Wenfei Huaxian Granules ameliorated pulmonary fibrosis through modulating the signaling pathway of TGF-β1/Smad
OBJECTIVE To explore the effect and mechanism of Wenfei Huaxian Granules for BLM-induced pulmonary fibrosis in mice.METHODS The C57BL/6J male mice were randomized into five groups of Control,Model,WFHX-low-dose(12.5 g·kg-1),WFHX-high-dose(50 g·kg-1)and Pirfenidone(180 mg·kg-1).The model of pulmonary fibrosis was estab-lished by an endotracheal intubation infusion of bleomycin(0.75 mg·kg1).The gavage intervention began from the second day of modeling and continued for 28 days.Degrees of alveolitis and fibrosis were detected after hematoxylin-eosin and Masson staining.The mRNA expressions of interleukin-6(IL-6),tumor necrosis factor α(TNF-α),collagen Ⅰ and epithelial-mesenchymal tran-sition(EMT)related markers in lung tissues were detected by real-time quantitative polymerase chain reaction(RT-qPCR).And the expressions of IL-6,TNF-α,collagen Ⅰ,EMT-related markers and transforming growth factor β1(TGF-β1)/Smad signaling pathway-related proteins in lung tissues were detected by Western blot.The expressions of p-Smad3 and Smad7 were detected by immunohistochemical(IHC)stain.RESULTS As compared with Control group,tissue fibrosis score was signifi-cantly higher in Model group and the mRNA and protein expression levels of IL-6,TNF-α,collagen Ⅰ,mesenchymal markers fibronectin(Fn),vimentin and α-smooth muscle actin(α-SMA)were significantly up-regulated(P＜0.01)while epithelial marker calmodulin E(E-cadherin)was markedly down-regulated(P＜0.05).Meanwhile,the protein expressions of TGF-β1,p-Smad2 and p-Smad3 spiked while Smad7 expression declined.As compared with Model group,intervention could lessen the degree of fibrosis in lung tissues.In all WFHX-dosed groups,the expressions of the above factors improved somewhat.The releases of IL-6 and TNF-α were blunted and inflammatory responses of lung tissues attenuated.Secretion of collagen Ⅰ and pro-duction of TGF-β1 became suppressed.EMT improved.Phosphorylation of Smad2/Smad3 was silenced and the expression of Smad7 became up-regulated.CONCLUSION Wenfei Huaxian Granules may exert anti-fibrotic effects through blunting inflam-matory responses,silencing EMT and suppressing the signaling pathway of TGF-β1/Smad.
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