Figure 1. Empagliflozin suppressed kidney fibrosis in association with inhibition of the EMT in kidneys from diabetic CD-1 mice.

Higher-magnification (original magnification, ×300) (A–D) and lower-magnification (original magnification, ×100) (E–H) images of Sirius Red staining for fibrosis. Scale bar: 60 μm (A–D); 20 μm (E–H). (I) Relative fibrosis areas were calculated using ImageJ software. Six independent high-magnification images of the staining were analyzed. n = 6. (J–N) PAS staining was performed in kidney paraffin sections. Six independent images of the staining were analyzed. n = 6 in each group. Scale bar: 80 μm. (N) Quantification of the relative surface area of glomeruli by ImageJ software. (O–Q) Electron microscopy (EM) was performed to evaluate glomerular damage. Representative images are presented. n = 2. Scale bar: 1 μm. (R) Representative Western blotting images of mesenchymal markers in kidney samples. β-Actin from same gel is shown under the corresponding blots as loading control. (S–U) Densitometric analysis of the Western blotting results normalized to β-actin. n = 5 in each group. (V–Y) Immunohistochemical analysis for vimentin. Deparaffinized sections were analyzed from each group of mice. n = 5. Scale bar: 50 μm. Representative data are shown. The data are expressed as mean ± SD. One-way ANOVA followed by Tukey’s multiple comparison test was used to determine significance, which was defined as P < 0.05. Empa, empagliflozin; NC, negative control.