortic aneurysms and aortic dissection, suggesting a role for eNOS in suppressing atherogenesis. To test whether accelerated atherosclerosis and aortic aneurysms were due to hypertension, we administered hydralazine to male apoE/eNOS DKO mice to reduce blood pressure. METHODS AND RESULTS: Male apoE/eNOS DKO mice were treated with hydralazine in their drinking water (250 mg/L) using a dose that lowers the blood pressure to levels seen in apoE KO mice. The mice were fed a Western-type diet for 16 weeks, and lesion formation was assessed by inspection of the vessel and staining with Sudan IV. Hydralazine-treated, normotensive male apoE/eNOS DKO mice developed increased aortic lesion areas (30.0+/-2.8%, n=11) compared with male apoE KO mice (14.6+/-0.8%, n=7). The extent of lesion formation was not significantly different from male apoE/eNOS DKO mice that were not given hydralazine (28.3+/-3.1%, n=9). Four of 11 hydralazine-treated male apoE/eNOS DKO mice developed abdominal aortic aneurysms. CONCLUSIONS: Hypertension is not required for the accelerated atherosclerosis seen in apoE/eNOS DKO animals, and control of hypertension during a 16-week period does not prevent aortic aneurysm formation.

Accelerated atherosclerosis, aortic aneurysm formation, and ischemic heart disease in apolipoprotein E/endothelial nitric oxide synthase double-knockout mice.

BACKGROUND: To test whether deficiency in endothelial nitric oxide synthase (eNOS) affects atherosclerosis development, we compared lesion formation in apolipoprotein E (apoE)/eNOS-double knockout (DKO) and apoE-knockout (KO) control animals. METHODS AND RESULTS: After 16 weeks of "Western-type" diet, apoE/eNOS-DKO males and females showed significant increases in lesion area of 93.6% and 59.2% compared with apoE-KO mice. All apoE/eNOS-DKO animals studied developed peripheral coronary arteriosclerosis, associated with perivascular and myocardial fibrosis, whereas none of the apoE-KO mice did. Transthoracic echocardiography showed a significantly increased left ventricular wall thickness and decreased fractional shortening in DKO animals. Mean arterial pressure was increased in DKO mice and was comparable in degree to eNOS-KO animals. Male DKO animals developed atherosclerotic abdominal aneurysms and aortic dissection. CONCLUSIONS: eNOS deficiency increases atherosclerosis in Western-type diet-fed apoE-KO animals and introduces coronary disease and an array of cardiovascular complications, including spontaneous aortic aneurysm and dissection. This phenotype constitutes the first murine model to demonstrate distal coronary arteriosclerosis associated with evidence of myocardial ischemia, infarction, and heart failure. Hypertrophy and reduced left ventricular function cannot be explained by increased blood pressure alone, because eNOS-KO animals do not develop these complications.

Genetic deficiency of inducible nitric oxide synthase reduces atherosclerosis and lowers plasma lipid peroxides in apolipoprotein E-knockout mice.

BACKGROUND: Inducible nitric oxide synthase (iNOS) is expressed by leukocytes and smooth muscle cells in atherosclerotic lesions. To test whether NO produced by iNOS deficiency affects atherosclerosis, we studied apoE/iNOS-double knockout (dKO) and apoE-knockout (KO) control animals fed a "Western-type" diet. METHODS AND RESULTS: After 16 weeks of Western-type diet, the aortic lesion area in apoE/iNOS-dKO males and females was significantly reduced, by 22% and 21%, respectively, compared with apoE-KO males and females. This effect was more pronounced after 24 weeks of Western-type diet, after which lesion formation in male and female dKO mice was reduced by 38% and 40%, respectively. Plasma levels of lipoperoxides in apoE/iNOS-dKO mice (2.0+/-0.23 micromol/L) were significantly lower than in apoE-KO control animals (3.2+/-0.44 micromol/L; P=0.02). To test whether substrate deficiency plays a role in the proatherogenic actions of iNOS, we administered L-arginine to apoE-KO animals for 16 and 24 weeks. L-Arginine treatment did not affect lesion formation in apoE-KO animals fed a Western-type diet. CONCLUSIONS: Genetic deficiency of iNOS decreases diet-induced atherosclerosis and lowers plasma levels of lipoperoxides, a marker for oxidative stress, in apoE-KO animals. Reduction in iNOS-mediated oxidative stress could partly explain protection from lesion formation in dKO animals. L-Arginine supplementation did not change lesion area in apoE-KO mice, indicating that substrate deficiency is not a likely cause for iNOS-mediated injury in this model of atherosclerosis.