Study On The Effect And Mechanism Of Total Flavonoids Of Astragalus On Hypertrophic Scar Fibroblasts

Objective: To investigate the effect and mechanism of total flavonoids of astragalus(TFA)on hypertrophic scar fibroblasts(HSF cells).Methods: 1.In this study,HSF cells were studied and a control group,TFA low dose group(10 μg/m L),TFA medium dose group(25 μg/m L)and TFA high dose group(50μg/m L)were set up and given TFA treatment for 24 h.Cell supernatants or cells were collected for subsequent experiments.2.The MTT method was used to determine the proliferation ability of each group of HSF cells in vitro and to evaluate the effect of TFA on cell proliferation.3.The expression of Col I and Col III in the cell supernatant was determined by ELISA;the protein expression levels of Col I,Col III and α-SMA in the cells were determined by Western blot;the m RNA expression levels of Col I,Col III and α-SMA in HSF cells were detected by RT-PCR to evaluate the effects of TFA on the extracellular matrix(ECM)of HSF cells.The effect of TFA on the extracellular matrix(ECM)of cells was evaluated.4.The expression of TGF-β1,p-Smad2 and p-Smad3 in HSF cells was measured by Western blot,and the role of TGF-β1/Smad signaling pathway in the inhibition of HSF cell proliferation and ECM deposition by TFA was analyzed.5.Western blot assay was performed to detect p-PI3 K,p-AKT and p-m TOR protein expression in the cells and analyze the role of PI3K/Akt/m TOR signaling pathway in the inhibition of HSF cell proliferation and ECM deposition by TFA.Results:1.The results of MTT assay showed that TFA inhibited the proliferation of HSF cells.2.ELISA results showed that the expression of Col I and Col III in the cell supernatants of the 10,25 and 50 μg/m L TFA groups was significantly decreased relative to the control group(P<0.01,P<0.05).Western blot results showed that the expression of Col I,Col III and a-SMA in the cells of the 10,25 and 50 μg/m L TFA groups was significantly decreased relative to the control group(P<0.01,P<0.05).RT-PCR results showed that relative to the control group,TFA significantly inhibited Col I(10,25 and 50 μg/m L TFA),Col III(10,25 and 50 μg/m L TFA)and a-SMA(25and 50 μg/m L TFA)m RNA expression(P<0.01,P<0.05).Thus,TFA inhibited ECM deposition in HSF cells.3.Western blot results showed that 10,25 and 50 μg/m L TFA significantly inhibited intracellular TGF-β1,p-smad2 and p-smad3 protein expression relative to the control group(P<0.01,P<0.05).Therefore,TFA may inhibit the proliferation and ECM deposition of HSF cells by regulating the TGF-β1/Smad signaling pathway.4.Western blot results showed that the protein expression levels of p-PI3 K,p-Akt and p-m TOR were significantly reduced in the 10,25 and 50 μg/m L TFA groups relative to the control group(P<0.01,P<0.05).Thus,TFA may reduce HSF cell proliferation and ECM deposition by inhibiting the PI3K/AKT/m TOR signaling pathway.Conclusion: TFA may inhibit HSF cell proliferation and ECM deposition by regulating TGF-β1/Smad and PI3K/Akt/m TOR signaling pathways,thereby ameliorating HS development and progression.Therefore,TFA can be used as a candidate drug for the treatment of HS.