Arterial tissues collected from Ossabaw swine bearing metabolic syndrome-induced cardiovascular plaques are characterized by multimodal nonlinear optical microscopy that allows coherent antiStokes Raman scattering, second-harmonic generation, and two-photon excitation fluorescence imaging on the same platform. Significant components of arterial walls and atherosclerotic lesions, including endothelial cells, extracellular lipid droplets, lipid-rich cells, low-density lipoprotein aggregates, collagen, and elastin are imaged without any labeling. Emission spectra of these components are obtained by nonlinear optical microspectrometry. The nonlinear optical contrast is compared with histology of the same sample. Multimodal nonlinear optical imaging of plaque composition also allows identification of atherosclerotic regions that are vulnerable to rupture risk. The demonstrated capability of nonlinear optical microscopy for label-free molecular imaging of atherosclerotic lesions with 3-D sub-micrometric resolution suggests its potential application to the diagnosis of atherosclerotic plaques, determination of their rupture risk, and design of individualized drug therapy based on plaque composition.

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