Magnetic Resonance Imaging of Endothelial Adhesion Molecules in Mouse Atherosclerosis Using Dual-Targeted Microparticles of Iron Oxide

Abstract

Objective-Microparticles of iron oxide (MPIO) distort magnetic field creating marked contrast effects far exceeding their physical size. We hypothesized that antibody-conjugated MPIO would enable magnetic resonance imaging (MRI) of endothelial cell adhesion molecules in mouse atherosclerosis. Methods and Results-MPIO (4.5 m) were conjugated to monoclonal antibodies against vascular cell adhesion molecule-1 (VCAM-MPIO) or P-selectin (P-selectin-MPIO). In vitro, VCAM-MPIO bound, in dose-dependent manner, to tumor necrosis factor (TNF)-␣ stimulated sEND-1 endothelial cells, as quantified by light microscopy (R 2 ϭ0.94, Pϭ0.03) and by MRI (R 2 ϭ0.98, Pϭ0.01). VCAM-MPIO binding was blocked by preincubation with soluble VCAM-1. To mimic leukocyte binding, MPIO targeting both VCAM-1 and P-selectin were administered in apolipoprotein E Ϫ/Ϫ mice. By light microscopy, dual-targeted MPIO binding to endothelium overlying aortic root atherosclerosis was 5-to 7-fold more than P-selectin-MPIO (PϽ0.05) or VCAM-MPIO (PϽ0.01) alone. Dual-targeted MPIO, injected intravenously in vivo bound aortic root endothelium and were quantifiable by MRI ex vivo (3.5-fold increase versus control; PϽ0.01). MPIO were well-tolerated in vivo, with sequestration in the spleen after 24 hours. Conclusions-Dual-ligand MPIO bound to endothelium over atherosclerosis in vivo, under flow conditions. MPIO may provide a functional MRI probe for detecting endothelial-specific markers in a range of vascular pathologies. Key Words: microparticles of iron oxide Ⅲ atherosclerosis Ⅲ magnetic resonance imaging Ⅲ molecular imaging M agnetic resonance imaging (MRI) has demonstrated substantial utility in phenotyping vascular disease. Using inherent physico-chemical properties that confer particular tissue relaxivities, it has been possible to characterize the vessel wall in atherosclerosis at a submillimeter level. 1,2 However, to capitalize fully on the diagnostic potential of MRI requires imaging at molecular and cellular levels. [3][4][5][6][7][8][9] To achieve this, purpose-built contrast agents are needed that can identify molecules of interest with high specificity, while conveying sufficient contrast to be easily distinguished from unenhanced tissue.Specificity can be achieved through conjugation of contrast agent with monoclonal antibodies or their immunospecific fragments F(ab), peptides, or peptide-mimetics. Previous approaches have included integrin-conjugated gadolinium-rich perfluorocarbon nanoparticles, 10 peptide-conjugated nanoparticles of iron oxide, 11 and fibrin-specific cyclic peptide labeled with gadolinium. 12 Gadolinium-based contrast agents shorten T1 providing positive contrast on T1-weighted images. However, for the quantities that can be delivered to an endothelial monolayer, the contrast effects are relatively modest. By comparison, iron oxide nanoparticles provide greater contrast effects, but require many particles to be delivered to a given voxel. Another potential drawback is that contrast effects are manifest in T 2 *-weighted images...

MRI-determined carotid artery flow velocities and wall shear stress in a mouse model of vulnerable and stable atherosclerotic plaque

“…MRI has been used to study plaque burden in the abdominal aorta of mice [12][13][14][15], and recently, several studies have focused on pre-clinical evaluation of novel targeted contrast agents to identify molecular fingerprints of plaque vulnerability [7,[16][17][18][19][20]. The apoE−/− mouse with tapered cast represents an extremely attractive model of atherosclerosis as it allows for studying plaques with both stable and vulnerable characteristics within one animal and within a single vascular segment.…”

Collagens in the progression and complications of atherosclerosis

“…Plaque macrophages are then imaged using a variety of techniques including micro-single-photon emission computed tomography (micro-SPECT), magnetic resonance imaging (MRI), and near infrared fluorescence (NIRF) microscopy. [82][83][84] NIRF microscopy has also been used to image proteolysis in the plaque by detecting the degradation of fluorescent substrates for the MMPs and cathepsins in atherosclerotic plaques in live mice. 85,86 Investigators have used these methods to study plaque growth in mice, and plaque regression after statin treatment.…”

Molecular MRI enables early and sensitive detection of brain metastases

“…This property has previously been used to track MPIO-labeled tumor cells in mouse models, thus enabling visualization of early metastasis distribution in the brain with sensitivity down to a single metastatic cell loaded with ∼50 pg iron at an imaging field strength of 1.5 T (17). Alternatively, conjugation of MPIO with targeting ligands enables their accumulation at sites where specific molecules are expressed (18)(19)(20)(21)(22)(23)(24)(25). Importantly, the targeted MPIO are not taken up by endothelial cells and do not enter the brain, thus providing an endovascular biomarker for pathology within the brain.…”