Mechanism of protopine in inhibiting pulmonary fibrosis based on network pharmacology and in vitro experiments
Objective:Exploring the mechanism of protopine(PTP)in inhibiting pulmonary fibrosis based on network pharma-cology and in vitro Experiments.Methods:Employing network pharmacology,we identified the core targets of PTP against pul-monary fibrosis and predicted the associated pathways.To validate these findings from network pharmacology,we constructed an in vitro model of lung fibrosis using human A549 cells induced by transforming growth factor beta1(TGF-β1).The experiment was divided into control group,TGF-β1 group,pirfenidone(PFD)intervention group,and PTP intervention group.Cell prolifer-ation activity was assessed using the CCK8 assay;cell morphology was observed under an inverted microscope;cell migration abil-ity was evaluated through the cell scratch assay;real-time quantitative PCR(RT-qPCR)was employed to detect the mRNA lev-els of E-cadherin,Vimentin,fibronectin(FN),and collagen Ⅰ;protein expression levels of E-cadherin,Vimentin,α-smooth muscle actin(α-SMA),and FN were analyzed by Western blot;and the expression of E-cadherin and FN proteins was observed under an upright fluorescence microscope.Results:The results of network pharmacology indicate that PTP primarily targets core such as ALB,AKT1,EGFR,HSP90AA1,and SRC,and modulates pathways including cancer pathways and the PI3K-Akt sig-naling pathway.The in vitro experimental results demonstrate that compared to the control group,the TGF-β1 group exhibits a morphological transition of A549 cells from epithelial to fibroblastic,with elevated expression of mesenchymal cell markers α-SMA,Vimentin,FN,and Collagen Ⅰ,and decreased expression of epithelial cell marker E-cadherin,accompanied by en-hanced cell migration capabilities.Following treatment with PTP,it effectively inhibits the proliferation and migration abilities in-duced by TGF-β1 in A549 cells.PTP also enhances the expression of E-cadherin in the induced cells while reducing the levels of Vimentin,α-SMA,FN,and Collagen Ⅰ.Conclusion:Research indicates that PTP may inhibit pulmonary fibrosis by suppressing the epithelial-mesenchymal transition induced by TGF-β1.
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