Neutralization of Interleukin-18 Inhibits Neointimal Formation in a Rat Model of Vascular Injury

Department of Experimental Pharmacology, University of Naples Federico II, Via D. Montesano 49, 80131 Naples, Italy.
Circulation (Impact Factor: 14.43). 09/2006; 114(5):430-7. DOI: 10.1161/CIRCULATIONAHA.105.602714
Source: PubMed


Studies in humans and animal models suggest that interleukin-18 (IL-18) plays a crucial role in vascular pathologies. IL-18 is a predictor of cardiovascular death in angina and is involved in atherotic plaque destabilization. Higher IL-18 plasma levels also are associated with restenosis after coronary artery angioplasty performed in patients with acute myocardial infarction. We investigated the effective role of IL-18 in neointimal formation in a balloon-induced rat model of vascular injury.
Endothelial denudation of the left carotid artery was performed by use of a balloon embolectomy catheter. Increased expression of IL-18 and IL-18Ralpha/beta mRNA was detectable in carotid arteries from days 2 to 14 after angioplasty. The active form of IL-18 was highly expressed in injured arteries. Strong immunoreactivity for IL-18 was detected in the medial smooth muscle cells at days 2 and 7 after balloon injury and in proliferating/migrating smooth muscle cells in neointima at day 14. Moreover, serum concentrations of IL-18 were significantly higher among rats subjected to vascular injury. Treatment with neutralizing rabbit anti-rat IL-18 immunoglobulin G significantly reduced neointimal formation (by 27%; P < 0.01), reduced the number of proliferating cells, and inhibited interferon-gamma, IL-6, and IL-8 mRNA expression and nuclear factor-kappaB activation in injured arteries. In addition, in vitro data show that IL-18 affects smooth muscle cell proliferation.
These results identify a critical role for IL-18 in neointimal formation in a rat model of vascular injury and suggest a potential role for IL-18 neutralization in the reduction of neointimal development.

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    • "Total extracts were prepared from liquid nitrogen frozen pooled carotid arteries (n = 2), crushed into powder, in a mortar with a pestle,and resuspended in 150 µl of lysis buffer (20 mM HEPES, 0.4 mM NaCl, 1.5 mM MgCl2, 1 mM EGTA, 1 mM EDTA, 1% Triton X-100, and 20% glycerol) containing protease inhibitors (1 mM DTT, 0.5 mM PMSF, 15 mg/mL Try-inhibitor, 3 mg/mL pepstatin-A, 2 mg/mL leupeptin, and 40 mM benzamidine) [20], [21]. After centrifugation at 13000×g at 4°C for 30 min, supernatants were collected and stored at −80°C until the assays. "
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    ABSTRACT: Bindarit, a selective inhibitor of monocyte chemotactic proteins (MCPs) synthesis, reduces neointimal formation in animal models of vascular injury and recently has been shown to inhibit in-stent late loss in a placebo-controlled phase II clinical trial. However, the mechanisms underlying the efficacy of bindarit in controlling neointimal formation/restenosis have not been fully elucidated. Therefore, we investigated the effect of bindarit on human coronary smooth muscle cells activation, drawing attention to the phenotypic modulation process, focusing on contractile proteins expression as well as proliferation and migration. The expression of contractile proteins was evaluated by western blot analysis on cultured human coronary smooth muscle cells stimulated with TNF-α (30 ng/mL) or fetal bovine serum (5%). Bindarit (100-300 µM) reduced the embryonic form of smooth muscle myosin heavy chain while increased smooth muscle α-actin and calponin in both TNF-α- and fetal bovine serum-stimulated cells. These effects were associated with the inhibition of human coronary smooth muscle cell proliferation/migration and both MCP-1 and MCP-3 production. The effect of bindarit on smooth muscle cells phenotypic switching was confirmed in vivo in the rat balloon angioplasty model. Bindarit (200 mg/Kg/day) significantly reduced the expression of the embryonic form of smooth muscle myosin heavy chain, and increased smooth muscle α-actin and calponin in the rat carodid arteries subjected to endothelial denudation. Our results demonstrate that bindarit induces the differentiated state of human coronary smooth muscle cells, suggesting a novel underlying mechanisms by which this drug inhibits neointimal formation.
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    • "Interestingly, Yamagami et al. (2005) demonstrated an association between higher serum IL-18 levels and greater intima-media thickness. Maffia et al. (2006) also reported that IL-18 play a critical role in neointimal formation. With respect to previous studies, our findings indicate that IL-18-mediated upregulation of dystrophin expression may play either a direct or indirect role in maintenance of BBB function following SE. "
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    • "Rats were anaesthetized with an intraperitoneal injection of ketamine (100 mg/kg) (Gellini International) and xylazine (5 mg/kg) (Sigma). Endothelial denudation of the left carotid artery was performed with a balloon embolectomy catheter (2 F, Fogarty, Edwards Lifesciences) according to the procedure well validated in our laboratories.18 Some animals were subjected to anaesthesia and surgical procedure, without balloon injury (sham-operated group). "
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