MRI detection of endothelial cell inflammation using targeted superparamagnetic particles of iron oxide (SPIO)
Atherosclerosis is a dynamic, progressive disease arising from the combination of endothelial dysfunction with a compensatory inflammatory response [1]. Inflammation is not only instrumental in the development of atheromatous plaques, but is also a key driver of plaque instability subsequently developing clinically significant symptoms such as stroke and myocardial infarction [1].
Current clinical techniques for imaging atherosclerotic disease include angiography, magnetic resonance angiography (MRA), computed tomography angiography (CTA) or ultrasound which principally gives an accurate assessment of degree of luminal stenosis caused by atheroma. However, none of these techniques offer insight about the biological behaviour of atheromatous plaques—particularly degree of inflammation—which cause significant clinical events such as plaque instability and rupture [2].
MRI (magnetic resonance imaging) has advantages over other imaging modalities in clinical use, as it is non-invasive, does not involve ionising radiation and is able to provide high resolution images of the vessel wall at a sub-millimetre level [3]. In addition, recent development of advanced MR contrast agents that selectively target molecules involved in plaque inflammation, offering the promise of in vivo molecular MR imaging of atherosclerosis [4] and ex vivo MR imaging of vulnerable carotid plaque [5].
By exploiting specific ‘inducible’ molecular targets, or cellular events in diseases, molecular imaging uses targeted imaging agents to generate the image contrast [6]. One of the earliest events in atherosclerosis is the over-expression of adhesion molecules on the activated endothelium following exposure to inflammatory cytokines [7]. These pathophysiologically ‘inducible’ endothelial adhesion molecules, such as VCAM-1 (vascular cell adhesion molecule-1) and P-selectin, have the potential to serve as attractive biomarkers for imaging inflammation in atherosclerosis. VCAM-1 (CD106) is an immunoglobulin superfamily glycoprotein (100–110 kDa) expressed on activated endothelial cells, macrophages and smooth muscle cells [8]. It has a vital role in monocyte recruitment in early stages of atherosclerosis [9]. Its expression is upregulated by activated endothelium under inflammation, unlike resting endothelial cells [8]. The increased level of expression appears to be correlated with the extent of exposure to the atherosclerotic risk factors [8]. P-selectin (CD62P, GMP-140, PADGEM), a single-chain glycoprotein, is an adhesion molecule expressed on the surface of activated endothelial cells, which line the luminal surface of blood vessels, and on activated platelets [10]. It mediates initial leukocyte rolling, preceding leukocyte diapedesis into the atherosclerotic lesions [10, 11]. Similar to VCAM-1, P-selectin is rapidly mobilized to the surface of endothelial cells and platelets in response to stimuli [10]. These features render VCAM-1 and P-selectin an ideal biomarker for functional molecular imaging and targeted therapeutics in early atherosclerosis.
To achieve targeted molecular MR imaging, smarter contrast agents are required to identify the molecules of interest (i.e. endothelial adhesion molecules in atherosclerosis) with high specificity. Specificity can be achieved through conjugation of contrast agents with monoclonal antibodies, or their immunospecific fragments F(ab) or peptides. Recent advances in this area include oxidized low-density lipoprotein (oxLDL)-targeted iron oxide nanoparticles [12] and scavenger receptor-AI-targeted iron oxide nanoparticles [13].
SPIO (superparamagnetic particles of iron oxide) nanoparticles are composed of an iron oxide core coated with a biocompatible polymer with size ranging from 50–300 nm. Various iron-oxide nanoparticle preparations are under clinical trials and their safety profile has been increasingly recognised [14]. Iron-oxide particles have become the favoured contrast agent for a number of reasons. Firstly, they have a known biocompatibility profile with their degradation occurring through normal physiologic iron metabolism pathways [15]. By contrast, gadolinium chelates, the clinically used MRI contrast agent, has been reported to have potential severe long-term toxicity effects, including nephrogenic sclerosing fibrosis in patients with impaired renal function [16]. Secondly, iron-oxide particles have high sensitivity due to iron-oxide’s inherent superparamagnetism. They distort magnetic fields, creating marked contrast effects far exceeding their physical size, thereby strongly enhancing the transverse relaxation times T2 and T2*. Hence, they act as contrast agents for T2 sequence [15]. T2 technique is a relaxation time measurement contributing to the transverse decay of the MR signal that arise from natural interactions at the atomic and molecular levels within the tissue or substance of interest. T2* can be considered an “observed” T2, whereas T2 can be considered the “true” T2 of the tissue being imaged [17].
Iron-oxide nanoparticles have the potential to open up new avenues into molecular MR imaging, in both pre-clinical and clinical studies. Indirect quantitative methods utilising the non-specific uptake of iron-oxide particles by macrophages to identify plaque inflammation, have been reported [18]. The present study is an exploratory qualitative study aimed at visualising and characterising atherosclerosis using targeted SPIO as an MRI probe for detecting inflamed endothelial cells. It is hypothesized that highly specific antibody-conjugated SPIO would enable MRI of inflamed endothelial cells.