Table 1.
Physical and clinical parameters.
Table 2.
PCR primer sequences.
Figure 1.
Blood glucose levels among diabetic mice were matched throughout the study and showed similar glucose levels among the diabetic mice at 1, 2, 3, 4 and 5 months post diabetes induction and were significantly higher than control mice (A).
Empagliflozin reduced insulin requirement in diabetic mice (B). The comparison was made between average insulin requirement per dose administered thrice weekly, before and after initiation of empagliflozin. Data are expressed as mean ± SEM with * = P<0.05 and ** = P<0.001 vs ctrl.
Figure 2.
Diabetic mice demonstrated increased glomerulosclerosis and glomerular fibronectin deposition, which was improved by telmisartan but not by empagliflozin.
Representative photographs of PAS stained sections for A) ctrl, B) ctrl + empa, C) dm, D) dm + empa and E) dm + tel groups and quantification of glomerulosclerosis by glomerulosclerotic index (F). Representative photographs of immunohistochemistry for glomerular fibronectin in G) ctrl, H) ctrl + empa, I) dm, J) dm + empa and K) dm + tel groups and quantification of glomerular fibronectin by Image J (L) (Magnification = original X400). Data are expressed as mean ± SEM with ** = P<0.001 vs ctrl, # = P<0.05 vs dm and ## = P<0.001 vs dm.
Figure 3.
Diabetic animals demonstrated increased tubulointerstitial inflammation, which was not ameliorated by empagliflozin.
Representative photographs of immunohistochemistry for tubulointerstitial F4/80 stain for activated macrophages in A) ctrl, B) ctrl + empa, C) dm, D) dm + empa, E) dm + tel groups (Magnification = original X 400) and quantification of F4/80 positive cells in tubulointerstitium (F) which was done by calculating the average number of F4/80 positive cells per high power field (HPF) in each group. Representative photographs of immunohistochemistry for tubular TLR2 in G) ctrl, H) ctrl + empa, I) dm, J) dm + empa, K) dm + tel groups (Magnification = original X 200) and quantification of tubular TLR2 expression by Image J (L). (Data are expressed as mean ± SEM with * = P<0.05 vs ctrl, ** = P<0.001 vs ctrl and # = P<0.05 vs dm).
Figure 4.
Diabetic mice demonstrated tubular atrophy and tubulointerstitial fibrosis, which was not improved by empagliflozin and partially reduced by telmisartan.
Representative photographs of PAS stained sections of tubulointerstitium in A) ctrl, B) ctrl + empa, C) dm, D) dm + empa, E) dm + tel groups and F) Quantification of tubular atrophy in all groups was done by counting the number of atrophic tubules per 400 tubule count (Data are expressed as mean ± SEM with * = P<0.05 vs ctrl). Representative photographs of tubulointerstitial picrosirius red stain in G) ctrl, H) ctrl + empa, I) dm, J) dm + empa, K) dm + tel groups and L) Quantification of tubulointerstitial Sirius red positive collagen content by Image J. Representative photographs of Masson's stain in M) ctrl, N) ctrl + empa, O) dm, P) dm + empa, Q) dm + tel groups and R) Quantification of Masson's positive collagen content by Image J (Magnification = original X 200). (Data are expressed as mean ± SEM with * = P<0.05 vs ctrl, ** = P<0.001 vs ctrl).
Figure 5.
Diabetic mice showed increased renal cortical transcription of inflammatory and fibrotic cytokines, which was not improved by empagliflozin.
Real time PCR results for Renal cortical transcription of A) MCP-1, B) Collagen 4, C) TGFβ and D) Fibronectin relative to actin (Data are expressed as mean ± SEM with * = P<0.05 vs ctrl and ** = P<0.001 vs ctrl).
Figure 6.
Empagliflozin increased renal GLUT1 transcription in comparison with untreated diabetic mice although renal protein expression of GLUT1 and SGLT2 were unchanged.
Real time PCR results for A) GLUT1, B) GLUT2, C) SGLT1 and D) SGLT2 expressed relative to actin. Expression levels of renal GLUT1 protein (E and F) and SGLT2 (G and H) relative to actin (Data are expressed as mean ± SEM with # = P<0.05 vs dm).