Radiofrequency endothelial ablation prevents recanalization after endovascular coil occlusion: in vitro and in vivo assessment.

PURPOSE: Coil embolization of intracranial aneurysms may be followed by recurrences. Radiofrequency (RF) ablation of the endothelium may prevent recanalization after coil embolization. MATERIALS AND METHODS: The authors performed in vitro experiments in chicken meat and egg white models to investigate the thermal distribution and geometry of lesions created with RF applied through standard coils alone or by using a prototype RF electrode inserted in a coil or a mass of coils. A mathematic model was designed to predict perianeurysmal isotherm lesions by using the bio-heat equation. In an in vivo coil arterial occlusion model (six dogs), the authors compared angiographic and pathologic results of coil embolization (n = 8) with those of coil embolization preceded by RF ablation (n = 7) by using a cardiac electrode at 1 month. RESULTS: Current coils offer high impedance (400 Omega) at high current frequencies and are damaged by RF transmission. A dedicated electrode generated reproducible lesions, but contact with coils interferes with lesion reproducibility. When the coil mass was used, a uniform RF lesion that conformed to the coil mass shape was produced. The mathematic model predicted a uniform heat distribution within 1 mm from the coil mass periphery. Arterial coil embolization led to occlusion followed by recanalization (n = 8), whereas RF ablation (20-30 W for 60 seconds) prevented recanalization in all coil-occluded arteries (P < .001, chi(2) test). Pathologic findings helped confirm complete arterial occlusion with RF ablation. One animal developed brachial plexus injury with excessive levels of RF ablation. CONCLUSIONS: RF ablation can prevent recanalization after coil occlusion-at least in the arterial model. Modifications of coils, dedicated neurovascular electrodes, and technique optimization remain necessary before considering a clinical application.

Deep vein thrombosis resolution is impaired in diet-induced type 2 diabetic mice.

OBJECTIVE: Type 2 diabetes mellitus results in a procoagulant and thrombogenic state that could predispose diabetic individuals to develop venous thrombosis. We sought to determine whether diet-induced type 2 diabetes mellitus affects deep venous thrombosis (DVT) resolution in a murine model. METHODS: C57Bl/6 mice were fed a low-fat or a high-fat diet (n = 10) for 10 weeks, after which DVT was created in the inferior vena cava (IVC) by a combination of low flow and endothelial damage. The IVC and thrombus were harvested at 1 and 2 weeks. Thrombus resolution and neovascularization were investigated through transfemoral angiography (n = 10), thrombus size (n = 4) and weight (n = 10), and nitric oxide synthase 3 immunoquantification (n = 4). Macrophage content was assessed by CD68 immunoreactivity (n = 4). The fibrinolytic system (urokinase plasminogen activator [uPA] and plasminogen activator inhibitor-1 [PAI-1]) was analyzed by Western immunoblotting (n = 6) and immunohistochemistry (n = 4). Total collagen was stained by Sirius red. Matrix metalloproteinases (MMP)-2 and MMP-9 activities were evaluated by zymography and their expressions by Western immunoblotting (n = 6) and immunohistochemistry (n = 4). RESULTS: Diabetic mice had significantly larger and heavier thrombi at 1 and 2 weeks (P < .05), threefold less neovascularization (P < .05), and 35-fold increase in macrophage content (P < .01), than control mice 2 weeks after surgery. IVC recanalization was documented in 90% of 2-week control mice and in 10% of 2-week diabetic mice (P < .01). Increased vein wall collagen and less uPA and more PAI-1 expressions with a decreased uPA/PAI-1 ratio (31%, P < .01) were documented at 2 weeks in diabetic mice. MMP-2 and MMP-9 activities and expressions were significantly increased in diabetic mice at 1 and 2 weeks (P < .05) compared with control mice. CONCLUSION: Diet-induced type 2 diabetes may impair DVT resolution through altered inflammatory, fibrinolytic, and MMP responses.

Matrix metalloproteinase-9 may play a role in recanalization and recurrence after therapeutic embolization of aneurysms or arteries.

PURPOSE: Matrix metalloproteinase (MMP)-9 plays various roles in vascular healing and angiogenesis. This study was conducted to determine if MMP-9 is involved in healing or recanalization after therapeutic occlusion of arteries or aneurysms. MATERIALS AND METHODS: Angiographic and pathologic changes were investigated in canine bilateral venous pouch carotid aneurysms embolized with gelatin sponges with or without previous endothelial denudation, a procedure that can prevent recanalization. To assess a potential role of MMP-9, messenger RNA (mRNA) and protein were compared in denuded and nondenuded aneurysms 4, 7, and 14 days after embolization. To assess if MMP-9 is essential to arterial recanalization, transmyocardial angiography and pathologic findings were compared 14 days after carotid occlusion with platinum coils in MMP-9-knockout and wild-type mice. RESULTS: Denudation of the endothelial lining led to improved angiographic results at 3 weeks (P < .001). Neointimal closure of the aneurysm neck was more complete in denuded versus nondenuded aneurysms. Denudation was followed by a decrease in MMP-9 mRNA (86%, P < .05) and protein (30%, P < .05) 7 days after embolization and a decrease in von Willebrand factor compared with nondenuded aneurysms. MMP-9 immunostaining of axial sections from embolized aneurysms confirmed MMP-9-positive endothelialized clefts, which were absent in denuded aneurysms. Transmyocardial angiography and pathologic examination showed recanalization of one of nine coiled carotid arteries of MMP-9-knockout mice, compared with five of seven controls (P = .035). CONCLUSIONS: MMP-9 may play a role in recanalization of arteries after coil occlusion and in recurrences after sponge embolization of aneurysms.

The effects of stenting and endothelial denudation on aneurysm and branch occlusion in experimental aneurysm models.

BACKGROUND: Stents are increasingly used in the endovascular treatment of intracranial aneurysms. We studied the effects of stenting and endothelial denudation on aneurysm and branch vessel occlusion. METHODS: Bilateral lingual bifurcation venous pouch aneurysms were created in eight dogs, surgically scraping the aneurysmal endothelial lining on one side. Both arteries were immediately stented using balloon-expandable stents. In four other dogs, a wide-neck carotid bifurcation aneurysm was created, with the vein pouch denuded or not (n=2 each), followed by immediate stenting. Results were compared using angiography and pathology at 10 days (n=2), 10 (n=8), and 20 weeks (n=2). Branch occlusion between initial and final angiograms was recorded. Pathological evaluation of aneurysms was studied, with attention to neointima formation at the aneurysm ostium and around branch vessel origins. RESULTS: All stented and denuded lingual aneurysms were obliterated compared with two of eight lingual aneurysms that were stented alone (P=.007). None of the carotid bifurcation aneurysms became obliterated (0/4), but denuded aneurysms showed partial thrombosis (2/2). Of 68 total stent-covered branches, 5 (7%) were occluded and 17 (27%) had altered angiographic flow. CONCLUSIONS: Stenting led to suboptimal results in the presence of an intact endothelial layer. Endothelial denudation can promote aneurysm occlusion when combined with stenting.

Thy-1 expression by cardiac fibroblasts: lack of association with myofibroblast contractile markers.

The objective of this study was to investigate the presence of Thy-1 in the myocardium and on cardiac fibroblasts and to determine whether or not cardiac fibroblasts form a heterogeneous population in term of Thy-1 expression. Thy-1 expression was examined by immunohistology of ventricular sections from normal and deoxycorticosterone acetate (DOCA)-salt hypertensive rats. Thy-1 immunostaining was detected in connective tissue on alpha8 integrin-positive and discoidin domain receptor 2 (DDR2)-positive fibroblasts. Enhanced Thy-1 staining was observed in the hearts of DOCA-salt rats particularly in areas of interstitial fibrosis. Cardiac mRNA analysis confirmed the increased Thy-1 expression. On cultured cardiac fibroblasts, flow cytofluorometry showed that cells, from primary culture to passage 4, were double positive for Thy-1 and for both alpha8 integrin and DDR2. Analysis of isolated lipid rafts by detergent-free sucrose gradient indicated that Thy-1 protein was probably located in these structures, but it may be located on a membrane microdomain slightly different from those of caveolin-1, as revealed by immunocytochemistry. Differentiation of fibroblasts into myofibroblasts being a characteristic of cardiac fibrosis and scarring, cardiac fibroblasts were stimulated in the presence of transforming growth factor-beta (TGF-beta) or connective tissue growth factor. While the expression of smooth muscle alpha-actin and alpha8 integrin doubled, Thy-1 level, measured by Western blotting and flow cytofluorometry, was not influenced by TGF-beta. These results demonstrate that cardiac fibroblasts express Thy-1 and form a homogeneous population. Thy-1 expression also appears to be independent of fibroblast differentiation. The dichotomy between the increased Tthy-1 expression in the fibrotic area and the lack of association with fibroblast differentiation suggests that Thy-1 may represent a marker of fibroblast proliferation in the myocardium.

Inhibition of angiogenic properties of brain endothelial cells by platelet-derived sphingosine-1-phosphate.

The platelet-derived lysophospholipid sphingosine-1-phosphate (S1P) is present in blood plasma and is one of the most potent growth factors displaying proangiogenic activity towards endothelial cells (EC) derived from various tissues. The paracrine regulation of brain angiogenesis by platelet-derived growth factors is, however, poorly understood. In the present study, we assessed the role of S1P on brain EC migration and tubulogenesis, using rat brain-derived (RBE4) EC as an in vitro model. We show that S1P inhibits brain EC migration and tubulogenesis, while it displays proangiogenic activity towards noncerebral EC. Overexpression of the S1P receptor S1P-1 in RBE4 cells potentiated all of the S1P-mediated events. We also show that the lack of expression of MT1-MMP, a membrane-bound matrix metalloproteinase that is thought to cooperate with S1P in tubulogenic processes, may explain the antiangiogenic activity of S1P on brain vasculature. Altogether our results support the hypothesis of a tissue-specific, antiangiogenic role of S1P in the brain, which may help to stabilize the cerebral vasculature and thus have crucial impact on the setting and regulation of normal brain vascularization.

Enhanced expression of fibrillin-1, a constituent of the myocardial extracellular matrix in fibrosis.

Fibrillin-1 localization in the myocardium and the modulation of its expression in cardiac fibrosis were examined. In normal rat hearts, fibrillin-1 was abundant throughout the myocardium as thin fibers that crossed over the perimysium and around arteries. After cardiac fibrosis was induced in rats by either 14-day ANG II infusion or 21-day DOCA-salt treatment [a high endothelin-1 (ET-1) model], fibrillin-1 immunostaining was stronger in the interstitium (2.8-fold and 4.4-fold increases, respectively, in each model), extended between myocytes, and accumulated in microscopic scars and in the perivascular area of both ventricles. mRNA analysis confirmed its enhanced ventricular expression in both groups of rats (2.5-fold and 6.6-fold increments, respectively, in each model). In 1B normotensive and 2C hypertensive transgenic mice, two lines expressing an ANG II fusion protein in cardiac myocytes, strong fibrillin-1 immunoreactivity was observed in the interstitium and around arteries (3.7-fold and 7-fold increases, respectively). ANG II and transforming growth factor-beta1 enhanced fibrillin-1 synthesis by cardiac fibroblasts. Some fibrillin-1 fragments interacted with RGD-dependent integrins, including alpha(8)beta(1)-integrin, of cardiac fibroblasts but not necessarily through the RGD motif. Our findings illustrate that fibrillin-1 is an important constituent of the myocardium. In vitro and in vivo evidence suggests that ANG II can directly induce fibrillin-1 expression in cardiac fibroblasts. This protein can thus contribute to reactive and reparative processes.