Modified Fangji Huangqi Decoction alleviates renal interstitial fibrosis by inhibiting TGF-β1/Smad/Snail signaling pathway during epithelial-mesenchymal transition
This study investigated the effects of modified Fangji Huangqi Decoction on the expression of proteins related to epithelial-mesenchymal transition(EMT)in a mouse model of unilateral ureteral obstruction(UUO)and in a rat renal tubular epithelial cell(NRK-52E)model of fibrosis induced by transforming growth factor β1(TGF-β1).It aims to decipher the molecular mechanism by which modified Fangji Huangqi Decoction alleviates renal interstitial fibrosis.C57/BL mice were subjected to UUO.After the surgery,the mice were treated with 0.5-fold and 2-fold concentrations of modified Fangji Huangqi Decoction and fosinopril sodium(positive control)for 7 days.The interstitial collagen deposition in the kidney was assessed by Masson staining.Western blot and RT-qPCR were employed to determine the expression levels of TGF-β1,phosphorylated Smad2/3(p-Smad2/3),Smad2/3,Snail,epithelial cadherin(E-cadherin),alpha smooth muscle actin(α-SMA),and vimentin.The NRK-52E cell model induced by TGF-β1 was treated with the serum samples collected from SD rats treated with different concentrations of modified Fangji Huangqi Decoction.The CCK-8 assay was employed to examine the effects of the serum samples on NRK-52E cell proliferation.The cell morphology in different groups was observed under a microscope.Furthermore,the modeled cells were treated with the serum containing 1-fold decoction.Western blot and RT-qPCR were then employed to measure the expression levels of p-Smad2/3,Smad2/3,Snail,E-cadherin,α-SMA,and vimentin in the cells.Under the same conditions,shRNA was used to silence the Snail gene,and measurements were repeated before and after treatment with the serum containing 1-fold decoction.The results indicated that modified Fangji Huangqi Decoction alleviated the fibrotic injury in the mouse model of UUO and the fibrosis in the NRK-52E cell model.The treatment with the decoction down-regulated the protein and mRNA levels of EMT-related indicators including p-Smad2/3,α-SMA,Snail,and vimentin,while it up-regulated the expression of E-cadherin.After shRNA silencing of the Snail gene,the protein and mRNA levels of E-cadherin,α-SMA,and vimentin showed no significant differences before and after treatment with the serum containing the decoction.The results suggest that modified Fangji Huangqi Decoction may alleviate renal interstitial fibrosis by inhibiting the TGF-β1/Smad/Snail signaling pathway and regulating the EMT process.
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