Journal: PeerJ

Article Title: Dapagliflozin attenuates diabetic renal fibrosis by inhibiting macrophage-myofibroblast transition via the TGF-β1-Smad3/7 pathway

doi: 10.7717/peerj.21321

Figure Lengend Snippet: (A) Representative IHC staining images of TGF-β1, p-Smad3, and Smad7 in renal tissues. Diabetic mice showed increased TGF-β1/p-Smad3 and decreased Smad7, which was reversed by DAPA. Scale bar = 2 0 µm. (B) Western blot analysis of TGF-β1, p-Smad3, and Smad7 protein expression in renal tissues (normalized to β-actin). TGF-β1 and p-Smad3/Smad3 ratio were increased and Smad7 was decreased in diabetic mice, and DAPA restored the Smad3/Smad7 balance. Data are presented as mean ± SD with individual data points (dots). n = 6 per group for I HC; n = 3 per group for Western blot. * p < 0.05 vs. Ctrl group; # p < 0.05 vs. DKD group.

Article Snippet: Membranes were blocked with 5% skim milk (1 h, room temperature), then incubated overnight at 4 °C with primary antibodies: TGF-β1 (A16640, 1:1000, Abclonal, Wuhan, China), p-Smad3 (AP0727, 1:1000, Abclonal, Wuhan, China), Smad3 (A22133, 1:2000, Abclonal, Wuhan, China), Smad7 (A16396, 1:2000, Abclonal, Wuhan, China), α-SMA (A17910, 1:2000, Abclonal, Wuhan, China), Col-I (14695-1-AP, 1:2000, Proteintech, Wuhan, China), and β-actin (db13986, 1:10,000, Diagbio, Hangzhou, China).

Techniques: Immunohistochemistry, Western Blot, Expressing

Journal: PeerJ

Article Title: Dapagliflozin attenuates diabetic renal fibrosis by inhibiting macrophage-myofibroblast transition via the TGF-β1-Smad3/7 pathway

doi: 10.7717/peerj.21321

Figure Lengend Snippet: In DKD, hyperglycemia induces renal inflammation and activation of the TGF-β1/Smad3 pathway, which drives macrophages (F4/80 + ) to undergo MMT, acquiring myofibroblast characteristics (α-SMA + ) and producing Col-I, leading to tubulointerstitial fibrosis and progressive kidney dysfunction. DAPA inhibits TGF-β1/Smad3 pathway activity, upregulates Smad7 expression, restores Smad3/Smad7 homeostatic balance, and is highly likely to block the MMT process through this mechanism, thereby reducing α-SMA and Col-I deposition, and ultimately ameliorating diabetic renal fibrosis. Created at BioRender.com.

Article Snippet: Membranes were blocked with 5% skim milk (1 h, room temperature), then incubated overnight at 4 °C with primary antibodies: TGF-β1 (A16640, 1:1000, Abclonal, Wuhan, China), p-Smad3 (AP0727, 1:1000, Abclonal, Wuhan, China), Smad3 (A22133, 1:2000, Abclonal, Wuhan, China), Smad7 (A16396, 1:2000, Abclonal, Wuhan, China), α-SMA (A17910, 1:2000, Abclonal, Wuhan, China), Col-I (14695-1-AP, 1:2000, Proteintech, Wuhan, China), and β-actin (db13986, 1:10,000, Diagbio, Hangzhou, China).

Techniques: Activation Assay, Activity Assay, Expressing, Blocking Assay