Characterization of vulnerable plaques by multiphoton microscopy

Abstract

Cardiovascular disease is the primary cause of death in the United States; the majority of these deaths are caused by the rupture of vulnerable plaques. An important feature of vulnerable plaques is the thickness of the fibrous cap that covers the necrotic core. A thickness of less than 65 microm has been proposed as a value that renders the plaque prone to rupture. This work shows that multiphoton microscopy (MPM) can image the plaque with microm resolution to a depth deeper than 65 microm. The fibrous cap emits primarily second harmonic generation due to collagen, in contrast to the necrotic core and healthy artery, which emits primarily two-photon excited fluorescence from elastin. This gives a good demarcation of the fibrous cap from underlying layers, facilitating the measurement of the fibrous cap thickness. Based on a measure of the collagen/elastin ratio, plaques were detected with a sensitivity of 65% and specificity of 81%. Furthermore, the technique gives detailed information on the structure of the collagen network in the fibrous cap. This network ultimately determines the mechanical strength of the plaque. A mechanical model based on this information could yield a measure of the propensity of the plaque to rupture.

Multimodal CARS microscopy determination of the impact of diet on macrophage infiltration and lipid accumulation on plaque formation in ApoE-deficient mice

“…In previous work, nonlinear microscopy, using combined SHG and TPEF imaging, was shown to reveal structural collagen and elastin fi bers of the extracellular matrix in the arterial wall, the site of atherosclerotic plaque development ( 4,12,24,25 ). In addition, Wang et al demonstrated that SHG/TPEF/CARS imaging can be used to identify endothelial and smooth muscle cells in the arterial wall ( 26 ).…”

Nonlinear optical microscopy in decoding arterial diseases

“…Megens et al (2008) and Doras et al (2011) investigated the changes in collagen structure inside an artery wall affected by atherosclerosis using reconstructions of multiphoton images as well as optical polarization state analysis. Using excised human aortas, Lilledahl et al (2007) demonstrated combined TPEF and SHG imaging of fibrous caps to predict plaque vulnerability. This work showed en face NLOM imaging of plaque to a depth beyond 65 μm from the lumen face through the fibrous cap in thincapped fibroatheroma and into the lesion core, clearly demonstrating the potential of NLOM to differentiate between vulnerable and stable plaques.…”

“…Maffia et al (2007) reported a threedimensional imaging of the entire structure of an isolated intact ApoE−/− mouse carotid artery, identifying a homed, fluorescently tagged, adoptively transferred lymphocytes. Based on a measure of the collagen to elastin density ratio (or SHG:TPEF intensity ratio), Lilledahl et al (2007) and Smith et al (2009) developed an approach to detect vulnerable plaques based on the assumption of a higher collagen density in atherosclerosis lesions, especially in fibroatheroma, compared to the normal vessel wall. Using histological specimens as a reference, Lilledahl et al (2007) reported a specificity of 81 % in detecting fibrous cap atheroma using collagen to elastin ratios derived from pixel-based image quantification.…”

“…Based on a measure of the collagen to elastin density ratio (or SHG:TPEF intensity ratio), Lilledahl et al (2007) and Smith et al (2009) developed an approach to detect vulnerable plaques based on the assumption of a higher collagen density in atherosclerosis lesions, especially in fibroatheroma, compared to the normal vessel wall. Using histological specimens as a reference, Lilledahl et al (2007) reported a specificity of 81 % in detecting fibrous cap atheroma using collagen to elastin ratios derived from pixel-based image quantification. In contrast, Smith et al (2009) reported an approach based on two-photon spectroscopic data.…”

“…In the context of characterizing atherosclerotic progression, Lilledahl et al (2007) were the first to use a measure of the SHG: TPEF ratio derived from a pixel-density based image quantification to detect atherosclerosis plaques with reasonable sensitivity and specificity. In another study, Doras et al (2011) performed multiphoton image reconstruction and also analyzed the change in optical polarization that occurs in atherosclerotic plaque development.…”