Ruiping Huang

Noninvasive Detection and Localization of Vulnerable Plaque and Arterial Thrombosis With Computed Tomography Angiography/Positron Emission Tomography

Background— It has been shown that plaque uptake of fluorodeoxyglucose is proportional to macrophage density. We tested the hypothesis that arterial thrombosis occurs in areas with high fluorodeoxyglucose uptake and that computed tomography angiography (CTA) can detect thrombi in vessels. Methods and Results— Twenty New Zealand White rabbits were studied before and after atherosclerosis induction through de-endothelialization and a high-cholesterol diet; 14 were then thrombus triggered. CTA/positron emission tomography scans were performed before cholesterol diet, at the middle diet feeding, at the end of diet feeding, and after triggering. Serum inflammatory markers were measured. Maximal standardized uptake value was measured over the thoracic and upper and lower abdominal aortas and correlated with thrombosis and macrophage density on sections from the same sites. Aortic diameters averaged 2.84±1.16 mm. The sensitivity, specificity, and accuracy of CTA for detecting thrombi were 92%, 89%, and 90%, respectively. Plasminogen activator inhibitor-1 and C-reactive protein levels increased with atherosclerosis and thrombosis triggering. Maximal standardized uptake value at baseline was 0.62±0.13, 0.96±0.33 at the middle of feeding, and 1.06±0.38 at the end of feeding. Segments that developed thrombosis had the highest maximal standardized uptake value of 1.32±0.69 (113% increase; P=0.002) and had a 129% increase in macrophage density compared with segments without thrombi (P=0.01). Conclusions— Fluorodeoxyglucose uptake was proportional to the duration of cholesterol feeding and peaked with plaque disruption and thrombosis. CTA was highly accurate in detecting thrombi. Our findings in this animal model of atherosclerotic plaques with high macrophage density showed that CTA/positron emission tomography can be used to identify and localize inflamed plaques and thrombosis. With the currently available technology and nuclear tracers, however, many challenges remain before clinical applications are possible.

Effect of statins on cholesterol crystallization and atherosclerotic plaque stabilization.

Pleiotropic effects of statins have not been fully elucidated. Recently we demonstrated that cholesterol expands when crystallizing and may trigger plaque rupture. The present study evaluated the potential direct effects of statins in altering cholesterol crystallization as a possible mechanism for plaque stabilization independent of cholesterol lowering. Cholesterol powder was dissolved in oil with and without pravastatin, simvastatin, or atorvastatin (10 to 90 mg) and then allowed to crystallize to measure peak volume expansion (ΔVE) in graduated cylinders. Effect of ΔVE on fibrous membrane damage was also evaluated. Human coronary, carotid, and peripheral arterial plaques (65 plaques from 55 patients) were incubated with statin or saline solution using matched plaque segments to evaluate direct effects of statins on preformed crystals. Also, the effect of in vivo use of oral statins on crystal structure was examined by scanning electron microscopy and crystal content in plaques scored from 0 to +3. For all statins, ΔVE decreased significantly in a dose-dependent fashion (0.76 ± 0.1 vs 0 ml at 60 mg, p <0.001). By scanning electron microscopy crystal structure with statins had loss of pointed tip geometries, averting fibrous membrane damage. Cholesterol crystal density was markedly decreased and appeared dissolved in human plaques incubated with statins (+2.1 ± 1.1 vs +1.3 ± 1.0, p = 0.0001). Also, plaques from patients taking oral statins compared to controls had significantly more dissolving crystals (p = 0.03). In conclusion, statins decreased ΔVE by altering cholesterol crystallization and blunting sharp-tipped crystal structure and dissolving cholesterol crystals in human arteries in vivo and in vitro, providing plaque stabilization.

Deletion of the mouse α-calcitonin gene-related peptide gene increases the vulnerability of the heart to ischemia-reperfusion injury

Calcitonin gene-related peptide (CGRP), a potent vasodilator released from capsaicin-sensitive C-fiber and Adelta-fiber sensory nerves, has been suggested to play a beneficial role in myocardial ischemia-reperfusion (I/R) injury. Because most previous studies showing a cardioprotective role of CGRP employed pharmacological experiments, the purpose of this study was to utilize a genetic approach by using mice with a targeted deletion of the alpha-CGRP gene to determine whether this neuropeptide had a modulatory function on the severity of I/R injury. To accomplish this goal, isolated, perfused hearts from alpha-CGRP knockout (KO) and wild-type (WT) mice were subjected to 30 min of ischemia followed by 5, 15, and 30 min of reperfusion. Cardiac functional parameters, including coronary flow rates, left ventricular developed pressure, maximum rates of pressure development, and left ventricular end-diastolic pressure, were measured before and after I/R injury, as were levels of creatine kinase, to assess myocardial damage, and malonaldehyde, to assess oxidative stress. Following I/R injury, cardiac performance was significantly reduced in the hearts from the alpha-CGRP KO mice compared with their WT counterparts. The marked reduction in myocardial function in the alpha-CGRP KO hearts compared with WT hearts after I/R injury was associated with a significant elevation in creatine kinase release into the perfusates and malonaldehyde production in the cardiac tissue. Therefore, these data indicate that, in this in vitro setting, deletion of alpha-CGRP makes the heart more vulnerable to I/R injury, possibly due, at least in part, to increased oxidative stress.

CD44 Targeting Magnetic Glyconanoparticles for Atherosclerotic Plaque Imaging

PurposeThe cell surface adhesion molecule CD44 plays important roles in the initiation and development of atherosclerotic plaques. We aim to develop nanoparticles that can selectively target CD44 for the non-invasive detection of atherosclerotic plaques by magnetic resonance imaging.MethodsMagnetic glyconanoparticles with hyaluronan immobilized on the surface have been prepared. The binding of these nanoparticles with CD44 was evaluated in vitro by enzyme linked immunosorbent assay, flow cytometry and confocal microscopy. In vivo magnetic resonance imaging of plaques was performed on an atherosclerotic rabbit model.ResultsThe magnetic glyconanoparticles can selectively bind CD44. In T2* weighted magnetic resonance images acquired in vivo, significant contrast changes in aorta walls were observed with a very low dose of the magnetic nanoparticles, allowing the detection of atherosclerotic plaques. Furthermore, imaging could be performed without significant delay after probe administration. The selectivity of hyaluronan nanoparticles in plaque imaging was established by several control experiments.ConclusionsMagnetic nanoparticles bearing surface hyaluronan enabled the imaging of atherosclerotic plaques in vivo by magnetic resonance imaging. The low dose of nanoparticles required, the possibility to image without much delay and the high biocompatibility are the advantages of these nanoparticles as contrast agents for plaque imaging.

Thrombostatin, a bradykinin metabolite, reduces platelet activation in a model of arterial wall injury

OBJECTIVE Thrombin activates platelets and contributes to the occlusion of arteries following thrombolytic therapy or angioplasty. Thrombostatin (RPPGF), the angiotensin converting enzyme degradation product of bradykinin, inhibits alpha-thrombin induced platelet activation. We hypothesized that thrombostatin prevents platelet aggregation and adhesion after balloon angioplasty (BA). METHODS Platelet-rich plasma (PRP) was obtained from 22 Beagle dogs before sacrifice and 10% of the PRP was labeled with 111In. Carotid arteries were then removed from each dog and mounted in a dual perfusion chamber and intimal injury was performed with BA. 111In-PRP with or without thrombostatin or aspirin alone was perfused through the arteries for 60 min. During perfusion, platelet volume was measured using a Coulter counter and a laser-light scattering technique. Platelet adhesion to arteries was measured by radioactivity count. RESULTS Arterial injury alone compared to non-injury increased platelet volume in the circuit by 1.4 times (x) (P<0.05) using a Coulter counter or 1.8x (P<0.05) using laser-light scattering and increased platelet adhesion by 2.3x (P<0.01). When compared to BA injury alone, the addition of thrombostatin reduced platelet volume by 1.8x (P<0.03) as measured by Coulter counter or 1.9x (P<0.01) by laser-light scattering and platelet adhesion by 4.2x (P<0.05). Compared to BA injury alone, aspirin reduced platelet volume by 1.2x (P<0.01) as assessed by Coulter counter or 1.5x (P<0.03) using laser-light scattering and platelet adhesion by 1.8x (P<0.02). CONCLUSION Thrombostatin or aspirin independently decreases evidence of platelet activation in the canine carotid artery model of BA injury.

The Impact of Circulating Cholesterol Crystals on Vasomotor Function: Implications for No-Reflow Phenomenon

OBJECTIVES The purpose of this study was to determine if cholesterol crystals can injure the endothelial surface by their jagged edges altering vasoreactivity and contributing to no-reflow after intervention. BACKGROUND After plaque rupture, cholesterol crystals are released into the circulation and flow downstream contacting the arterial wall. METHODS Both carotid arteries from 22 rabbits were placed in a dual perfusion chamber and challenged with norepinephrine followed by acetylcholine and nitroprusside. Arterial diameters were measured before and after exposure to cholesterol crystals or microspheres and compared with diameters of normal control arteries. Arteries were examined by light, confocal, atomic force and scanning electron microscopy. RESULTS Pre-exposure mean arterial diameter was 2.33 ± 0.27 mm. With baseline norepinephrine there was vasoconstriction of 0.82 ± 0.19 mm, 0.79 ± 0.18 mm, and 0.83 ± 0.16 mm in lumen diameter in the crystal, microsphere, and control groups, respectively. After cholesterol crystals or microspheres, vasoconstriction was significantly less for cholesterol crystals but not for microspheres (0.71 ± 0.28 mm and 0.81 ± 0.15 mm; p < 0.02 and p = 0.68). After acetylcholine in the same artery, there was significantly less dilation before versus after crystals (0.49 ± 0.24 mm vs. 0.38 ± 0.22 mm, p = 0.04) but not with microspheres or in the control group. There was no significant difference after nitroprusside in any group, suggesting endothelial injury. By scanning electron microscopy, cholesterol crystals were found embedded in the intima with endothelial cell tears whereas the microsphere treatment and control groups had minimal or no injury (93% vs. 31% vs. 14%; p < 0.01). By atomic force microscopy, surface roughness was significantly greater with cholesterol crystals compared with microspheres or in control arteries (p < 0.05). CONCLUSIONS Cholesterol crystals damaged the endothelium and reduced vasodilator response, potentially aggravating myocardial ischemia after interventions.

Arterial wall cholesterol content is a predictor of development and severity of arterial thrombosis

Background: It is unclear if total cholesterol content contributes to the severity of cardiovascular events by affecting the amount of thrombosis. This study evaluated relationships between cholesterol levels and the amount of thrombosis in an atherosclerotic rabbit model of plaque disruption and thrombosis.Methods: Three groups of NZW rabbits were used: normal rabbits (Group I, n = 4); atherosclerotic rabbits (Group II, n = 4); and atherosclerotic rabbits with pharmacologically triggered thrombosis (Group III, n = 16). Atherosclerosis was induced by feeding a cholesterol enriched diet and balloon deendothelialization. At post-mortem, platelet-rich thrombus and arterial wall cholesterol were quantified and histology performed by light and electron microscopy.Results: Arterial wall cholesterol was strongly correlated to serum cholesterol in all groups (r = 0.94, p < 0.0001). There was a significant correlation between the thrombus surface area with arterial wall cholesterol in Group III (r = 0.71, p < 0.002). Serum cholesterol, arterial wall cholesterol, and thrombus surface area were all significantly correlated but only arterial wall cholesterol was an independent predictor of thrombosis. A threshold specific for this model was noted for serum and arterial cholesterol levels above which thrombosis consistently occurred.Conclusions: Arterial wall cholesterol was strongly correlated to serum cholesterol and thrombosis severity. Serum cholesterol, arterial wall cholesterol and thrombus surface area were all integrally related.Condensed AbstractA model of plaque disruption and thrombosis was used to demonstrate a correlation between serum and arterial wall cholesterol (r = 0.94; p < 0.0001); arterial wall cholesterol and the amount of thrombosis (surface area; r = 0.71, p < 0.002). A threshold of serum and arterial cholesterol was determined at which thrombosis occurred in this model.