Faculty Bibliography

The vulnerability or destabilization of atherosclerotic plaques has been directly linked to plaque composition. Imaging modalities, such as magnetic resonance (MR) imaging, that allow for evaluation of plaque composition at a cellular and molecular level, could further improve the detection of vulnerable plaque and may allow for monitoring the efficacy of antiatherosclerotic therapies. In this review we focus on MR imaging strategies for the detection and evaluation of atherosclerotic plaques and their composition. We highlight recent advancements in the development of MR pulse sequences, computer image analysis, and the use of commercially available MR contrast agents, such as gadopentic acid (Gd-DTPA), for plaque characterization. We also discuss molecular imaging strategies that are currently being used to design specific imaging probes targeted to biochemical and cellular markers of atherosclerotic plaque vulnerability.

PURPOSE OF REVIEW: Decades of literature have unambiguously demonstrated regression and remodeling of atherosclerotic lesions, including advanced plaques. Recent insights into underlying mechanisms are reviewed. RECENT FINDINGS: Factors promoting regression include decreased apolipoprotein B-lipoprotein retention within the arterial wall, efflux of cholesterol and other harmful lipids from plaques, and emigration of lesional foam cells followed by entry of healthy phagocytes that remove necrotic debris and other plaque components. Cellular lipid efflux and foam cell emigration can occur surprisingly rapidly once the plaque milieu is improved. Lipid efflux and foam cell emigration each involve specific molecular mediators, many of which have been identified. Necrotic debris removal can be surprisingly comprehensive, with essentially full disappearance documented in animal models. SUMMARY: The essential prerequisite for regression is robust improvement in plaque milieu, meaning large plasma reductions in atherogenic apolipoprotein B-lipoproteins or brisk enhancements in 'reverse' lipid transport from plaque into liver. Importantly, the processes of regression are consistent with rapid correction of features characteristic of the rupture-prone, vulnerable plaques responsible for acute coronary syndromes. New interventions to lower apolipoprotein B-lipoprotein levels and enhance reverse lipid transport may allow regression to become a widespread clinical goal. Strategies based on recent mechanistic insights may facilitate further therapeutic progress

Epidemiological studies support an association between vascular risk factors, including hypercholesterolemia, and Alzheimer's disease (AD). Recently, there has been much interest in the possibility that hypercholesterolemia might directly promote beta-amyloid (Abeta) production. Indeed, in vitro studies have shown that increasing cellular cholesterol levels enhances Abeta production. However, studies in AD transgenic mouse models have not consistently found that elevated plasma cholesterol leads to increased Abeta production or deposition in vivo. In this study, we determined whether elevated peripheral cholesterol influences Abeta production in mice with a null mutation of the low-density lipoprotein receptor (LDLR). We show that dramatically elevated plasma cholesterol levels, whether induced by high cholesterol, high fat, or high fat/high cholesterol diets, did not affect either levels of brain Abeta40, Abeta42, or APP, or the Abeta42/40 or APP-CTF/APP ratios, nor substantially alter brain cholesterol levels. ApoE protein levels in brain were, however, elevated, in LDLR-/- mice by post-transcriptional mechanisms. Collectively, these studies argue that plasma cholesterol levels do not normally regulate production of brain Abeta

The major protein component in secreted very low density lipoproteins (VLDL) is apoB, and it is established that these particles can reach sizes approaching 100 nm. We previously employed a cell-free system to investigate the nature of the vesicles in which this large cargo exits the endoplasmic reticulum (ER) (Gusarova, V., Brodsky, J. L., and Fisher, E. A. (2003) J. Biol. Chem. 278, 48051-48058). We found that apoB-containing lipoproteins exit the ER as dense lipid-protein complexes regardless of the final sizes of the particles and that further expansion occurs via post-ER lipidation. Here, we focused on maturation in the Golgi apparatus. In three separate approaches, we found that VLDL maturation (as assessed by changes in buoyant density) was associated with conformational changes in apoB. In addition, as the size of VLDL expanded, apoE concentrated in a subclass of Golgi microsomes or Golgi-derived vesicles that co-migrated with apoB-containing microsomes or vesicles, respectively. A relationship between apoB and apoE was further confirmed in co-localization studies by immunoelectron microscopy. These combined results are consistent with previous suggestions that apoE is required for VLDL maturation. To our surprise, however, we observed robust secretion of mature VLDL when apoE synthesis was inhibited in either rat hepatoma cells or apoE(-/-) mouse primary hepatocytes. We conclude that VLDL maturation in the Golgi involves apoB conformational changes and that the expansion of the lipoprotein does not require apoE; rather, the increase in VLDL surface area favors apoE binding.

For more than 50 years, investigators have unsuccessfully tried to recreate in experimental animals the cardiovascular complications of diabetes seen in humans. In particular, accelerated atherosclerosis and dilated cardiomyopathy, the major causes of mortality in patients with diabetes, have been conspicuously absent in many mouse models of the disease. Under the auspices of the NIH, the Animal Models of Diabetic Complications Consortium has worked to address this issue. This effort has focused on the development of mouse models because of the high level of genomic information available and the many well-developed genetic manipulations that may be performed in mice. Importantly, the consortium has also worked to standardize many methods to assess metabolic and cardiovascular end points for measurement of the diabetic state and its macrovascular complications. Finally, for maximum benefits from these animal models in the study of atherosclerosis and of other diabetic complications, the consortium has created a system for sharing both the animal models and the accumulated phenotypic data with the greater scientific community.

Early detection and characterization of atherosclerotic lesions susceptible to sudden rupture and thrombosis may decrease morbidity and mortality. Plaque development has been extensively studied using MRI in animal models of rapidly progressing atherosclerosis. These transgenic mice develop atherosclerotic plaques in the aortic root by 10 weeks of age and throughout the vasculature thereafter. Transplantation of lesion-containing segments of the thoracic aorta into wild-type mice results in nearly total reversal of atherosclerosis, making it possible to study both progression and regression of plaques in this model. MRI permits the non-invasive accurate assessment of atherosclerotic plaque burden and the differentiation between the lipid and fibrous content of individual plaques, thus providing a non-invasive approach to serially monitor the evolution of individual plaques in the mouse models. Emergence of novel contrast agents that target a diverse set of molecules within the plaque are now helping to elucidate the changes at the cellular and molecular levels during plaque progression and regression.

Sudden fibrous cap disruption of 'high-risk' atherosclerotic plaques can trigger the formation of an occlusive thrombus in coronary arteries, causing acute coronary syndromes. High-risk atherosclerotic plaques are characterized by their specific cellular and biological content (in particular, a high density of macrophages), rather than by their impact on the vessel lumen. Early identification of high-risk plaques may be useful for preventing ischemic events. One major hurdle in detecting high-risk atherosclerotic plaques in coronary arteries is the lack of an imaging modality that allows for the identification of atherosclerotic plaque composition with high spatial and temporal resolutions. Here we show that macrophages in atherosclerotic plaques of rabbits can be detected with a clinical X-ray computed tomography (CT) scanner after the intravenous injection of a contrast agent formed of iodinated nanoparticles dispersed with surfactant. This contrast agent may become an important adjunct to the clinical evaluation of coronary arteries with CT.

We investigated the ability of targeted immunomicelles to detect and assess macrophages in atherosclerotic plaque using MRI in vivo. There is a large clinical need for a noninvasive tool to assess atherosclerosis from a molecular and cellular standpoint. Macrophages play a central role in atherosclerosis and are associated with plaques vulnerable to rupture. Therefore, macrophage scavenger receptor (MSR) was chosen as a target for molecular MRI. MSR-targeted immunomicelles, micelles, and gadolinium-diethyltriaminepentaacetic acid (DTPA) were tested in ApoE-/- and WT mice by using in vivo MRI. Confocal laser-scanning microscopy colocalization, macrophage immunostaining and MRI correlation, competitive inhibition, and various other analyses were performed. In vivo MRI revealed that at 24 h postinjection, immunomicelles provided a 79% increase in signal intensity of atherosclerotic aortas in ApoE-/- mice compared with only 34% using untargeted micelles and no enhancement using gadolinium-DTPA. Confocal laser-scanning microscopy revealed colocalization between fluorescent immunomicelles and macrophages in plaques. There was a strong correlation between macrophage content in atherosclerotic plaques and the matched in vivo MRI results as measured by the percent normalized enhancement ratio. Monoclonal antibodies to MSR were able to significantly hinder immunomicelles from providing contrast enhancement of atherosclerotic vessels in vivo. Immunomicelles provided excellent validated in vivo enhancement of atherosclerotic plaques. The enhancement seen is related to the macrophage content of the atherosclerotic vessel areas imaged. Immunomicelles may aid in the detection of high macrophage content associated with plaques vulnerable to rupture.