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Aims The adenylate cyclase type 9 (ADCY9) gene appears to determine atherosclerotic outcomes in patients treated with dalcetrapib. In mice, we recently demonstrated that Adcy9 inactivation potentiates endothelial function and inhibits atherogenesis. The objective of this study was to characterize the contribution of ADCY9 to the regulation of endothelial signalling pathways involved in atherosclerosis. Methods and results We show that ADCY9 is expressed in the endothelium of mouse aorta and femoral arteries. We demonstrate that ADCY9 inactivation in cultured endothelial cells paradoxically increases cAMP accumulation in response to the adenylate cyclase activators forskolin and vasoactive intestinal peptide (VIP). Reciprocally, ADCY9 overexpression decreases cAMP production. Using mouse femoral artery arteriography, we show that Adcy9 inactivation potentiates VIP-induced endothelial-dependent vasodilation. Moreover, Adcy9 inactivation reduces mouse atheroma endothelial permeability in different vascular beds. ADCY9 overexpression reduces forskolin-induced phosphorylation of Ser157-vasodilator-stimulated phosphoprotein (VASP) and worsens thrombin-induced fall of RAP1 activity, both leading to increased endothelial permeability. ADCY9 inactivation in thrombin-stimulated human coronary artery endothelial cells results in cAMP accumulation, increases p-Ser157-VASP, and inhibits endothelial permeability. MLC2 phosphorylation and actin stress fibre increases in response to thrombin were reduced by ADCY9 inactivation, suggesting actin cytoskeleton regulation. Finally, using the Miles assay, we demonstrate that Adcy9 regulates thrombin-induced endothelial permeability in vivo in normal and atherosclerotic animals. Conclusion Adcy9 is expressed in endothelial cells and regulates local cAMP and endothelial functions including permeability relevant to atherogenesis.

Background and aims: The anti-inflammatory agent colchicine is gaining interest as a treatment for coronary artery disease. However, the effects of colchicine in atherosclerotic animal models are mostly unknown. This study aimed to evaluate colchicine in a rabbit model of atherosclerosis. Methods: Twenty-two rabbits were fed a 0.5% cholesterol-enriched diet for 10 weeks and then randomized to receive either oral saline (n=11) or colchicine (350 μg/kg/day; n=11) for 6 weeks, with 0.2% cholesterol-diet during the treatment period. We performed intravascular ultrasound imaging (at start and end of treatment) and histology analyses of the descending thoracic aorta. Leucocyte activation was assessed in vitro on blood samples obtained during treatment. Results: Colchicine prevented positive aortic vascular remodelling (p=0.029 vs placebo). This effect was even more marked at high plasma cholesterol level (third quartile of plasma cholesterol, p=0.020). At high cholesterol level, both atherosclerotic plaque and media areas on histomorphology were reduced by colchicine compared to placebo (p=0.031 and p=0.039, respectively). Plaque fibrosis and macrophage area were reduced by colchicine (Masson's trichrome stain: p=0.038; RAM-11: p=0.026). The plaque vulnerability index, assessed by histology, was reduced by colchicine (p=0.040). Elastin/type I collagen ratio in media was significantly higher with colchicine compared to placebo (p=0.013). At a high level of plasma cholesterol, in vitro LPS challenge revealed a decrease in monocyte activation following treatment with colchicine (p

The large HDL particles generated by administration of cholesteryl ester transfer protein inhibitors (CETPi) remain poorly characterized, despite their potential importance in the routing of cholesterol to the liver for excretion, which is the last step of the reverse cholesterol transport. Thus, the effects of the CETPi dalcetrapib and anacetrapib on HDL particle composition were studied in rabbits and humans. The association of rabbit HDL to the LDL receptor (LDLr) in vitro was also evaluated. New Zealand White rabbits receiving atorvastatin were treated with dalcetrapib or anacetrapib. A subset of patients from the dal-PLAQUE-2 study treated with dalcetrapib or placebo were also studied. In rabbits, dalcetrapib and anacetrapib increased HDL-C by more than 58% (P0.01) and in turn raised large apo E-containing HDL by 66% (P0.001) and 59% (P0.01), respectively. Additionally, HDL from CETPi-treated rabbits competed with human LDL for binding to the LDLr on HepG2 cells more than control HDL (P0.01). In humans, dalcetrapib increased concentrations of large HDL particles (+69%, P0.001) and apo B-depleted plasma apo E (+24%, P0.001), leading to the formation of apo E-containing HDL (+47%, P0.001) devoid of apo A-I. Overall, in rabbits and humans, CETPi increased large apo E-containing HDL particle concentration, which can interact with hepatic LDLr. The catabolism of these particles may depend on an adequate level of LDLr to contribute to reverse cholesterol transport.

Background We previously demonstrated that high-density lipoprotein (HDL) infusions may improve left ventricular diastolic dysfunction (LVDD) in an aortic valve stenosis (AVS) model. Whether the benefit was direct or mediated by the observed reduction in AVS severity is not clear. Here, we aimed to test the direct effect of an ApoA-I mimetic on LVDD in the absence of AVS. Methods Rabbits were exposed to three different protocols to develop LVDD. First, rabbits were exposed to 0.5% cholesterol-rich diet for an average of 17 weeks. Second, rabbits were subjected to surgical ascending aortic constriction (AAC), to mimic the effect of fixed reduced aortic valve area, and studied after 10 weeks. The third model combined both cholesterol-enriched diet (for 12 weeks) and surgical AAC. The control group consisted of age-matched rabbits under normal diet. After development of LVDD, rabbits were randomized to receive infusions of saline or apoA-I mimetic (25 mg/kg) 3 times per week for 4 weeks. Detailed cardiac structure and function measurements were assessed at baseline and weekly during treatment period. Histological and molecular analyses were performed on LV samples. Results In the three models, echocardiographic results showed development of LVDD over time, with preserved LV systolic and aortic valve functions versus controls. ApoA-I mimetic infusions did not significantly improve echocardiographic parameters nor molecular markers of cardiac inflammation, oxidative stress and fibrosis. Conclusion ApoA-I mimetic therapy did not directly improve LVDD. These results indicate that previously observed changes of LVDD were caused by AVS improvement induced by this treatment.

Background: Pharmacogenomic studies have shown that ADCY9 genotype determines the effects of the CETP (cholesteryl ester transfer protein) inhibitor dalcetrapib on cardiovascular events and atherosclerosis imaging. The underlying mechanisms responsible for the interactions between ADCY9 and CETP activity have not yet been determined. Methods: Adcy9 -inactivated ( Adcy9 Gt/Gt ) and wild-type (WT) mice, that were or not transgenic for the CETP gene (CETPtg Adcy9 Gt/Gt and CETPtg Adcy9 WT ), were submitted to an atherogenic protocol (injection of an AAV8 [adeno-associated virus serotype 8] expressing a PCSK9 [proprotein convertase subtilisin/kexin type 9] gain-of-function variant and 0.75% cholesterol diet for 16 weeks). Atherosclerosis, vasorelaxation, telemetry, and adipose tissue magnetic resonance imaging were evaluated. Results: Adcy9 Gt/Gt mice had a 65% reduction in aortic atherosclerosis compared to WT ( P Adcy9 Gt/Gt mice compared to WT animals ( P Adcy9 Gt/Gt mice (versus WT, P Adcy9 Gt/Gt ( P Adcy9 Gt/Gt mice allowed significantly less adhesion of splenocytes compared to WT ( P Adcy9 Gt/Gt mice gained more weight than WT with the atherogenic diet; this was associated with an increase in whole body adipose tissue volume ( P Adcy9 Gt/Gt compared to WT mice ( P P P =0.0572). Adcy9 inactivation–induced effects on atherosclerosis, endothelial function, weight gain, adipose tissue volume, and feed efficiency were lost in CETPtg Adcy9 Gt/Gt mice ( P >0.05 versus CETPtg Adcy9 WT ). Conclusions: Adcy9 inactivation protects against atherosclerosis, but only in the absence of CETP activity. This atheroprotection may be explained by decreased macrophage accumulation and proliferation in the arterial wall, and improved endothelial function and autonomic tone.

Garden potato varieties grown on more than 1.5 ha are subject to variety registration to be sold as seed potato in Canada. Prerequisites for the registration of varieties include the pedigree and breeding history as well as a morphological description. For most varieties, no pedigree or morphological description is available, and furthermore, some groups are essentially derived from one another. Therefore, a review to determine the genetic and morphological similarities and uniqueness of some of the garden potato varieties was initiated. Seven garden potato varieties grown over the largest areas in Canada were genotyped using a set of nine microsatellite [simple sequence repeat (SSR)] markers, resulting in six distinct SSR profiles. These varieties were also grown out for morphological description, providing supplementary information for the national registration and their monitoring during the seed certification process. To further assess the genetic similarities of garden or heritage varieties, 84 potato varieties were genotyped, and the unique SSR profiles of 62 varieties and six SSR profiles shared among the other 22 varieties were added to the Canadian Food Inspection Agency potato variety reference SSR profile collection. Furthermore, a set of blue varieties were described morphologically to evaluate the nature of morphological characteristic variation of varieties sharing the same SSR profile. Although all 14 varieties grown show some morphological differences, all varieties within each group sharing an SSR profile also clustered together based on morphological characteristics.

BackgroundLeft ventricular diastolic dysfunction (LVDD) is present in more than 50% of patients suffering from heart failure. LVDD animal models are limited and its underlying mechanisms remain largely unknown. Aortic valve stenosis (AVS) may cause LVDD, and we recently reported LVDD in an AVS rabbit model. Here we aimed to develop a rabbit model of LVDD without AVS.MethodsRabbits were fed with a 0.5% cholesterol-enriched diet (n = 9) or normal diet (n = 8) until they developed LVDD defined by a value of the echocardiographic parameter E/Em ratio higher than the mean at baseline + 2SD. Rabbits were then fed a 0.2% cholesterol-enriched diet for 4 weeks (average total diet duration: 20 weeks). Detailed cardiac structure and function measurements were assessed by echocardiography at baseline, weeks 8, 12 and 14 to 20, when applicable. Histological analyses and RT-qPCR were performed on LV samples.ResultsThe hypercholesterolemic diet induced LVDD without systolic dysfunction or AVS, as shown by multiple echocardiographic parameters, including early filling mitral peak velocity and deceleration rate, Em/Am ratio and E/Em ratio (all pConclusionRabbits fed with a cholesterol-enriched diet develop LVDD with preserved systolic function and evidence of cardiac inflammation and oxidative stress. This rabbit model may be used in future studies to test treatment strategies against LVDD.

Aortic valve stenosis (AVS) is the most common valvular heart disease in the Western world. Therapy based on apolipoprotein A-I (apoA-I), the major protein component of high-density lipoproteins, results in AVS regression in experimental models. Nevertheless, apoA-I degradation by proteases might lead to suboptimal efficacy of such therapy. An activatable probe using a quenched fluorescently labeled full-length apoA-I protein was generated to assess apoA-I-degrading protease activity in plasma derived from 44 men and 20 women with severe AVS (age 65.0 ± 10.4 years) as well as from a rabbit model of AVS. In human and rabbit AVS plasma, apoA-I-degrading protease activity was significantly higher than in controls (humans: 0.038 ± 0.009 vs 0.022 ± 0.005 RFU/s, p

Pharmacogenomic studies have shown that ADCY9 genotype determines the effects of the cholesteryl ester transfer protein (CETP) inhibitor dalcetrapib on cardiovascular events, atherosclerosis imaging and body weight variation. The underlying mechanisms responsible for the interactions between ADCY9 and CETP have not yet been determined. Adcy9 -inactivated ( Adcy9 Gt/Gt ) and wild-type (WT) mice, that were or not transgenic for the CETP gene (CETP Gt and CETP WT ), were submitted to an atherogenic protocol (injection of an AAV8 expressing a PCSK9 gain-of-function variant and 0.75% cholesterol diet for 16 weeks). Atherosclerosis, cell adhesion, vasorelaxation, telemetry and adipose tissue MRI were evaluated. Adcy9 Gt/Gt mice had a 65% reduction in aortic atherosclerosis compared to WT ( P Adcy9 Gt/Gt mice compared to WT animals ( P Adcy9 inactivation did not change counts of blood monocytes and their subsets. Splenocytes showed reduced adhesion to native aortic endothelium from Adcy9 Gt/Gt mice ( P Adcy9 Gt/Gt mice (versus WT, P Adcy9 Gt/Gt versus WT ( P Adcy9 Gt/Gt mice gained more weight than WT with the atherogenic diet, and this was associated with an increase in whole body adipose tissue volume ( P Adcy9 Gt/Gt compared to WT mice ( P P =0.0572) and prolonged cardiac RR interval ( P Adcy9 inactivation-induced effects on atherosclerosis, endothelium-dependent vasodilation, weight gain and feed efficiency were lost in CETP Gt mice ( P >0.05 vs CETP WT ). Adcy9 inactivation protects against atherosclerosis, but only in the absence of CETP activity. This atheroprotection may be explained by decreased macrophage accumulation and proliferation in the arterial wall, improved endothelial function and autonomic tone.

Background Infusions of apolipoprotein A-I (apoA-I), the major protein component of high-density lipoproteins (HDL), result in aortic valve stenosis (AVS) regression in experimental models. Severe AVS can be complicated by acquired von Willebrand syndrome, a haemorrhagic disorder associated with loss of high-molecular-weight von Willebrand factor (vWF) multimers (HMWM), the latter being a consequence of increased shear stress and enhanced vWF-cleaving protease (ADAMTS-13) activity. Although antithrombotic actions of HDL have been described, its effects on ADAMTS-13 and vWF in AVS are unknown. Methods and Results We assessed ADAMTS-13 activity in plasma derived from a rabbit model of AVS (n = 29) as well as in plasma collected from 64 patients with severe AVS (age 65.0 ± 10.4 years, 44 males) undergoing aortic valve replacement (AVR). In both human and rabbit AVS plasma, ADAMTS-13 activity was higher than that in controls (p Conclusion Our data indicate that HDL levels are associated with reduced ADAMTS-13 activity and increased HMWM. HDL-based therapies may reduce the haematologic abnormalities of the acquired von Willebrand syndrome in AVS.

Polymorphisms in the ADCY9 gene, coding for adenylate cyclase type 9, determine atherosclerotic cardiovascular responses to the CETP inhibitor dalcetrapib in patients. ADCY9 is broadly expressed and involved in various immune cell functions and inflammatory responses. Our objective is to determine the role of ADCY9 in the development of atherosclerosis in the absence of CETP. We used 8-12 week-old wild-type (WT, n=25) or Adcy9 -inactivated ( Adcy9 Gt , n=21) male mice. To induce hypercholesterolemia and atherosclerosis, mice were infected with an adeno-associated virus coding for a gain-of-function mutant of Pcsk9 ( Pcsk9 D377Y ) and were fed an atherogenic high-cholesterol diet for 16 weeks in absence or presence of dalcetrapib treatment (200 mg/kg/day). Percent atherosclerotic lesion area (PALA) in the whole aorta was quantified using en face Oil Red O plaque staining. We used VCAM-1 immunofluorescence staining (n=6 per group) in atherosclerotic lesions of the aortic root to quantify inflammation. We also measured aortic root accumulation of macrophages and vascular smooth muscle cells by immunofluorescence imaging of CD68 and smooth muscle actin (SMA), respectively, in plaque lesions. We also quantified blood leukocytes in normocholesterolemic WT and Adcy9 Gt mice by flow cytometry analysis of CD45 + cells. In hypercholesterolemic animals, Adcy9 Gt mice showed a 60% decrease in atherosclerotic lesions (PALA: 2.7±0.5%) compared to WT mice (6.5±0.7%, PAdcy9 Gt mice (2.9±0.4%) compared to WT (7.6±1.6%, PAdcy9 Gt mice (PAdcy9 Gt mice (P=0.0598). CD45 + blood leukocytes were reduced by 35.4% in Adcy9 Gt compared to WT normocholesterolemic mice (P Adcy9 inactivation in mice (without CETP) results in large reductions of atherosclerosis and plaque macrophage and in decreased VCAM-1 expression. These results support the ADCY9 genotype-dependent clinical effects of dalcetrapib.

Atherosclerotic cardiovascular diseases are characterized by the formation of a plaque in the arterial wall. Intravascular ultrasound (IVUS) provides high-resolution images allowing delineation of atherosclerotic plaques. When combined with near infrared fluorescence (NIRF), the plaque can also be studied at a molecular level with a large variety of biomarkers. In this work, we present a system enabling automated volumetric histology imaging of excised aortas that can spatially correlate results with combined IVUS/NIRF imaging of lipid-rich atheroma in cholesterol-fed rabbits. Pullbacks in the rabbit aortas were performed with a dual modality IVUS/NIRF catheter developed by our group. Ex vivo three-dimensional (3D) histology was performed combining optical coherence tomography (OCT) and confocal fluorescence microscopy, providing high-resolution anatomical and molecular information, respectively, to validate in vivo findings. The microscope was combined with a serial slicer allowing for the imaging of the whole vessel automatically. Colocalization of in vivo and ex vivo results is demonstrated. Slices can then be recovered to be tested in conventional histology.

Objectives: To determine if heart rate (HR) reduction with ivabradine (IVA), a selective inhibitor of the pacemaker If current, prevents cardiac dysfunction associated with dyslipidemia. Methods: New Zealand White rabbits received either a standard diet, a 0.5% cholesterol-enriched diet only (CD), or a 0.5% CD with IVA (17 mg/kg/day) for 12 weeks. HR, left ventricular (LV) systolic function, diastolic function and LV regional myocardial performance index (MPI) were studied using echocardiography. Histological analysis included cardiac interstitial fibrosis and collagen type I fibers. Plasma levels of angiotensin II and aldosterone were quantified by immunoassays. Results: IVA reduced HR by approximately 11%. IVA improved MPI and attenuated LV diastolic dysfunction (DD) (92% mild and 8% moderate DD with IVA vs. 54% mild and 46% moderate DD in CD group). IVA also reduced atrial fibrosis (p = 0.027), ventricular fibrosis (p = 0.0002) and ventricular collagen type I (p = 0.0042). IVA decreased plasma angiotensin II levels (p = 0.042), and both angiotensin II and aldosterone levels were correlated with HR (p = 0.038 and 0.008). Conclusion: Selective HR reduction with IVA reduces DD and cardiac fibrosis in hypercholesterolemic rabbits. These beneficial effects of IVA support testing pure HR reduction in patients with diastolic heart failure.

Background and purpose: Aortic valve stenosis (AVS) is the most common valvular heart disease, and standard curative therapy remains open heart surgical valve replacement. The aim of our experimental study was to determine if apolipoprotein A-I (ApoA-I) mimetic peptide infusions could induce regression of AVS. Experimental approach: Fifteen New Zealand White male rabbits received a cholesterol-enriched diet and vitamin D2 until significant AVS was detected by echocardiography. The enriched diet was then stopped to mimic cholesterol-lowering therapy and animals were allocated randomly to receive saline (control group, n=8) or an ApoA-I mimetic peptide (treated group, n=7), three times per week for 2 weeks. Serial echocardiograms and post mortem valve histology were performed. Key results: Aortic valve area increased significantly by 25% in the treated group after 14 days of treatment (P=0.012). Likewise, aortic valve thickness decreased by 21% in the treated group, whereas it was unchanged in controls (P=0.0006). Histological analysis revealed that the extent of lesions at the base of valve leaflets and sinuses of Valsalva was smaller in the treated group compared with controls (P=0.032). The treatment also reduced calcification, as revealed by the loss of the positive relationship observed in the control group (r=0.87, P=0.004) between calcification area and aortic valve thickness. Conclusions and implications: Infusions of ApoA-I mimetic peptide lead to regression of experimental AVS. These positive results justify the further testing of high-density lipoprotein (HDL)-based therapies in patients with valvular aortic stenosis. Regression of aortic stenosis, if achieved safely, could transform the clinical treatment of this disease. British Journal of Pharmacology (2008) 154, 765–773; doi:10.1038/bjp.2008.122; published online 14 April 2008

Introduction: Left ventricular diastolic dysfunction (LVDD) is characterized by the disturbance of ventricle’s performance due to its abnormal relaxation or to its increased stiffness. Hypertrophy, inflammation, fibrosis and oxidative stress are often associated with LVDD. Hypothesis: Given the reported anti-inflammatory and anti-oxidant properties of HDL, we assessed the potential of an apoA-I mimetic peptide to improve LVDD in a recently developed rabbit model. Methods: Forty-two rabbits were fed a normal diet (n=7) or a 0.5% cholesterol-enriched diet supplemented with vitamin D2 (n=35) for an average of 16 weeks. The hypercholesterolemic rabbits were randomised to receive 6 infusions of saline (n=11), 10 mg/kg (n=12) or 30 mg/kg of an apoA-I mimetic peptide over a period of 2 weeks. Serial echocardiography was used to assess LVDD. RT-qPCR and histological staining were performed on left ventricular homogenates and sections, respectively. Results: LVDD was improved in treated groups compared to saline as shown by decreased E/Em and increased Em/Am ratio assessed by echocardiography (p≤0.05). The apoA-I mimetic decreased transferrin receptor mRNA expression (p=0.039 for 10 mg/kg group) and increased ferritin heavy chain mRNA (p=0.053 for 30 mg/kg group) compared to saline. These changes were associated with a large decrease of non-heme iron content in LV sections of the treated groups (-90% in 10 mg/kg group, p≤0.01; -72% in 30 mg/kg group, p=0.1) compared to the saline-treated group as detected by Prussian blue staining. Significant reduction of RAM-11 macrophage staining was observed in treated groups compared to saline. A correlation was observed between iron and macrophage contents in LV (R=0.59 and p=0.0002). Most of the non-heme iron and macrophage stainings were seen in coronary arteries and plaques. Anti-oxidant enzyme (SOD2) mRNA was numerically increased in the 30 mg/kg group compared to saline (p=0.08). Conclusion: ApoA-I mimetic treatment improves LVDD. The observed decrease of left ventricular iron content may lead to reduce oxidative stress as iron contributes to reactive oxygen species formation. Further work is ongoing to validate whether decreased iron-mediated toxicity is a new benefit of apoA-I mimetic therapy.

Introduction: In the dal-OUTCOMES trial, cholesteryl ester transfer protein (CETP) inhibition by dalcetrapib increased HDL-C but had a neutral effect on CV risk. However, these patients were also treated with standard-of-care therapies including statins, a drug class shown to reduce ABCA1 mRNA expression in human peripheral blood mononuclear cells (PBMC). Hypothesis: Given the role of ABCA1 in cholesterol efflux and HDL biogenesis, we determined the impact of atorvastatin on the potential beneficial effects provided by dalcetrapib and anacetrapib, two CETP inhibitors. Methods: Dalcetrapib (300 mg/kg) or anacetrapib (30 mg/kg) were administered to New Zealand White rabbits receiving or not atorvastatin (2.5 mg/kg) for 14 days. Lipid profiles were measured biochemically. ApoA-I distribution in HDL subclasses was evaluated by 1D-non-denaturing gradient gel electrophoresis (1D-NDGGE). BHK cells with inducible expression of human ABCA1 and HepG2 hepatocytes were used to measure ABCA1- and SR-BI-dépendent cholesterol efflux. Human THP-1 macrophages and PBMC isolated from rabbits were also used for efflux assays. Results: Dalcetrapib increased HDL-C by +81% (p Conclusion: CETP inhibition increased HDL-C, apoA-I levels and cholesterol efflux from different cell types. Atorvastatin had no effect on these parameters, suggesting that the increased cholesterol efflux capacity induced by CETPi is not hampered by statins.

Background and Purpose We have shown that infusions of apolipoprotein A-I (ApoA-I) mimetic peptide induced regression of aortic valve stenosis (AVS) in rabbits. This study aimed at determining the effects of ApoA-I mimetic therapy in mice with calcific or fibrotic AVS. Experimental Approach Apolipoprotein E-deficient (ApoE−/−) mice and mice with Werner progeria gene deletion (WrnΔhel/Δhel) received high-fat diets for 20 weeks. After developing AVS, mice were randomized to receive saline (placebo group) or ApoA-I mimetic peptide infusions (ApoA-I treated groups, 100 mg·kg−1 for ApoE−/− mice; 50 mg·kg−1 for Wrn mice), three times per week for 4 weeks. We evaluated effects on AVS using serial echocardiograms and valve histology. Key Results Aortic valve area (AVA) increased in both ApoE−/− and Wrn mice treated with the ApoA-I mimetic compared with placebo. Maximal sinus wall thickness was lower in ApoA-I treated ApoE−/− mice. The type I/III collagen ratio was lower in the sinus wall of ApoA-I treated ApoE−/− mice compared with placebo. Total collagen content was reduced in aortic valves of ApoA-I treated Wrn mice. Our 3D computer model and numerical simulations confirmed that the reduction in aortic root wall thickness resulted in improved AVA. Conclusions and Implications ApoA-I mimetic treatment reduced AVS by decreasing remodelling and fibrosis of the aortic root and valve in mice.

Excessive neointima formation often occurs after arterial injury. Interleukin-1β (IL-1β) is a potent pleiotropic cytokine that has been shown to regulate neointimal proliferation. We investigated the effects of the IL-1β modulator gevokizumab in a rat carotid denudation model.Sprague-Dawley rats were subjected to balloon denudation of the right carotid artery and were then randomized to receive a single subcutaneous infusion immediately after balloon injury of saline (control group, n = 13) or gevokizumab (gevokizumab groups, n = 15 in each group: 1, 10 and 50 mg/kg). We evaluated the treatment effects on carotid intima-media thickness (IMT) using ultrasonography, on endothelial regrowth using Evans Blue staining and on inflammatory response using histology. We also assessed the effects of IL-1β and gevokizumab on human umbilical vein endothelial cells (HUVEC) and rat smooth muscle cells.We found that carotid IMT, in the proximal part of the denuded artery at day 28, was decreased by gevokizumab 1 mg/kg compared with controls. Neointima area and the intima/media area ratio were both reduced in the gevokizumab 1 mg/kg-treated group. Gevokizumab at the 1 mg/kg dose also improved endothelial regrowth. No effect was observed with gevokizumab 10 or 50 mg/kg. Gevokizumab also decreased the inflammatory effect of IL-1β in in vitro cell experiments and protected HUVECs from IL-1β's deleterious effects on cell migration, apoptosis and proliferation.A single administration of gevokizumab 1 mg/kg improves endothelial regrowth and reduces neointima formation in rats following carotid denudation, at least in part through its beneficial effects on endothelial cells.

Purpose : Aortic valve stenosis is the most common valvular heart disease, and standard curative therapy remains open-heart surgical valve replacement. The aim of our experimental study was to determine if ApoA-I mimetic peptide infusions could induce regression of aortic valve stenosis. Methods : Twenty-seven New-Zealand White male rabbits received a cholesterol-enriched diet and vitamin D 2 until significant aortic valve stenosis was detected by echocardiography. The enriched diet was then stopped to mimic cholesterol-lowering therapy and animals were randomized to receive saline (control group, n =14) or an ApoA-I mimetic peptide (treated group, n =13), 3 times per week for 2 weeks. Serial echocardiograms and post mortem valve histology were performed. Results : Aortic valve area improved significantly in the treated group compared to controls after 7 days (20.9±0.9 mm 2 vs. 18.2±0.6 mm 2 , P 2 vs. 19.5±0.6 mm 2 , P =0.0032) (increases of 19.2% vs. 9.1%), and 14 days of treatment (22.4±0.9 mm 2 vs. 20.4±0.6 mm 2 , P =0.0028) (increases of 24.4% vs. 14.2%). Likewise, aortic valve thickness decreased by 19% after 14 days of treatment in the treated group (0.951±0.070 mm at baseline vs. 0.768±0.074 mm at follow-up) whereas it was unchanged in controls ( P P =0.032). The ApoA-I mimetic peptide treatment also leads to a reduction in aortic valve calcifications as revealed by the loss of the positive relationship observed in the control group ( r =0.87, P =0.004) between calcifications area and aortic valve thickness. Conclusions : Infusions of an ApoA-I mimetic peptide lead to regression of experimental aortic valve stenosis. These positive results justify the further testing of HDL-based therapies in patients with valvular aortic stenosis. Regression of aortic stenosis, if achieved safely, could transform our clinical approach of this disease.

Coronary artery disease is characterized by atherosclerotic plaque formation. Despite impressive advances in intravascular imaging modalities, in vivo molecular plaque characterization remains challenging, and different multimodality imaging systems have been proposed. We validated an engineered bimodal intravascular ultrasound imaging (IVUS) / near-infrared fluorescence (NIRF) imaging catheter in vivo using a balloon injury atherosclerosis rabbit model. Rabbit aortas and right iliac arteries were scanned in vivo after indocyanine green (ICG) injection, and compared to corresponding ex vivo fluorescence and white light images. Areas of ICG accumulation were colocalized with macroscopic atherosclerotic plaque formation. In vivo imaging was performed with the bimodal catheter integrating ICG-induced fluorescence signals into cross-sectional IVUS imaging. In vivo ICG accumulation corresponded to ex vivo fluorescence signal intensity and IVUS identified plaques.

Purpose: High-density lipoprotein (HDL) infusions induce rapid improvement of atherosclerosis and aortic valve stenosis (AVS) in animals but their effect on left ventricular diastolic dysfunction (LVDD) remains unknown. Our study aimed at evaluating the effects of the HDL mimetic peptide CER-522 on LVDD in a rabbit model of AVS. Methods: Thirty-five (35) male New-Zealand White rabbits were fed with a 0.5% cholesterol- and vitamin D2-enriched diet until mild AVS was detected by echocardiography (mean duration of diet: 16 weeks). Animals were then randomized to receive saline (control group, n = 11) or the HDL mimetic peptide CER-522 at doses of 10 mg/kg (n = 12) or 30 mg/kg (n = 12) 3 times per week for 2 weeks. Serial echocardiograms and end-of-study histological examination of the aortic valve and left ventricle were performed. Results: Aortic valve area increased at day 10 in the CER-522 10 mg/kg group (P = 0.016) and at day 14 in both the CER-522 10 and 30 mg/kg treated groups compared to controls (P = 0.011 and P = 0.033, respectively). Aortic root strain index was higher in both CER-522 treated groups vs controls at 14 days of treatment (P = 0.007 and P = 0.047, respectively). At day 14, compared to controls, CER-522 normalized early filling deceleration time (P < 0.001 for both treated groups) and E wave deceleration rate (P < 0.001 for both treated groups), decreased E/Em ratio (P < 0.001 and P = 0.001 for the 10 and 30 mg/kg groups, respectively) and increased Em/Am ratio (P < 0.001 and P = 0.025). Although all groups had similar amount of global LVDD before treatment, moderate LVDD was present in 36.4, 16.7 and 0% of animals in the control, CER-522 10 and 30 mg/kg groups respectively at study end (P = 0.216 and P < 0.001 for 10 and 30 mg/kg groups vs controls). Histological analyses showed that RAM-11 positive cells (macrophages) in LV myocardium and pericoronary fibrosis were reduced with CER-522 10 mg/kg compared to controls (P = 0.006 and P = 0.018). The number of apoptotic cardiomyocytes was reduced by CER-522 30 mg/kg vs controls (P = 0.010; P=NS for 10 mg/kg). Coronary artery atheroma area was decreased in both CER-522 groups vs controls (P = 0.028 and P = 0.044 for 10 and 30 mg/kg groups), as was the case for the coronary external/internal elastic lamina area ratio (P = 0.003 and P = 0.010). Conclusions: The HDL mimetic peptide CER-522 improves LVDD in rabbits while also reducing cardiomyocyte apoptosis, mild AVS, aortic root rigidity and coronary atherosclerosis.