Label-free visualization of acetaminophen-induced liver injury by high-speed stimulated Raman scattering spectral microscopy and multivariate image analysis

We recently established a high‐speed, label‐free, spectral imaging method based on stimulated Raman scattering (SRS). This method enables examination of cellular features within relatively short periods, thus enabling new imaging applications in pathology. Previously, we reported on label‐free visualization of mouse tissue using SRS spectral microscopy combined with multivariate image analysis, but the feasibility of applying this approach to diseased tissues with diverse morphology and irregular chemical species has not been examined. We, therefore, assessed acetaminophen‐induced liver injury to evaluate the potential use of Raman spectral microscopy for visualizing histopathologic specimens. Acetaminophen‐overdosed mouse liver was prepared and the pathologic changes including centrilobular necrosis were confirmed. Multi‐colored images were reconstructed through principal component analysis (PCA) of a multi‐band SRS dataset, which provided rich information compared with a monochrome single‐band SRS dataset. A wide view of the multi‐colored principal component (PC) images showed the distribution of cellular constituents, which was similar to that observed by fat staining. In addition, different types of cells in liver parenchyma were also demonstrated. In conclusion, the combination of SRS spectral microscopy and PCA has the potential to reveal both the morphological and chemical features of specimens and therefore has potential utility in diagnostic pathology.