Rabbit Atherosclerotic Plaques Research Articles

Molecular Imaging of Fibroblast Activation in Rabbit Atherosclerotic Plaques: a Preclinical PET/CT Study.

Atherosclerosis remains the pathological basis of myocardial infarction and ischemic stroke. Early and accurate identification of plauqes is crucial to improve clinical outcomes of atherosclerosis patients. Our study aims to evaluate the potential value of fibroblast activation protein inhibitor (FAPI)-04 PET/CT in identifying plaques via a preclinical rabbit model of atherosclerosis. New Zealand white rabbits were fed high-fat diet (HFD), and randomly divided into the model group injured by the balloon, and the sham group only with incisions. Ultrasound was performed to detect plaques, and FAPI-avid was determined through Al18F-NOTA-FAPI-04 PET/CT. Mean standardized uptake values (SUVmean) in lesions were compared, and biodistribution of Al18F-NOTA-FAPI-04 and target-to-background ratios (TBRs) were calculated. Histological staining was performed to display arterial plaques, and autoradiography (ARG) was employed to measure the in vitro intensity of Al18F-NOTA-FAPI-04. At last, the correlation among FAP levels, plaque area, SUVmean values and fibrous cap thickness was assessed. The rabbit carotid and abdominal atherosclerosis model was established. Al18F-NOTA-FAPI-04 showed a higher uptake in carotid plaques (SUVmean 1.32 ± 0.11) and abdominal plaques (SUVmean 0.73 ± 0.13) compared to corresponding controls (SUVmean 1.07 ± 0.06; 0.46 ± 0.03) (P < 0.05). Biodistribution analysis of Al18F-NOTA-FAPI-04 revealed that the bigger plaques were delineated with higher TBRs. Pathological staining showed the formation of arterial plaques, and ARG staining exhibited a higher intensity of Al18F-NOTA-FAPI-04 in the bigger plaques. Lastly, plaque area was found to be positively correlated to FAP expression and SUVmean, while FAP expression was negatively correlated to fibrous cap thickness of plaques. We successfully achieve molecular imaging of fibroblast activation in atherosclerotic lesions of rabbits, suggesting Al18F-NOTA-FAPI-04 PET/CT may be a potentially valuable tool to identify plaques.

Accumulation of Iron Oxide-Based Contrast Agents in Rabbit Atherosclerotic Plaques in Relation to Plaque Age and Vulnerability Features.

In this study, a detailed characterization of a rabbit model of atherosclerosis was performed to assess the optimal time frame for evaluating plaque vulnerability using superparamagnetic iron oxide nanoparticle (SPION)-enhanced magnetic resonance imaging (MRI). The progression of atherosclerosis induced by ballooning and a high-cholesterol diet was monitored using angiography, and the resulting plaques were characterized using immunohistochemistry and histology. Morphometric analyses were performed to evaluate plaque size and vulnerability features. The accumulation of SPIONs (novel dextran-coated SPIONDex and ferumoxytol) in atherosclerotic plaques was investigated by histology and MRI and correlated with plaque age and vulnerability. Toxicity of SPIONDex was evaluated in rats. Weak positive correlations were detected between plaque age and intima thickness, and total macrophage load. A strong negative correlation was observed between the minimum fibrous cap thickness and plaque age as well as the mean macrophage load. The accumulation of SPION in the atherosclerotic plaques was detected by MRI 24 h after administration and was subsequently confirmed by Prussian blue staining of histological specimens. Positive correlations between Prussian blue signal in atherosclerotic plaques, plaque age, and macrophage load were detected. Very little iron was observed in the histological sections of the heart and kidney, whereas strong staining of SPIONDex and ferumoxytol was detected in the spleen and liver. In contrast to ferumoxytol, SPIONDex administration in rabbits was well tolerated without inducing hypersensitivity. The maximum tolerated dose in rat model was higher than 100 mg Fe/kg. Older atherosclerotic plaques with vulnerable features in rabbits are a useful tool for investigating iron oxide-based contrast agents for MRI. Based on the experimental data, SPIONDex particles constitute a promising candidate for further clinical translation as a safe formulation that offers the possibility of repeated administration free from the risks associated with other types of magnetic contrast agents.

Experimental Study of Ultrafine Superparamagnetic Iron Oxide-Enhanced MRI in an Atherosclerotic Plaque Model.

Unstable atherosclerotic (AS) plaques contain numerous macrophages that can phagocytose a specific contrast agent, namely ultrafine superparamagnetic iron oxide (USPIO). This study prepared USPIO nanoparticles, defined their physical and magnetic properties and transmission characteristics, explored the feasibility of their use as a magnetic resonance negative contrast agent, and used them to detect unstable AS plaque as a specific marker of macrophages. We randomly divided 20 healthy male New Zealand white rabbits into two groups, experimental and control (n = 10 each). The experimental group was fed pure high-fat feed. The magnetic resonance imaging (MRI) performance of arterial plaque before and after USPIO enhancement compare with pathological results. The experimental group successfully established a rabbit AS plaque model, and 8 of 10 rabbits developed AS plaque. USPIO-enhanced T2W1 sequence showed that the central signal of the plaque was reduced, and the signal-to-noise ratio of the vascular wall was the lowest at 96 h. The control group showed no change in the wall signal before and after enhancement. The USPIO-enhanced PJN2D-TOF sequence showed dotted filling defects on the tube wall. Pathological examination showed that USPIO nanoparticles were mainly deposited under the arterial intima. A simple high-fat diet can establish a rabbit AS plaque model, and USPIO-enhanced MRI reflects the condition of rabbit AS plaque, which is helpful in evaluating AS lesion diagnosis.

Validation of Ultrasound Super-Resolution Imaging of Vasa Vasorum in Rabbit Atherosclerotic Plaques.

Acute coronary syndromes and strokes are mainly caused by atherosclerotic plaque (AP) rupture. Abnormal increase of vasa vasorum (VV) is reported as a key evidence of plaque progression and vulnerability. However, due to their tiny size, it is still challenging to noninvasively identify VV near the major vessels. Ultrasound super resolution (USR), a technique that provides high spatial resolution beyond the acoustic diffraction limit, demonstrated an adequate spatial resolution for VV detection in early studies. However, a thorough validation of this technology in the plaque model is particularly needed in order to continue further extended preclinical studies. In this letter, we present an experiment protocol that verifies the USR technology for VV identification with subsequent histology and ex vivo micro-computed tomography ( μ CT). Deconvolution-based USR imaging was applied on two rabbits to identify the VV near the AP in the femoral artery. Histology and ex vivo μ CT imaging were performed on excised femoral tissue to validate the USR technique both pathologically and morphologically. This established validation protocol could facilitate future extended preclinical studies toward the clinical translation of USR imaging for VV identification.

Super-resolution ultrasound imaging beyond the acoustic diffraction limit

Contrast enhanced ultrasound (CEU) imaging technologies using microbubbles (MBs) provide superior contrast of vasculatures, effectively suppressing the surrounding tissue signals, but the spatial resolution remains to the acoustic diffraction limit. By localizing the center of each MBs unprecedented high spatial resolution beyond the acoustic diffraction limit can be achieved. However, some methods of localizing each center of the signals from individual MBs that only applies symmetrically distributed signal amplitude require a large number of imaging frames, especially when MBs are densely clumped, therefore result in a long scan time that is not ideal for in vivo scan under physiologic conditions. In this paper, we present an innovative approach using deconvolution technique that will allow for identifying signals from individual MBs from dense population in any forms even grouped together within the full-width-at-half-maximum (FWHM) of the point spread function (PSF) of the US probe. In this way, no collected frame sets require to be excluded for image reconstruction, therefore scan time can be reduced significantly. In vivo application of this new approach in identifying vasa vasorum in rabbit atherosclerotic plaque model will be presented. Some technical limitations including background noise as well as motion artifact will be discussed.

Detection of Atherosclerotic Plaques in the Rabbit Aorta Using Ultrasound Microbubbles Conjugated to Interleukin-18 Antibodies.

BackgroundThe purpose of the study was to investigate the ability of microbubbles (MBs) targeting interleukin-18 (IL-18) to detect plaques in a rabbit atherosclerotic plaque model.Material/MethodsA rabbit atherosclerotic plaque model was established. The locations of the atherosclerotic plaques were verified by two-dimensional scanning and color Doppler flow imaging. An IL-18 antibody was conjugated to naked MBs (MBc) using the biotin-streptavidin conjugation method, resulting in the formation of MBIL-18. MBc and MBIL-18 were then used for contrast-enhanced ultrasound (CEUS) studies. The locations of CD34 and IL-18 within the plaques were determined by immunohistochemistry, and IL-18 expression levels in the plaques were determined by Western blot analysis. The relationships between IL-18 expression and the contrast intensity of the 2 MBs were analyzed.ResultsMBc and MBIL-18 were both uniformly dispersed. Fluorescence microscopy and flow cytometry revealed that IL-18 was successfully conjugated to MBs. CEUS images showed that the intensity of the MBIL-18 signal was substantially enhanced and prolonged compared with that of the MBc signal. Immunohistochemistry showed that CD34 expression was significantly increased in the plaques and that IL-18 was mainly located in the inner parts and base of the atherosclerotic plaques. Western blot analysis revealed that IL-18 expression was higher in the plaque regions. Correlation analysis showed that IL-18 expression was correlated with the contrast intensity of MBIL-18 (r=0.903, P<0.05) but not with MBc (r=0.540, P>0.05).ConclusionsMBs targeting IL-18 may be a novel, noninvasive method of diagnosing atherosclerotic plaques.

Radiofrequency Ablation of the Atherosclerotic Plaque: a Proof of Concept Study in an Atherosclerotic Model

Increased plaque vascularization is causatively associated with the progression of unstable atherosclerotic vessel disease. We investigated the safety and efficacy of heat-generating radiofrequency ablation (RFA) in reducing the number of vessels in the plaque and adventitia and its effect on plaque size and composition. To this end, New Zealand White rabbits were fed a cholesterol-enriched diet and subjected to balloon denudation of the infrarenal aorta to induce atherosclerotic plaque formation. After 13 weeks, the proximal or distal half of the infrarenal aorta was exposed to transluminal RFA. The untreated half served as an intra-individual control. Optical coherence tomography (OCT) was performed directly after RFA. We found that RFA on the rabbit atherosclerotic plaque is safe and leads to decreased intraplaque vessel density and smooth muscle cell content but does not affect other components of plaque composition or size.

Polyglucose nanoparticles with renal elimination and macrophage avidity facilitate PET imaging in ischaemic heart disease

Tissue macrophage numbers vary during health versus disease. Abundant inflammatory macrophages destruct tissues, leading to atherosclerosis, myocardial infarction and heart failure. Emerging therapeutic options create interest in monitoring macrophages in patients. Here we describe positron emission tomography (PET) imaging with 18F-Macroflor, a modified polyglucose nanoparticle with high avidity for macrophages. Due to its small size, Macroflor is excreted renally, a prerequisite for imaging with the isotope flourine-18. The particle’s short blood half-life, measured in three species, including a primate, enables macrophage imaging in inflamed cardiovascular tissues. Macroflor enriches in cardiac and plaque macrophages, thereby increasing PET signal in murine infarcts and both mouse and rabbit atherosclerotic plaques. In PET/magnetic resonance imaging (MRI) experiments, Macroflor PET imaging detects changes in macrophage population size while molecular MRI reports on increasing or resolving inflammation. These data suggest that Macroflor PET/MRI could be a clinical tool to non-invasively monitor macrophage biology.

Abstract 12082: Radiofrequency Ablation of the Atherosclerotic Plaque: A Safety and Feasibility Study in an Atherosclerotic Rabbit Model

Introduction: Angiogenesis from the vasa vasorum plays a key role in atheroprogression and plaque vulnerability. Immature neovessels are prone to leakage, hence facilitating additional lipid and inflammatory cell accumulation and increasing intraplaque hemorrhage risk. Hypothesis: Heat-generating radiofrequency ablation (RFA) safely reduces vasa vasorum, decreases inflammatory cell influx and stabilizes the plaque. Methods: Thirteen New-Zealand White rabbits on a 0.5% cholesterol-enriched diet were subjected to balloon denudation of the infrarenal aorta. After 13 weeks, either the proximal or distal half of the infrarenal aorta was exposed to transluminal RFA, the untreated half served as an intra-individual control. Optical coherence tomography (OCT) was performed directly after RFA. Results: Within 24 hours after RFA, no plaque ruptures or adverse events were observed by OCT or histology. Treated regions showed widespread decellularization of intimal, medial and (to a lesser extent) adventitial layers, which were recellularized after 7 and 28 days. Vasa vasorum density was not different between both regions at short-term (24 hours; RFA 19.8±5.1 vs. control 29.2±8.3/mm 2 ; p=0.15) or long-term follow-up (7 days; 39.1±11.5 vs. 41.7±8.6/mm 2 ; p=0.81 and 28 days; 46.4±13.4 vs. 38.5±6.6/mm 2 ; p=0.06). After 7 days, treated regions were comparable to control areas with respect to plaque size (5.6±1.7 vs. 5.5±1.5mm 2 ; p=0.62), intimal collagen (34.6±11.9 vs. 32.7±13.0%; p=0.22) and macrophage content (14.3±2.8 vs. 18.5±5.8%; p=0.32). RFA-treated regions did show a major decrease in intimal smooth muscle cell (αSMA) content (1.8±0.4 vs. 18.0±5.9%; p=0.038), which remained decreased after 28 days (6.8±3.8 vs. 12.6±1.6%; p=0.020). Again, after 28 days no differences were observed in plaque area (4.6±0.9 vs. 4.4±1.1 mm 2 ; p=0.46), intimal collagen (39.2±9.1 vs. 40.3±12.4%; p=0.85) or macrophage content (13.2±2.5 vs. 12.6±2.0%; p=0.73). Conclusion: Radiofrequency ablation of the rabbit atherosclerotic plaque is safe and leads to a major reduction in αSMA content, but does not affect other aspects of plaque stability over time. It may thus serve as a therapeutic option for major (re)stenosis in peripheral arterial disease.

Polymer-Lipid Hybrid Theranostic Nanoparticles Co-Delivering Ultrasmall Superparamagnetic Iron Oxide and Paclitaxel for Targeted Magnetic Resonance Imaging and Therapy in Atherosclerotic Plaque.

Magnetic resonance imaging (MRI) combined with ultrasmall superparamagnetic iron oxide (USPIO) is effective for the detection of atherosclerotic (AS) plaque, and paclitaxel is effective for the treatment of AS. C11 is a polypeptide with high affinity and specificity for collagen IV. It is abundantly expressed in the outer layer of AS plaque. This study aimed to develop USPIO + paclitaxel-loaded polymer-lipid hybrid theranostic nanoparticles conjugated with C11 (UP-NP-C11) for simultaneous imaging and treatment AS plaque. UP-NP-C11 was developed by the nanoprecipitation method, and the theranostics of AS plaque by UP-NP-C11 were evaluated both in vitro and in the rabbit AS model. UP-NP-C11 was of desired particle size (140.2 nm), showed encapsulation efficiency of 35.5% and 55.2% for USPIO and paclitaxel, respectively, and had drug release profile. The accumulation of USPIO in Matrigel (containing abundant collagen IV) and macrophages coated on the Matrigel was significantly higher in the UP-NP-C11-treated group than in the group treated by UP-NP (USPIO + paclitaxel-loaded nanoparticles). Thus, UP-NP-C11 exerted better growth inhibitory effect and MRI ability in macrophages than UP-NP. Significantly, UP-NP-C11 showed better in vivo MRI ability and therapeutic effect in rabbit AS plaque than UP-NP and commercial USPIO + paclitaxel, and Prussian blue staining revealed significantly greater accumulation of USPIOs in the UP-NP-C11-treated group than in the control group. Furthermore, UP-NP-C11 did not cause severe toxicity to the rabbits. UP-NP-C11 represents a potential approach for targeted MRI and therapy in AS plaque.

Effect of rosuvastatin on atherosclerotic plaque stability: An intravascular ultrasound elastography study

ObjectivesThe present study aimed to investigate the effect of potent rosuvastatin therapy on plaque mechanical stabilization as seen on IVUSE. Methods14 purebred New Zealand rabbits were fed a high-cholesterol diet; the abdominal aorta endothelium was balloon-injured after 2 weeks; at week 13, 7 rabbits received rosuvastatin (1.5 mg/kg/day), and the other 7 received an equal volume of saline. IVUS images of abdominal aortas were acquired, and 2 consecutive frames near the end-diastole images in situ were used to construct an IVUS elastogram. ResultsControl rabbits showed a significant increase in shear strain (SS) and area strain (AS) in total plaques. The rosuvastatin group showed no change in SS and AS, but serum TG and LDL-C levels were reduced, with less lipid deposition, macrophage infiltration, production of proinflammatory cytokines and apoptosis in plaques. The changes in SS and AS from baseline between groups significantly differed (SS: 1.15 (1.96) % vs. −0.99 ± 2.83%, p = 0.013; AS: 1.25 (2.29) % vs. −1.67 ± 5.05%, p = 0.022). At follow-up, for controls, strain values were increased in the shoulder of eccentric plaques (SS: 2.66 ± 1.31% vs. 4.86 ± 1.93%, p = 0.016; AS: 4.45 ± 2.33% vs. 7.91 ± 2.74%, p = 0.009) but not the plaque body. Changes in SS and AS in the plaque shoulder differed between the control and rosuvastatin groups (SS: 2.20 ± 2.17% vs. −0.87 ± 3.31%, p = 0.028; AS: 2.10 (4.61) % vs. −2.75 ± 5.97%, p = 0.009). ConclusionRosuvastatin therapy in rabbits with atherosclerotic plaques led to less vulnerable plaque features. IVUSE is a very sensitive technique for detecting pharmacologically-induced mechanical changes in rabbit atherosclerotic plaques.

Relationship between arterial atheromatous plaque morphology and platelet–associated miR–126 and miR–223 expressions

To study the expression of miR-126 and miR-223 in platelet of rabbit arterial plaque models, and explore its correlation with plaque morphology. Rabbit arterial plaque models were established, peripheral blood of models and control animals was collected. Plaque morphologies were divided into type I, type II and type III based on angiography plaque morphology and Ambrose method. Platelet isolation kit was applied to isolate and purify peripheral blood platelets, CD45 immunomagnetic beads were used to remove the residual white blood cells. The miRNAs of platelets was extracted by miRNA Isolation Kit, and expressions of miR-126 and miR-223 of the platelets samples were detected by Real-time PCR. The correlation between plaque morphology and platelet-associated miR-126 and miR-223 expressions were analyzed. Expressions of target gene VCAM-1 and P2Y12 receptors of miR-126 and miR-223 in the atherosclerosis plaque of rabbit model were detected by Western blot. Relative expression levels of miR-126 and miR-223 in the model group were 0.27±0.10 and 0.71±0.14, respectively. Plaque morphology was divided into types I, II and III; and miR-126 and miR-223 expression levels were detected in each type. Expression levels of miR-126 in each type were 0.42±0.07, 0.17±0.11 and 0.22±0.15, respectively; and expression levels of miR-223 in each type are 0.68±0.02, 0.57±0.06 and 0.88±0.10, respectively. Relative to the control group, miR-126 and miR-223 known target genes in VCAM-1 and P2Y12 receptors increased platelets in rabbit atherosclerotic plaque models (P<0.05). Relative to normal control animals, miR-126 and miR-223 platelets were reduced in the rabbit atherosclerotic plaque model group (P<0.05). In the type II plaque morphology group, miR-126 was greatly reduced; and there is no significant correlation between miR-223 and plaque morphology.

Tiaozhi Tongmai Granules reduce atherogenesis and promote the expression of ATP-binding cassette transporter A1 in rabbit atherosclerotic plaque macrophages and the liver

ObjectiveATP-binding cassette transporter A1 (ABCA1) is an integral membrane protein that plays a key role in cellular lipid metabolism, preventing the accumulation of lipids that contribute to the initiation and progression of atherosclerosis. Tiaozhi Tongmai Granules are a Chinese herbal compound that is capable of treating atherosclerosis. This study was designed to explore the potential pharmacological mechanism by which Tiaozhi Tongmai Granules protect against atherosclerosis. MethodsForty-nine male New Zealand rabbits were randomly divided into seven groups: normal control group, normal diet; model groups 1 and 2: balloon injury and high-fat diet for 6 or 12 weeks; statin groups 1 and 2: balloon injury and high-fat diet plus atorvastatin for 6 or 12 weeks; and Chinese herb groups 1 and 2: balloon injury and high-fat diet plus Tiaozhi Tongmai Granules for 6 or12 weeks. The granules were administered at a dose of 1.14 g/kg/d, with atorvastatin (1.14 mg/kg/d) serving as positive control. Serum lipid profiles and liver function indices were measured. Atherogenesis was viewed after H&E staining and quantified by thickened intimal area percentage and maximal intimal thickness percentage. The ABCA1 protein expression in atherosclerotic plaque macrophages of the common carotid arteries (CCA), thoracic aortae (TA), and liver tissues were observed by immunohistochemical staining and evaluated using mean optical density (OD) value in macrophages and ABCA1-positive hepatocyte number. ResultsCompared with model group 1 at week 6, Chinese herb group 1 and statin group 1 displayed significant reductions in total cholesterol (TC) (P = 0.027, 0.012) and low-density lipoprotein cholesterol (LDL-C) (P = 0.039, 0.028) levels, as well as marked increases in ABCA1-positive hepatocyte numbers (P all <0.001), and only statin group 1 displayed a markedly reduced maximal intimal thickness percentage in the CCA (P = 0.018). Compared with model group 2 at week 12, Chinese herb group 2 and statin group 2 all presented significant reductions in TC (P = 0.011, 0.003), LDL-C (P = 0.017, 0.010) and thickened intimal area percentage in the CCA (P = 0.001, 0.022), as well as prominent increases in the ABCA1 OD value of both the CCA (P = 0.001, 0.039) and TA (P = 0.001, 0.025) and positive hepatocyte number (P all <0.001). Chinese herb group 2 had a markedly reduced maximal intimal thickness percentage compared with model group 2 (P = 0.006) and a higher positive hepatocytes number than statin group 2 (P = 0.001). ConclusionsTiaozhi Tongmai Granules appear to have an anti-atherogenic effect that is most likely mediated by simultaneously upregulating the protein expression of ABCA1 in rabbit atherosclerotic plaque macrophages and in the liver.

Extracellular Acidosis Is a Novel Danger Signal Alerting Innate Immunity via the NLRP3 Inflammasome

Local acidosis has been demonstrated in ischemic tissues and at inflammatory sites. Acidic extracellular pH triggers NLRP3 inflammasome activation and interleukin-1β secretion in human macrophages. Acidic pH represents a novel danger signal alerting the innate immunity. Local acidosis may promote inflammation at ischemic and inflammatory sites. Local extracellular acidification has been demonstrated at sites of ischemia and inflammation. IL-1β is one of the key proinflammatory cytokines, and thus, its synthesis and secretion are tightly regulated. The NLRP3 (nucleotide-binding domain leucine-rich repeat containing family, pyrin domain containing 3) inflammasome complex, assembled in response to microbial components or endogenous danger signals, triggers caspase-1-mediated maturation and secretion of IL-1β. In this study, we explored whether acidic environment is sensed by immune cells as an inflammasome-activating danger signal. Human macrophages were exposed to custom cell culture media at pH 7.5-6.0. Acidic medium triggered pH-dependent secretion of IL-1β and activation of caspase-1 via a mechanism involving potassium efflux from the cells. Acidic extracellular pH caused rapid intracellular acidification, and the IL-1β-inducing effect of acidic medium could be mimicked by acidifying the cytosol with bafilomycin A1, a proton pump inhibitor. Knocking down the mRNA expression of NLRP3 receptor abolished IL-1β secretion at acidic pH. Remarkably, alkaline extracellular pH strongly inhibited the IL-1β response to several known NLRP3 activators, demonstrating bipartite regulatory potential of pH on the activity of this inflammasome. The data suggest that acidic environment represents a novel endogenous danger signal alerting the innate immunity. Low pH may thus contribute to inflammation in acidosis-associated pathologies such as atherosclerosis and post-ischemic inflammatory responses.

Pravastatin Inhibits Plaque Rupture and Subsequent Thrombus Formation in Atherosclerotic Rabbits with Hyperlipidemia

Previous studies have demonstrated that statin can reduce the risk of acute coronary syndrome. In order to explore the mechanism, we observed the effects of pravastatin on plaque stability in atherosclerotic rabbits. Sixteen male rabbits were fed with a high fat diet following their damaged abdominal aortic endothelium by using catheter. Eight of them were administered with pravastatin (10 mg·kg(-1)·d(-1)) for 4 weeks. Then the rabbit atherosclerotic plaque rupture and thrombosis were triggered by injection of viper venom and histamine. Compared with model group, the thrombus area on aorta in pravastatin-treated group was reduced. Fibre cap on plaque was more thick and integrant, and inflammatory cell infiltration was also decreased. Serum total cholesterol, triglyceride, low density lipoprotein-cholesterol and contents of cholesterol in abdominal aorta were decreased. 6-Keto-prostaglandin F(1α) (6-keto-PGF(1α)) level and ratio of 6-keto-PGF(1α)/thromboxane B(2) (TXB(2)) in aorta were significantly increased. These results suggested that pravastatin could increase plaque stability and inhibit thrombosis through both lipid-dependent and lipid-independent way.

Correlation between parameters of contrast-enhanced ultrasound and microvessel density in rabbit atherosclerotic plaques

Objective To evaluate the correlation between quantitative parameters of contrastenhanced ultrasound(CEUS) and microvessel density (MVD) of rabbit atherosclerotic plaques.Methods Twenty-six male New Zealand rabbits were damaged abdominal aortic by balloon expanded,fed with high fat 12 weeks later.Thirty two plaques were detected using conventional ultrasound examination.The maximum area,luminal area stenosis rate and the maximum thickness of each plaque were recorded.CEUS was performed on 32 plaques and enhancement images were offline analyzed quantitatively by ACQ software and the parameters such as arrival time(AT),time to peak(TTP),basic intensity(BI),and peak intensity(PI) were acquired.Enhanced intensity (EI) was calculated according to the formula:EI =PI- BI.Then the experimental rabbits were killed for pathological examination and CD34 monoclonal antibody immunohistochemical detection.MVD was counted under the microscope.The correlation between the parameters of CEUS and MVD was analyzed.Based on pathological findings,the plaques were divided into vulnerable plaques and stable plaques.Results EI and MVD of vulnerable plaques group were significantly higher than that of stable plaques group [EI:(26.36 ± 1.44) dB vs (23.90 ± 2.92)dB,t =- 3.243,P =0.001 ；MVD:(5.23 ± 1.16)/mm2 vs (2.47 ± 1.12)/mm2,Z =- 4.378,P ＜0.001].The maximum area,luminal area stenosis rate,the maximum thickness and PI,AT,TTP had no significant difference between two groups( P ＞0.05).The value of EI showed significant positive correlation with MVD ( r =0.676,P ＜0.001).Conclusions EI can accurately evaluate the angiogenesis of rabbit atherosclerotic plaques,which can be regarded as a useful index to distinguish between vulnerable and stable plaques. Key words: Ultrasonography; Microbubbles; Aorta,abdominal; Atherosclerosis; Microvessel density

Rabbit atherosclerotic vulnerable plaques features of MRI: an experimental study

ObjectiveTo investigate the difference between the vulnerable plaques and stable plaques in MR images.MethodsAtherosclerosis was induced in twenty male New Zealand White rabbits by high cholesterol diet and balloon injury...

Inhibition of inositol monophosphatase by lithium chloride induces selective macrophage apoptosis in atherosclerotic plaques

Lithium chloride (LiCl) inhibits inositol monophosphatase (IMPase) at therapeutic concentrations. Given that LiCl induces death in cultured macrophages and that macrophages play an active role in atherosclerotic plaque destabilization, we investigated whether LiCl would induce selective macrophage death to stabilize the structure of the plaque. The effect of LiCl was assessed on macrophages and smooth muscle cells (SMCs) in culture, in isolated atherosclerotic carotid arteries from rabbits and after local in vivo treatment via osmotic minipumps to rabbits with collared atherosclerotic carotid arteries. In addition, in vitro experiments were performed to elucidate the mechanism of LiCl-induced macrophage death. In vitro, whereas SMCs were highly resistant, LiCl induced macrophage death characterized by externalization of phosphatidylserine, caspase-3 cleavage and DNA fragmentation, all indicative of apoptosis. LiCl reduced inositol-1,4,5-trisphosphate levels in macrophages. Moreover, the IMPase inhibitor L-690 330 as well as IMPase gene silencing induced macrophage apoptosis. Both in vitro treatment of rabbit atherosclerotic carotid arteries with LiCl and local in vivo administration of LiCl to the plaques decreased plaque macrophages through apoptosis, as shown by terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick-end labelling (TUNEL), without affecting SMCs. Vasomotor studies in vitro showed that LiCl did not affect the functionality of SMCs and endothelial cells. LiCl selectively decreased the macrophage load in rabbit atherosclerotic plaques via IMPase inhibition without affecting the viability or functionality of SMCs and endothelial cells. These data provide evidence for local administration of an IMPase inhibitor to stabilize atherosclerotic plaques.

Use of contrast-enhancement and high-resolution 3D black-blood MR Imaging to identify inflammation in rabbit atherosclerotic plaques

Methods Ten atherosclerotic rabbits and three normal control rabbits underwent high-resolution 3D contrast-enhanced black-blood MRI. MR images and the corresponding histopathological sections were divided into four quadrants. Plaque composition was analyzed for each quadrant according to histopathological (percent of lipid-rich, fibrous, macrophage area and microvessel density) and imaging criteria (enhancement ratio (ER), ER=SIpost/SIpre).

Sphingosine kinase regulates oxidized low density lipoprotein-mediated calcium oscillations and macrophage survival

We recently reported that oxidized LDL (oxLDL) induces an oscillatory increase in intracellular calcium ([Ca(2+)](i)) levels in macrophages. Furthermore, we have shown that these [Ca(2+)](i) oscillations mediate oxLDL's ability to inhibit macrophage apoptosis in response to growth factor deprivation. However, the signal transduction pathways by which oxLDL induces [Ca(2+)](i) oscillations have not been elucidated. In this study, we show that these oscillations are mediated in part by intracellular mechanisms, as depleting extracellular Ca(2+) did not completely abolish the effect. Inhibiting sarco-endoplasmic reticulum ATPase (SERCA) completely blocked [Ca(2+)](i) oscillations, suggesting a role for Ca(2+) reuptake by the ER. The addition of oxLDL resulted in an almost immediate activation of sphingosine kinase (SK), which can increase sphingosine-1-phosphate (S1P) levels by phosphorylating sphingosine. Moreover, S1P was shown to be as effective as oxLDL in blocking macrophage apoptosis and producing [Ca(2+)](i) oscillations. This suggests that the mechanism in which oxLDL generates [Ca(2+)](i) oscillations may be 1) activation of SK, 2) SK-mediated increase in S1P levels, 3) S1P-mediated Ca(2+) release from intracellular stores, and 4) SERCA-mediated Ca(2+) reuptake back into the ER.