This Article Abstract Full Text (PDF) All Versions of this Article: 108/18/2270    most recent 01.CIR.0000093185.16083.95v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Winter, P. M. Articles by Wickline, S. A. Search for Related Content PubMed PubMed Citation Articles by Winter, P. M. Articles by Wickline, S. A. Pubmed/NCBI databases Substance via MeSH Related Collections Cardiovascular imaging agents/Techniques Pathophysiology

(Circulation. 2003;108:2270.)
© 2003 American Heart Association, Inc.

Basic Science Reports

Molecular Imaging of Angiogenesis in Early-Stage Atherosclerosis With _vß₃-Integrin–Targeted Nanoparticles

Patrick M. Winter, PhD; Anne M. Morawski, BS; Shelton D. Caruthers, PhD; Ralph W. Fuhrhop; Huiying Zhang, MD; Todd A. Williams, BS; John S. Allen, BS; Elizabeth K. Lacy, BS; J. David Robertson, PhD; Gregory M. Lanza, MD, PhD; Samuel A. Wickline, MD

From the Cardiovascular Magnetic Resonance Laboratories (P.M.W., A.M.M., S.D.C., R.W.F., H.Z., T.A.W., J.S.A., E.K.L., G.M.L., S.A.W.), Department of Medicine, Washington University School of Medicine, St Louis, Mo; Philips Medical Systems (S.D.C.), Best, the Netherlands; and the University of Missouri Research Reactor (J.D.R.), Columbia, Mo.

Correspondence to Samuel A. Wickline, MD, or Gregory M. Lanza, MD, PhD, Washington University School of Medicine, Campus Box 8086, 660 S Euclid Ave, St Louis, MO 63110. E-mail saw{at}howdy.wustl.edu

Received April 18, 2003; revision received July 7, 2003; accepted July 7, 2003.

	Abstract

Top Abstract Introduction Methods Results Discussion References

 
Background— Angiogenesis is a critical feature of plaque development in atherosclerosis and might play a key role in both the initiation and later rupture of plaques that lead to myocardial infarction and stroke. The precursory molecular or cellular events that initiate plaque growth and that ultimately contribute to plaque instability, however, cannot be detected directly with any current diagnostic modality.

Methods and Results— Atherosclerosis was induced in New Zealand White rabbits fed 1% cholesterol for 80 days. _vß₃-Integrin–targeted, paramagnetic nanoparticles were injected intravenously and provided specific detection of the neovasculature within 2 hours by routine magnetic resonance imaging (MRI) at a clinically relevant field strength (1.5 T). Increased angiogenesis was detected as a 47±5% enhancement in MRI signal averaged throughout the abdominal aortic wall among rabbits that received _vß₃-targeted, paramagnetic nanoparticles. Pretreatment of atherosclerotic rabbits with _vß₃-targeted, nonparamagnetic nanoparticles competitively blocked specific contrast enhancement of the _vß₃-targeted paramagnetic agent. MRI revealed a pattern of increased _vß₃-integrin distribution within the atherosclerotic wall that was spatially heterogeneous along both transverse and longitudinal planes of the abdominal aorta. Histology and immunohistochemistry confirmed marked proliferation of angiogenic vessels within the aortic adventitia, coincident with prominent, neointimal proliferation among cholesterol-fed, atherosclerotic rabbits in comparison with sparse incidence of neovasculature in the control animals.

Conclusions— This molecular imaging approach might provide a method for defining the burden and evolution of atherosclerosis in susceptible individuals as well as responsiveness of individual patients to antiatherosclerotic therapies.

Key Words: magnetic resonance imaging • atherosclerosis • contrast media • angiogenesis

	Introduction

Top Abstract Introduction Methods Results Discussion References

 
The preponderance of annual mortality in the Western world is caused by atherosclerosis, which presents as cerebrovascular accident or myocardial infarction. In the United States alone, nearly 500 000 cardiac deaths occur each year without warning.¹ Unfortunately, comprehensive evaluation of atherosclerosis generally ensues only after a clinical event is caused by plaque rupture in the later stages of this disease process. Efforts to prevent the development of clinically significant atherosclerotic disease through dietary, lifestyle, and pharmaceutical means abound, but sensitive and specific metrics of early disease are essentially unavailable.

Angiography, the gold standard technique for the detection and characterization of severe atherosclerotic stenosis, has notable limitations inherent to all lumenographic techniques. Other invasive approaches, ie, elastography,² angioscopy,³ thermography,⁴ and intravascular ultrasound,^5,6 are designed to characterize late-stage vascular disease. In contrast, magnetic resonance imaging (MRI) offers a unique, noninvasive opportunity for sensitive detection and characterization of atherosclerotic disease.

The inherently different T₂ relaxation times of atheromatous plaque allow MRI to distinguish cholesterol crystals, collagenous cap, and normal media components.⁷ High-resolution black-blood techniques provide detailed images of carotid and coronary arterial walls and might anatomically resolve the thinning fibrous caps of vulnerable plaques.⁸ Delayed⁹ and dynamic¹⁰ contrast-enhanced MRI can provide an index of vascularity in patients with late carotid disease. These measurements focus on later stages of plaque development that portend or indicate plaque rupture.

In this report, we describe an in vivo, molecular diagnostic imaging approach for quantifying very early plaque development. Angiogenic biomarkers are induced in widespread vascular territories in response to cholesterol feeding, which results in early and critical expansion of the vasa vasorum to support plaque growth.¹¹ Integrins, such as _vß₃, are associated with angiogenesis,¹² and atherosclerotic lesions are highly vascular compared with normal vessel tissues.¹¹ Accordingly, we proposed and developed a paramagnetic nanoparticle contrast agent targeted specifically to _vß₃-integrins to permit noninvasive molecular imaging of plaque-associated angiogenesis.

	Methods

Top Abstract Introduction Methods Results Discussion References

 
Targeted Nanoparticles
For in vivo imaging of sparse molecular epitopes of angiogenesis associated with atherosclerosis, we formulated paramagnetic nanoparticles targeted to _vß₃-integrins. Methods developed in our laboratories were used to prepare perfluorocarbon (perfluorooctylbromide) emulsions encapsulated by a lipid-surfactant monolayer.^13,14 Approximately 90 000 paramagnetic chelates (gadolinium–diethylenetriamine pentaacetic acid-bis-olcate [Gd-DTPA-BOA]) were formulated onto the surface of each particle, as previously described.¹⁵ Paramagnetic nanoparticles were directed to the _vß₃-integrin with a peptidomimetic vitronectin antagonist (US patent No. 6,130,231). The Arg-Gly-Asp mimetic was coupled at a 1:1 molar ratio to malcimidophenyl-butyramide-polyethylene glycol₂₀₀₀-phosphatidylethanolamine resuspended from a dry lipid film in 3 mL of N₂-purged, 6 mmol/L EDTA by water-bath sonication for 30 minutes at 37°C to 40°C. This ligand premix was added to the remaining surfactant components, perfluorooctylbromide, and water for emulsification.¹⁵ Nontargeted, paramagnetic particles were prepared by excluding the targeting ligand, and _vß₃-targeted, nonparamagnetic nanoparticles were produced by omitting the lipophilic Gd³⁺ chelate. The nominal sizes for each formulation were measured with a submicron particle analyzer (Malvern Zetasizer, Malvern Instruments).

The gadolinium concentrations were quantified by neutron activation analysis conducted at the Research Reactor facility at the University of Missouri. In brief, samples were placed in specialized plastic cuvettes that contained no interfering metals, weighed wet, lyophilized, and reweighed. The mass of gadolinium was determined by measuring the 361-keV -rays from the ß-decay of ¹⁶¹Gd (t_1/2=3.66 minutes) produced through neutron capture on ¹⁶⁰Gd. Individual samples and standards were irradiated in a thermal neutron flux of 5x10¹³ n · cm^-2 · s^-1 for 7 seconds, allowed to decay for 30 seconds, and counted on a high-resolution -ray spectrometer for 300 seconds. The minimum detectable amount of gadolinium with this method is reported to be several nanograms.¹⁶

The actual T₁ and T₂ relaxivities (r₁ and r₂, respectively) of the particle formulations were determined with the use of standard inversion recovery and multiecho pulse sequences¹³ applied to pure samples (nanoparticles present at 59 nmol/L). Samples were placed in a quadrature birdcage coil and imaged with a clinical 1.5-T system (Philips NT Intera CV, Philips Medical Systems).

Experimental Design
Male New Zealand White rabbits (Charles River Laboratories, Cambridge, Mass) were fed either 1% cholesterol (n=16) or standard rabbit chow (n=4) for 80 days. Cholesterol-fed rabbits were divided into 3 groups, each of which received 1 of the following: (1) _vß₃-targeted, paramagnetic nanoparticles (n=6); (2) nontargeted, paramagnetic nanoparticles (n=4); or (3) pretreatment with _vß₃-targeted, nonparamagnetic nanoparticles 2 hours before delivery of _vß₃-targeted, paramagnetic nanoparticles (n=6). The last group was designed to demonstrate the specificity of the _vß₃-integrin targeting by in vivo competitive blocking of the binding of targeted, paramagnetic nanoparticles. All control diet animals received _vß₃-targeted, paramagnetic nanoparticles (n=4).

After baseline MRI, all contrast agents were injected peripherally into the ear vein at a dose of 0.5 mL/kg body weight, and contrast enhancement was followed for 2 hours. Nonspecific delayed enhancement between control and fat-fed rabbits was quantified 20 minutes after injection of 0.1 mmol/kg gadolinium-DTPA (Magnevist, Berlex Laboratories, Inc). After imaging, aortas were extracted for histologic assessment, as described later. The experimental protocol was approved by the Animal Studies Committee of the Washington University School of Medicine.

MRI Protocol
A 1.5 T clinical magnet (NT Intera CV, Philips Medical Systems) was used, along with a quadrature birdcage radiofrequency receive coil to image the abdominal aorta in vivo before and at 15, 60, and 120 minutes after treatment with paramagnetic nanoparticles. Multislice, T₁-weighted, spin-echo, fat-suppressed, black-blood imaging of the aorta was performed from the renal arteries to the diaphragm (repetition time=380 ms, echo time=11 ms, 250x250-µm in-plane resolution, 5-mm slice thickness, number of signals averaged=8). To null the blood signal, sliding radiofrequency saturation bands were placed proximal and distal to the region of image acquisition and moved with the selected imaging plane.

Image Analysis
We developed an objective and quantitative method for calculating contrast enhancement within the aortic wall for each image slice, based on a customized, semiautomated segmentation program.¹⁷ The aortic lumen was isolated in each 2-dimensional image through the use of a seeded region-growing algorithm. The segmented lumen was dilated 5 pixels to encompass the entire vessel wall. Further thresholding was used to eliminate background pixels, leaving a binary mask of the aortic wall for each slice and time-point combination. Signal intensity was normalized to the signal from a fiduciary marker (a gadolinium-DTPA/saline solution in a test tube phantom) that was placed within the field of view. The signal enhancement of the aortic wall and adjacent muscle was calculated slice by slice with respect to the baseline images. General linear modeling with Duncan’s multiple-range testing of group differences (SAS, Inc) was used to determine the significance of differences in MRI signals (P<0.05).

Histology
Routine hematoxylin/eosin staining was performed on formalin-fixed, paraffin-embedded sections (4 µm) of aortas. Expression of _vß₃-integrin in the aortic wall was confirmed by immunohistochemistry of formalin-fixed sections with use of a specific primary antibody (LM609, Chemicon International, Inc) and a secondary antibody developed with an aminoethylcarbazole substrate kit. Platelet and endothelial cell adhesion molecule (PECAM) was stained similarly with CD31 primary antibody (Chemicon International, Inc). Images of neovasculature were digitized under high power (600x) with a Nikon microscope and Nikon DXM1200 camera.

	Results

Top Abstract Introduction Methods Results Discussion References

 
Nanoparticle Characteristics
Nanoparticles were 273 nm in diameter (polydispersity=0.15) and contained 6.17 mmol/L gadolinium, or 94 200 Gd³⁺ atoms per particle. The ionic-based r₁ and r₂ values for paramagnetic nanoparticles expressed per millimole Gd³⁺ per liter were 17.7±0.2 and 25.3±0.6 (s · mmol/L)^-1, respectively, and the particle-based relaxivities that reflect the signal effects achievable per individual binding site (ie, particle) were 1 670 000±100 000 and 2 380 000±120 000 (sec · mmol particle/L)^-1 for r₁ and r₂, respectively. Targeted and nontargeted particles exhibited equivalent paramagnetic properties.

_vß₃-Integrin Within Atherosclerotic Aortic Wall
Figure 1 (top) shows the imaged portion of the aorta in longitudinal profile for a selected animal and transverse slices (bottom) at baseline (Pre) and 120 minutes after (Post) treatment with targeted nanoparticles and the segmented aortic wall mask (Segmented) used to quantify the contrast enhancement (Enhancement) for this slice. The signal in the aortic wall increased after contrast injection (labeled Post and Enhancement), indicating the presence of targeted nanoparticles bound to _vß₃-integrin epitopes on the neovasculature. Furthermore, note that the aortic blood pool background was not confounded by signal originating from circulating nanoparticles, which enabled rapid detection of contrast enhancement in the aortic wall. Aortic wall contrast enhancement was variable along the circumference and length of the aorta, as illustrated by the color-coded signal enhancement maps (Figure 2) and longitudinally along the aorta for 3 selected rabbits (Figure 3). However, greater signal enhancement was observed in the cholesterol-fed, targeted rabbits for practically every aortic segment.

View larger version (50K): [in this window] [in a new window]   Figure 1. In vivo spin-echo image reformatted to display long axis of aorta from renal arteries to diaphragm of 1 cholesterol-fed rabbit (top) and at single transverse level (bottom) before (Pre) and after (Post) treatment, after semiautomated segmentation (Segmented, grayish ring; see text for description), and with color-coded signal enhancement (Enhancement) above baseline (in percent).

View larger version (37K): [in this window] [in a new window]   Figure 2. Percent enhancement maps (false-colored from blue to red) from individual aortic segments at renal artery (A), mid-aorta (B), and diaphragm (C) 2 hours after treatment in cholesterol-fed rabbit given vß3-targeted nanoparticles.

View larger version (23K): [in this window] [in a new window]   Figure 3. Spatial variation of contrast enhancement after treatment showing longitudinal signal enhancement from renal arteries to diaphragm for 1 representative animal selected from 3 experimental groups: cholesterol (Chol)-fed/targeted, control diet (CNT), and cholesterol-fed/nontargeted.

Histology of cholesterol-fed rabbit aortas revealed mild intimal thickening after 80 days, which was not appreciated in animals that received the control diet (Figure 4). Estimation of lumen area and wall thickness by MRI was similar for all treatment groups, reflecting the early stage of vascular disease in these animals. Immunohistochemistry revealed expansion of the aortic vasa vasorum (PECAM-positive) among atherosclerotic rabbits in comparison with the controls. Angiogenic vessels, delineated by colocalized staining of _vß₃-integrin and PECAM, formed a subpopulation of the total expanded vasculature in the hyperlipidemic aortas, predominately localized to the adventitia-media interface (Figure 4). In control rabbits, a sparse neovasculature was noted in the same regions.

View larger version (118K): [in this window] [in a new window]   Figure 4. Immunohistochemistry of vß3-integrin in aorta sections from cholesterol-fed (A, C, D) and control diet (B) rabbits. Thickened intima (I) is observed in cholesterol-fed animal (A, 200x) but not in control animal (B, 200x). Immunostaining of neovascular vß3-integrin (C) and PECAM (D) in aorta from cholesterol-fed animal in A at 600x. Solid arrows delineate vß3-integrin expression, and open arrows mark PECAM expression at interface between media (M) and adventitia (Av). Prevalence of vß3-integrin staining in control rabbit (solid arrows in B) is far less prominent than for cholesterol-fed animal (A).

Quantification of the aortic signal enhancement among cholesterol-fed rabbits that received _vß₃-targeted, paramagnetic nanoparticles showed a 26±4% and 47±5% increase over baseline at 15 and 120 minutes, respectively, when averaged across the expanse of the aorta for each rabbit (Figure 5, bottom). In cholesterol-fed rabbits that received nontargeted nanoparticles, the aortic wall was enhanced by 19±1% within 15 minutes, but it stabilized from 60 to 120 minutes at 26±1%, which represents about half of the signal augmentation observed for the specific _vß₃-targeted enhancement. Competitive blockade of angiogenic _vß₃-integrins with targeted, nonparamagnetic nanoparticles reduced the signal enhancement of _vß₃-targeted, paramagnetic nanoparticles by at least 50%. In control diet rabbits, aortic wall enhancement from _vß₃-targeted nanoparticles paralleled the contrast effect noted among the nontargeted, cholesterol-fed animals. Thus, the overall average signal enhancement in the aortic wall for cholesterol-fed animals was approximately twice that for control diet animals at 120 minutes. The signal enhancement observed in the adjacent skeletal muscle (Figure 5, top) owing to nanoparticles in all treatment groups was negligible relative to that exhibited by the aortic wall.

View larger version (23K): [in this window] [in a new window]   Figure 5. Quantitative analysis of MRI signal enhancement (percent) from aorta (bottom) and skeletal muscle (top) after treatment with vß3-targeted or nontargeted nanoparticles in cholesterol-fed or control diet groups. *P<0.05 for cholesterol-fed/targeted vs all other groups.

In contradistinction to the specific MRI enhancement observed with _vß₃-targeted nanoparticles, delayed contrast enhancement of the vessel wall with gadolinium-DTPA revealed no significant difference between cholesterol-fed (91±7%) and control diet (87±3%) animals. Although nonspecific contrast agents have been used to distinguish fibrous, necrotic, and calcified tissues associated with complex plaques,⁹ they were insensitive to the subtle vascular changes associated with the early development of atherosclerosis.

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
Plaque angiogenesis and expansion of the vasa vasorum are critical processes for the initiation of vascular lesions and the progression of early atherosclerotic inflammatory events driven by cholesterol feeding.^18,19 These data indicate that targeted, paramagnetic nanoparticles might be used for molecular imaging of inducible _vß₃-integrins throughout the vascular wall and reaffirm that early atherosclerosis evolves as a diffuse process in response to elevated cholesterol. Targeted characterization of _vß₃-integrin expression revealed that the early atherosclerotic process manifests considerable heterogeneity within individual aortic segments. Variation in the gross severity of atherosclerosis from the diaphragm to the renal arteries generally corresponded to the calculated MRI signal enhancement and the magnitude of neovascular proliferation observed histologically. These findings are in agreement with an analogous heterogeneous distribution of aortic foam cells in cholesterol-fed rabbits illustrated with cold-spot imaging of susceptibility artifacts created by the cellular uptake of nontargeted, iron oxide particles.²⁰ Although the precise role of _vß₃-integrin expression in the expansion of the vasa vasorum was not elucidated by these data, its detection by noninvasive imaging methods could serve as a useful biochemical signature of the early atherosclerotic condition that might demarcate lesion-prone sites of focal plaque growth.

In the present study, we confirmed the in vivo binding specificity of _vß₃-targeted nanoparticles with in vivo competition studies. Pretreatment of cholesterol-fed rabbits with _vß₃-targeted, nonparamagnetic nanoparticles blocked the increased signal enhancement from _vß₃-targeted, paramagnetic nanoparticles to a level slightly below that obtained for nontargeted, paramagnetic nanoparticles. Passive contrast enhancement among atherosclerotic animals injected with nontargeted nanoparticles might reflect neovasculature-related leakage within the aorta. In rabbits fed a control diet and given _vß₃-targeted, paramagnetic nanoparticles, signal enhancement was similar to the nonspecific background level. This low-contrast effect might be partially attributable to a combination of specific targeting and passive leakage associated with sparse neovascular vessels observed histologically (Figure 4).

In the present study conducted at 1.5 T, molecular imaging of angiogenesis in hyperlipidemic rabbits was appreciated within the aortic wall but could not be spatially resolved to a specific vascular layer (eg, adventitia). With the anticipated clinical adoption of 3 T and higher-field MR scanners, greater image resolution is expected, which should improve noninvasive neovascular localization and facilitate clinical risk assessments.

We have previously reported that this nanoparticle emulsion allows sensitive molecular imaging of abundant vascular epitopes (eg, fibrin) with clinical ultrasound and MRI systems.^13,14,21 In this study, we have shown that paramagnetic nanoparticles can provide sensitive in vivo detection and characterization of molecular epitopes of lesser abundance, such as _vß₃-integrin, in early atherosclerosis. As powerful serum markers for the generalized state of inflammation become widely available for clinical use (eg, C-reactive protein), high-resolution in vivo molecular imaging methods could provide adjunctive data to precisely localize and quantify early or unstable vascular lesions that can be triaged rationally to medical treatment and then followed up serially for therapeutic response. Furthermore, as recently reported,²² this ligand-targeted nanoparticle system might carry a variety of drugs in its lipid membrane, rendering it an intriguing candidate for simultaneous molecular imaging and site-directed drug delivery for early atherosclerotic disease.

	Acknowledgments

 
We acknowledge grant support from the National Institutes of Health (HL-42950, HL-59865, and NO1-C0–07121), the American Heart Association, the Barnes-Jewish Research Foundation, the Edith and Alan Wolff Charitable Trust, and Philips Medical Systems. We appreciate the use of the vitronectin antagonist provided by Bristol-Myers Squibb Medical Imaging (Billerica, Mass).

	Footnotes

 
Dr Caruthers is an employee of Philips Medical Systems. Drs Lanza and Wickline are cofounders, board members, and equity holders of Kereos, Inc. R.W. Fuhrhop and J.S. Allen are consultants to Kereos, Inc.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Zheng Z-J, Croft JB, Giles WH, et al. Sudden cardiac death in the United States, 1989 to 1998. Circulation. 2001; 104: 2158–2163.[Abstract/Free Full Text]

2. Carlier SG, de Korte CL, Brusseau E, et al. Imaging of atherosclerosis: elastography. J Cardiovasc Risk. 2002; 9: 237–245.[CrossRef][Medline] [Order article via Infotrieve]

3. Unno N, Mitsuoka H, Takei Y, et al. Virtual angioscopy using 3-dimensional rotational digital subtraction angiography for endovascular assessment. J Endovasc Ther. 2002; 9: 529–534.[CrossRef][Medline] [Order article via Infotrieve]

4. Madjid M, Naghavi M, Malik BA, et al. Thermal detection of vulnerable plaque. Am J Cardiol. 2002; 90: 36L–39L.[CrossRef][Medline] [Order article via Infotrieve]

5. Demos S, Alkan-Onyuksel H, Kane B, et al. In vivo targeting of acoustically reflective liposomes for intravascular and transvascular ultrasonic enhancement. J Am Coll Cardiol. 1999; 33: 867–875.[Abstract/Free Full Text]

6. Tardif JC, Gregoire J, Schwartz L, et al. Effects of AGI-1067 and probucol after percutaneous coronary interventions. Circulation. 2003; 107: 552–558.[Abstract/Free Full Text]

7. Kramer CM. Magnetic resonance imaging to identify the high-risk plaque. Am J Cardiol. 2002; 90: 15L–17L.[CrossRef][Medline] [Order article via Infotrieve]

8. Fayad ZA, Fuster V, Nikolaou K, et al. Computed tomography and magnetic resonance imaging for noninvasive coronary angiography and plaque imaging: current and potential future concepts. Circulation. 2002; 106: 2026–2034.[Free Full Text]

9. Yuan C, Kerwin WS, Ferguson MS, et al. Contrast-enhanced high resolution MRI for atherosclerotic carotid artery tissue characterization. J Magn Reson Imaging. 2002; 15: 62–67.[CrossRef][Medline] [Order article via Infotrieve]

10. Kerwin W, Hooker A, Spilker M, et al. Quantitative magnetic resonance imaging analysis of neovasculature volume in carotid atherosclerotic plaque. Circulation. 2003; 107: 851–856.[Abstract/Free Full Text]

11. O’Brien ER, Garvin MR, Dev R, et al. Angiogenesis in human coronary atherosclerotic plaque. Am J Pathol. 1994; 145: 883–894.[Abstract]

12. Brooks PC, Montgomery AMP, Rosenfeld M, et al. Integrin alpha v beta 3 antagonists promote tumor regression by inducing apoptosis of angiogenic blood vessels. Cell. 1994; 79: 1157–1164.[CrossRef][Medline] [Order article via Infotrieve]

13. Flacke S, Fischer S, Scott MJ, et al. Novel MRI contrast agent for molecular imaging of fibrin: implications for detecting vulnerable plaques. Circulation. 2001; 104: 1280–1285.[Abstract/Free Full Text]

14. Lanza G, Wallace K, Scott M, et al. A novel site-targeted ultrasonic contrast agent with broad biomedical application. Circulation. 1996; 94: 3334–3340.[Abstract/Free Full Text]

15. Winter PM, Caruthers SD, Yu X, et al. Improved molecular imaging contrast agent for detection of human thrombus. Magn Reson Med. 2003; 50: 411–416.[CrossRef][Medline] [Order article via Infotrieve]

16. Landsberger S. Delayed instrumental neutron activation analysis. In: Alfassi ZB, ed. Chemical Analysis by Nuclear Methods. New York, NY: Wiley; 1994: 122–140.

17. Nappi J, Dachman A, MacEneaney P, et al. Automated knowledge-guided segmentation of colonic walls for computerized detection of polyps in CT colonography. J Comput Assist Tomogr. 2002; 26: 493–504.[CrossRef][Medline] [Order article via Infotrieve]

18. Moulton KS, Heller E, Konerding MA, et al. Angiogenesis inhibitors endostatin or TNP-470 reduce intimal neovascularization and plaque growth in apolipoprotein E–deficient mice. Circulation. 1999; 99: 1653–1655.[Free Full Text]

19. Wilson SH, Herrmann J, Lerman LO, et al. Simvastatin preserves the structure of coronary adventitial vasa vasorum in experimental hypercholesterolemia independent of lipid lowering. Circulation. 2002; 105: 415–418.[Abstract/Free Full Text]

20. Ruehm SG, Corot C, Vogt P, et al. Magnetic resonance imaging of atherosclerotic plaque with ultrasmall superparamagnetic particles of iron oxide in hyperlipidemic rabbits. Circulation. 2001; 103: 415–422.[Abstract/Free Full Text]

21. Yu X, Song S-K, Chen J, et al. High-resolution MRI characterization of human thrombus using a novel fibrin-targeted paramagnetic nanoparticle contrast agent. Magn Reson Med. 2000; 44: 867–872.[CrossRef][Medline] [Order article via Infotrieve]

22. Lanza GM, Yu X, Winter PM, et al. Targeted antiproliferative drug delivery to vascular smooth muscle cells with an MRI nanoparticle contrast agent: implications for rational therapy of restenosis. Circulation. 2002; 106: 2842–2847.[Abstract/Free Full Text]

This article has been cited by other articles:

				H.-f. Zhou, H. W. Chan, S. A. Wickline, G. M. Lanza, and C. T. N. Pham {alpha}v{beta}3-Targeted nanotherapy suppresses inflammatory arthritis in mice FASEB J, September 1, 2009; 23(9): 2978 - 2985. [Abstract] [Full Text] [PDF]

				M. Sirol, P. R. Moreno, K.-R. Purushothaman, E. Vucic, V. Amirbekian, H.-J. Weinmann, P. Muntner, V. Fuster, and Z. A. Fayad Increased Neovascularization in Advanced Lipid-Rich Atherosclerotic Lesions Detected by Gadofluorine-M-Enhanced MRI: Implications for Plaque Vulnerability Circ Cardiovasc Imaging, September 1, 2009; 2(5): 391 - 396. [Abstract] [Full Text] [PDF]

				A. Saraste, S. G. Nekolla, and M. Schwaiger Cardiovascular molecular imaging: an overview Cardiovasc Res, September 1, 2009; 83(4): 643 - 652. [Abstract] [Full Text] [PDF]

				R. P. Choudhury and E. A. Fisher Molecular Imaging in Atherosclerosis, Thrombosis, and Vascular Inflammation Arterioscler Thromb Vasc Biol, July 1, 2009; 29(7): 983 - 991. [Abstract] [Full Text] [PDF]

				I. Laitinen, A. Saraste, E. Weidl, T. Poethko, A. W. Weber, S. G. Nekolla, P. Leppanen, S. Yla-Herttuala, G. Holzlwimmer, A. Walch, et al. Evaluation of {alpha}v{beta}3 Integrin-Targeted Positron Emission Tomography Tracer 18F-Galacto-RGD for Imaging of Vascular Inflammation in Atherosclerotic Mice Circ Cardiovasc Imaging, July 1, 2009; 2(4): 331 - 338. [Abstract] [Full Text] [PDF]

				F. A. Jaffer, P. Libby, and R. Weissleder Optical and Multimodality Molecular Imaging: Insights Into Atherosclerosis Arterioscler Thromb Vasc Biol, July 1, 2009; 29(7): 1017 - 1024. [Abstract] [Full Text] [PDF]

				M. Oostendorp, M. J. Post, and W. H. Backes Vessel Growth and Function: Depiction with Contrast-enhanced MR Imaging Radiology, May 1, 2009; 251(2): 317 - 335. [Abstract] [Full Text] [PDF]

				V. Amirbekian, J. G. S. Aguinaldo, S. Amirbekian, F. Hyafil, E. Vucic, M. Sirol, D. B. Weinreb, S. Le Greneur, E. Lancelot, C. Corot, et al. Atherosclerosis and Matrix Metalloproteinases: Experimental Molecular MR Imaging in Vivo Radiology, May 1, 2009; 251(2): 429 - 438. [Abstract] [Full Text] [PDF]

				M. Nahrendorf, D. E. Sosnovik, B. A. French, F. K. Swirski, F. Bengel, M. M. Sadeghi, J. R. Lindner, J. C. Wu, D. L. Kraitchman, Z. A. Fayad, et al. Multimodality Cardiovascular Molecular Imaging, Part II Circ Cardiovasc Imaging, January 1, 2009; 2(1): 56 - 70. [Full Text] [PDF]

				A. L. Doiron, K. Chu, A. Ali, and L. Brannon-Peppas Nanomaterials in Medicine Special Feature Sackler Colloquium: Preparation and initial characterization of biodegradable particles containing gadolinium-DTPA contrast agent for enhanced MRI PNAS, November 11, 2008; 105(45): 17232 - 17237. [Abstract] [Full Text] [PDF]

				P. M. Winter, S. D. Caruthers, H. Zhang, T. A. Williams, S. A. Wickline, and G. M. Lanza Antiangiogenic synergism of integrin-targeted fumagillin nanoparticles and atorvastatin in atherosclerosis. J. Am. Coll. Cardiol. Img., September 1, 2008; 1(5): 624 - 634. [Abstract] [Full Text] [PDF]

				E. Arbustini and F. I. Gambarin Theranostic strategy against plaque angiogenesis. J. Am. Coll. Cardiol. Img., September 1, 2008; 1(5): 635 - 637. [Full Text] [PDF]

				J. Schwitter Extending the Frontiers of Cardiac Magnetic Resonance Circulation, July 8, 2008; 118(2): 109 - 112. [Full Text] [PDF]

				H. F. Langer, R. Haubner, B. J. Pichler, and M. Gawaz Radionuclide imaging a molecular key to the atherosclerotic plaque. J. Am. Coll. Cardiol., July 1, 2008; 52(1): 1 - 12. [Abstract] [Full Text] [PDF]

				T. H. Marwick and M. Schwaiger The Future of Cardiovascular Imaging in the Diagnosis and Management of Heart Failure, Part 1: Tasks and Tools Circ Cardiovasc Imaging, July 1, 2008; 1(1): 58 - 69. [Full Text] [PDF]

				W. J.M. Mulder and Z. A. Fayad Nanomedicine Captures Cardiovascular Disease Arterioscler Thromb Vasc Biol, May 1, 2008; 28(5): 801 - 802. [Full Text] [PDF]

				C. Burtea, S. Laurent, O. Murariu, D. Rattat, G. Toubeau, A. Verbruggen, D. Vansthertem, L. Vander Elst, and R. N. Muller Molecular imaging of {alpha}v{beta}3 integrin expression in atherosclerotic plaques with a mimetic of RGD peptide grafted to Gd-DTPA Cardiovasc Res, April 1, 2008; 78(1): 148 - 157. [Abstract] [Full Text] [PDF]

				E. Lancelot, V. Amirbekian, I. Brigger, J.-S. Raynaud, S. Ballet, C. David, O. Rousseaux, S. Le Greneur, M. Port, H. R. Lijnen, et al. Evaluation of Matrix Metalloproteinases in Atherosclerosis Using a Novel Noninvasive Imaging Approach Arterioscler Thromb Vasc Biol, March 1, 2008; 28(3): 425 - 432. [Abstract] [Full Text] [PDF]

				L. L. Johnson, L. Schofield, T. Donahay, M. Bouchard, A. Poppas, and R. Haubner Radiolabeled RGD Peptides to Image Angiogenesis in Swine Model of Hibernating Myocardium. J. Am. Coll. Cardiol. Img., January 1, 2008; 1: 500 - 510. [Abstract] [Full Text] [PDF]

				M. A. McAteer, J. E. Schneider, Z. A. Ali, N. Warrick, C. A. Bursill, C. von zur Muhlen, D. R. Greaves, S. Neubauer, K. M. Channon, and R. P. Choudhury Magnetic Resonance Imaging of Endothelial Adhesion Molecules in Mouse Atherosclerosis Using Dual-Targeted Microparticles of Iron Oxide Arterioscler Thromb Vasc Biol, January 1, 2008; 28(1): 77 - 83. [Abstract] [Full Text] [PDF]

				A. K. Bajpai, E. Blaskova, S. B. Pakala, T. Zhao, W. C. Glasgow, J. S. Penn, D. A. Johnson, and G. N. Rao 15(S)-HETE Production in Human Retinal Microvascular Endothelial Cells by Hypoxia: Novel Role for MEK1 in 15(S)-HETE Induced Angiogenesis Invest. Ophthalmol. Vis. Sci., November 1, 2007; 48(11): 4930 - 4938. [Abstract] [Full Text] [PDF]

				F. A. Jaffer, P. Libby, and R. Weissleder Molecular Imaging of Cardiovascular Disease Circulation, August 28, 2007; 116(9): 1052 - 1061. [Full Text] [PDF]

				T. Saam, T. S. Hatsukami, N. Takaya, B. Chu, H. Underhill, W. S. Kerwin, J. Cai, M. S. Ferguson, and C. Yuan The Vulnerable, or High-Risk, Atherosclerotic Plaque: Noninvasive MR Imaging for Characterization and Assessment Radiology, July 1, 2007; 244(1): 64 - 77. [Abstract] [Full Text] [PDF]

				B. Doyle and N. Caplice Plaque Neovascularization and Antiangiogenic Therapy for Atherosclerosis J. Am. Coll. Cardiol., May 29, 2007; 49(21): 2073 - 2080. [Abstract] [Full Text] [PDF]

				K. Srivastava, V. Kundumani-Sridharan, B. Zhang, A. K. Bajpai, and G. N. Rao 15(S)-Hydroxyeicosatetraenoic Acid-Induced Angiogenesis Requires STAT3-Dependent Expression of VEGF Cancer Res., May 1, 2007; 67(9): 4328 - 4336. [Abstract] [Full Text] [PDF]

				X. Yang Nano- and Microparticle-based Imaging of Cardiovascular Interventions: Overview Radiology, May 1, 2007; 243(2): 340 - 347. [Abstract] [Full Text] [PDF]

				D. E. Sosnovik, M. Nahrendorf, and R. Weissleder Molecular Magnetic Resonance Imaging in Cardiovascular Medicine Circulation, April 17, 2007; 115(15): 2076 - 2086. [Full Text] [PDF]

				L. Zhao, J. A. Hall, N. Levenkova, E. Lee, M. K. Middleton, A. M. Zukas, D. J. Rader, J. J. Rux, and E. Pure CD44 Regulates Vascular Gene Expression in a Proatherogenic Environment Arterioscler Thromb Vasc Biol, April 1, 2007; 27(4): 886 - 892. [Abstract] [Full Text] [PDF]

				P. M. Winter, A. M. Neubauer, S. D. Caruthers, T. D. Harris, J. D. Robertson, T. A. Williams, A. H. Schmieder, G. Hu, J. S. Allen, E. K. Lacy, et al. Endothelial {alpha}{nu}{beta}3 Integrin-Targeted Fumagillin Nanoparticles Inhibit Angiogenesis in Atherosclerosis Arterioscler Thromb Vasc Biol, September 1, 2006; 26(9): 2103 - 2109. [Abstract] [Full Text] [PDF]

				R. S. Vasan Biomarkers of Cardiovascular Disease: Molecular Basis and Practical Considerations Circulation, May 16, 2006; 113(19): 2335 - 2362. [Full Text] [PDF]

				P. R. Moreno, K-R. Purushothaman, M. Sirol, A. P. Levy, and V. Fuster Neovascularization in Human Atherosclerosis Circulation, May 9, 2006; 113(18): 2245 - 2252. [Full Text] [PDF]

				R. L. Wilensky, H. K. Song, and V. A. Ferrari Role of magnetic resonance and intravascular magnetic resonance in the detection of vulnerable plaques. J. Am. Coll. Cardiol., April 18, 2006; 47(8 Suppl): C48 - C56. [Abstract] [Full Text] [PDF]

				F. A. Jaffer, P. Libby, and R. Weissleder Molecular and Cellular Imaging of Atherosclerosis: Emerging Applications J. Am. Coll. Cardiol., April 4, 2006; 47(7): 1328 - 1338. [Abstract] [Full Text] [PDF]

				S. A. Wickline, A. M. Neubauer, P. Winter, S. Caruthers, and G. Lanza Applications of Nanotechnology to Atherosclerosis, Thrombosis, and Vascular Biology Arterioscler Thromb Vasc Biol, March 1, 2006; 26(3): 435 - 441. [Abstract] [Full Text] [PDF]

				F. Hyafil, J.-P. Laissy, M. Mazighi, D. Tchetche, L. Louedec, H. Adle-Biassette, S. Chillon, D. Henin, M.-P. Jacob, D. Letourneur, et al. Ferumoxtran-10-Enhanced MRI of the Hypercholesterolemic Rabbit Aorta: Relationship Between Signal Loss and Macrophage Infiltration Arterioscler Thromb Vasc Biol, January 1, 2006; 26(1): 176 - 181. [Abstract] [Full Text] [PDF]

				H.-J. Wester and H. Kessler Molecular Targeting with Peptides or Peptide-Polymer Conjugates: Just a Question of Size? J. Nucl. Med., December 1, 2005; 46(12): 1940 - 1945. [Full Text] [PDF]

				K. A. Kelly, J. R. Allport, A. Tsourkas, V. R. Shinde-Patil, L. Josephson, and R. Weissleder Detection of Vascular Adhesion Molecule-1 Expression Using a Novel Multimodal Nanoparticle Circ. Res., February 18, 2005; 96(3): 327 - 336. [Abstract] [Full Text] [PDF]

				F. A. Jaffer and R. Weissleder Molecular Imaging in the Clinical Arena JAMA, February 16, 2005; 293(7): 855 - 862. [Abstract] [Full Text] [PDF]

				J. C. Miller, H. H. Pien, D. Sahani, A. G. Sorensen, and J. H. Thrall Imaging Angiogenesis: Applications and Potential for Drug Development J Natl Cancer Inst, February 2, 2005; 97(3): 172 - 187. [Abstract] [Full Text] [PDF]

				C. M. Matter, P. K. Schuler, P. Alessi, P. Meier, R. Ricci, D. Zhang, C. Halin, P. Castellani, L. Zardi, C. K. Hofer, et al. Molecular Imaging of Atherosclerotic Plaques Using a Human Antibody Against the Extra-Domain B of Fibronectin Circ. Res., December 10, 2004; 95(12): 1225 - 1233. [Abstract] [Full Text] [PDF]

				P. R. Moreno and V. Fuster The year in atherothrombosis J. Am. Coll. Cardiol., December 7, 2004; 44(11): 2099 - 2110. [Full Text] [PDF]

				M. A. McAteer, J. E. Schneider, K. Clarke, S. Neubauer, K. M. Channon, and R. P. Choudhury Quantification and 3D Reconstruction of Atherosclerotic Plaque Components in Apolipoprotein E Knockout Mice Using Ex Vivo High-Resolution MRI Arterioscler Thromb Vasc Biol, December 1, 2004; 24(12): 2384 - 2390. [Abstract] [Full Text] [PDF]

				R. J. Gibbons and P. A. Araoz The year in cardiac imaging J. Am. Coll. Cardiol., November 16, 2004; 44(10): 1937 - 1944. [Full Text] [PDF]

				S. A. Wickline Plaque characterization: surrogate markers or the real thing? J. Am. Coll. Cardiol., April 7, 2004; 43(7): 1185 - 1187. [Full Text] [PDF]

				F. A. Jaffer and R. Weissleder Seeing Within: Molecular Imaging of the Cardiovascular System Circ. Res., March 5, 2004; 94(4): 433 - 445. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) All Versions of this Article: 108/18/2270    most recent 01.CIR.0000093185.16083.95v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Winter, P. M. Articles by Wickline, S. A. Search for Related Content PubMed PubMed Citation Articles by Winter, P. M. Articles by Wickline, S. A. Pubmed/NCBI databases Substance via MeSH Related Collections Cardiovascular imaging agents/Techniques Pathophysiology