The pathologic features that predict atherosclerotic plaque rupture are large lipid collections, thinning of the fibrous cap, and infiltration of macrophages. Optical Coherence Tomography (OCT) has already been demonstrated to accurately image thin fibrous caps and large lipid cores. We now propose an approach which will detect macrophages in vulnerable plaque with OCT in patients at the time of catheterization. We demonstrate that by labeling plaque-based macrophages with intravenous gold nanoparticles we can detect the presence of macrophages for the first time using phase-sensitive OCT coupled with a stimulating laser. In this approach, optical nanoparticles engulfed by plaque-based macrophages are put into nanometer (nm) motion via thermal expansion and relaxation due to application of a pulsed laser, and this motion detected with a phase-sensitive OCT system. We have also developed novel optical nanoparticles for this approach which are excited by light maximally at a wavelength of 700-800 nm, to prevent laser interaction with competing plaque components such as hemoglobin, lipid, and arterial wall which maximally absorb light at 500-600 nm. Due to their multi-faceted surface, we have coined the term "nanorose" to describe these nanoparticles. To demonstrate cellular imaging of macrophages in intact plaque, we propose the following specific aims:
Specific Aim # 1 - Specificity and sensitivity of nanoparticle (nanorose) uptake. 1.1 We will synthesize and fully characterize nanorose with controlled size, and gold and dextran amounts, to achieve strong NIR absorbance and specific cell (macrophage) uptake. 1.2 We will demonstrate specific uptake of nanorose by macrophages as opposed to endothelial and smooth muscle cells in culture via the use of flow cytometry. 1.3 We will inject nanorose iv into atherosclerotic rabbits, and perform detailed histology with fluorescence microcopy to determine both the minimal detectable dose and the specificity of nanorose uptake in plaque.
Specific Aim # 2 - Ex vivo and in vivo OCT imaging of macrophages in rabbit atherosclerotic aortas with nanorose. 2.1 We will inject nanorose iv into atherosclerotic rabbits, perform OCT imaging of ex vivo aortic tissue, and compare these images with histology via RAM-11 identification of macrophages, and hyperspectral imaging of nanorose. 2.2 We will inject nanorose iv into atherosclerotic rabbits, repeat each of the histological experiments in section 2.1, but with in vivo OCT imaging.
Specific Aim # 3 - Excretion and toxicity of nanorose. 3.1 We will characterize the excretion pathway of nanorose (hepatic vs. renal, organs of distribution, and blood half-life). 3.2 We will demonstrate in rabbits the absence of organ toxicity via blood chemistry for evidence of hepatic and renal damage, eosinophil production, and post-mortem examination. These studies will also be performed in collaboration with the Nanotechnology Characterization Laboratory (see letter of support).
Development of a combined contrast agent - light based approach to identify vulnerable plaques will have a significant impact on the public health. Inasmuch as atherosclerosis remains the leading cause of death in Western society, application of the proposed technique that can image plaques at risk for rupture can reduce the number of deaths from heart attack, stroke and peripheral vascular disease. Successful completion of the proposed research will provide a basis for expanded clinical studies to validate the proposed methodology in a large patient population.