Nanozoomer digital pathology virtual slide viewer software program

After fixation in 10% neutral-buffered formalin for 24 h, paraffin-embedded liver specimens were systematically cut into sequential 4-μm-thick sections. For histological analyses of the liver, images of hematoxylin and eosin (H&E), oil red-O, and immunohistochemistry.
IHC sections were captured and quantified using the NanoZoomer Digital Pathology Virtual Slide Viewer software program (Hamamatsu Photonics Corp, Hamamatsu, Japan). To evaluate the degree of lipid accumulation (steatosis score and lipid accumulation score for the liver), we performed oil red-O staining using frozen liver sections and categorized the tissues into 4 grades, as follows: no lipid droplets (score = 0); lipid droplets in <33% of hepatocytes (score = 1); lipid droplets 33%–66% of hepatocytes (score = 2) and lipid droplets in >66% of hepatocytes (score = 3). In addition, the degree of liver cell ballooning injury (ballooning score) was classified into three grades as follows: none (score = 0); few balloon cells (score = 1) or many balloon cells/prominent ballooning (score = 2).

After fixation in 10% neutral-buffered formalin over 24 h, the same liver lobe of all mice was selected, embedded in paraffin, cut into sequential sections of 3–5 μm in thickness and subjected to Hematoxylin-Eosin (HE) staining and immunohistochemistry (IHC). The frozen sections were cut into 5- to 8-μm sequential sections and subjected to Oil Red O staining (Oil Red O Stain Kit; Polysciences, Inc., Warrington, PA, USA). All section images were captured with the Nano Zoomer Digital Pathology Virtual Slide Viewer software program (Hamamatsu Photonics Corp, Hamamatsu, Japan). As described in a previous study [11 (link)], we used the NAFLD score (steatosis score and ballooning score) to assess the condition of the mouse liver (Table 1).