Near-infrared spectroscopic characterization of human advanced atherosclerotic plaques
Jing Wang, PhD*,
Yong-Jian Geng, MD, PhD ,
Bujin Guo, PhD*,
Tomas Klima, MD*,
Birendra N. Lal, MD,
James T. Willerson, MD* and
Ward Casscells, MD*,*
* Vascular Cell Biology Laboratory, Texas Heart Institute, Houston, Texas, USA
Division of Cardiology, Department of Internal Medicine, University of Texas Medical School at Houston, Houston, Texas, USA
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Figure 1 Spectroscopic images of a thin (25 µm), freeze-dried section of vulnerable human carotid atherosclerotic plaque in the near-infrared radiation region. (A) Regular light image of a plaque section. (B to D) Color near-infrared images illustrating the pattern of spectroscopic alteration. Major biochemical components are visualized by wavelength. (B) At 2,225 to 2,550 nm, the contents of cholesterol and cholesterol esters are shown from high to low in red to orange to yellow to green to blue. (C) At 2,200 to 2,225 nm, the contents of protein components (mainly collagen and other matrix proteins) are shown. (D) At 1,720 nm, lipid-rich regions (red) tend to co-localize with regions not in cholesterol (as in B).
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Figure 2 The near-infrared spectra of normal fresh (unfixed) canine aorta and canine normal aortic tissue overlaid with various amounts of lipid deposits (from carotid plaques in the same animal). (A) Overview of second-derivative near-infrared spectra in the region of 1,100 to 2,350 nm. Characteristic lipid shifts at 1,200, 1,720 and 2,300 nm were noted in all other animals, as well. Magnified views are shown in parts B to D. Spectra in the (B) long-range wavelength region (2,200 to 2,330 nm) and mid-range regions (C: 1,620 to 1,820 nm; D: 1,130–1,260 nm).
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Figure 3 The near-infrared spectral features corresponding to the histologic features of human carotid atherosclerotic plaque. (A) The near-infrared spectra. The dotted line corresponds to the fibrous region; the dashed line to the calcified region; and the solid line to the yellow, soft region. The second derivative was also analyzed (B), thereby reducing the heavy influence of the water signal. The three plaque regions became more homogeneous in signal, but some differences remained, particularly in the region of 1,660 to 1,750 nm. A.U. = absorbance units.
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Figure 4 Photomicrographs illustrating the histopathologic findings of a portion of a stable, firm plaque, mostly collagen, with a few small areas of lipid deposits (a), and an advanced atherosclerotic lesion in an unstable, soft plaque (b). A thin cap (arrow) of  150 µm is mainly composed of collagen layers. The necrotic core consists of lipids, cholesterol crystals (clefts), fibrin, lymphocytes and macrophages. Hematoxylin-eosin staining; magnification x50. C = calcium; E = erosion; FC = fibrous cap; I = inflammation; L = lipid pool; M = medial layer; N = necrosis; P = platelet clump; T = thrombus.
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Figure 5 Cluster analysis yields an empiric classification of the NIR reflectance spectra of fresh atherosclerotic plaques from 25 patients. The plaques were divided into 145 fresh (living, unfixed) specimens and examined by near-infrared reflectance spectroscopy. Cluster analysis reveals three distinct groups (right panels) in each of two spectral regions: 2,240 to 2,320 nm (A) and 1,665 to 1,800 nm (B). For each of the six cluster groups (right panels), an average spectrum was calculated (A and B). Group 1 specimens in the top and bottom panels correspond to American Heart Association (AHA) type I and II stable lesions. Group 2 specimens correspond to AHA type III and IV lesions. Group 3 specimens correspond to AHA type V and VI lesions.
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