碧萝芷通过下调ERK磷酸化及自噬水平抑制TGF-β1诱导的肝星状细胞活化

Pycnogenol suppresses TGF-β1-induced hepatic stellate cell activation via ERK-mediated autophagy inhibition

目的:探究碧萝芷对转化生长因子β1(TGF-β1)诱导的肝星状细胞活化的影响。方法:5μg/L TGF-β1和不同浓度碧萝芷(0、10、25、50 mg/L)分别作用于LX-2细胞,在有或无自噬抑制剂3-MA和ERK抑制剂PD98059的情况下,用MTT法检测细胞活力的变化,Western blot实验检测α-SMA、ColⅠ、TIMP-1、LC3-Ⅱ/Ⅰ、beclin1、p-ERK1/2和ERK1/2蛋白水平的变化。结果:与对照组相比,5μg/L TGF-β1组的LX-2细胞活力以及α-SMA、ColⅠ、TIMP-1、LC3-Ⅱ/Ⅰ、beclin 1、p-ERK1/2和ERK1/2蛋白水平明显增加(P<0.05)。而碧萝芷预处理能逆转上述效应,并呈现一定的剂量依赖性,50 mg/L碧萝芷的抑制效果最为显著(P<0.05)。而且,与TGF-β1组相比较,50 mg/L碧萝芷、5 mmol/L 3-MA或者20μmol/L PD98059预处理下,TGF-β1诱导的LX-2细胞活力以及α-SMA和LC3-Ⅱ/Ⅰ蛋白表达均明显下调(P<0.05)。结论:碧萝芷通过下调ERK磷酸化及自噬水平抑制TGF-β1诱导的肝星状细胞活化。

AIM：To explore the effect of Pycnogenol on transforming growth factor-β1（ TGF-β1）-induced hepatic stellate cell activation.METHODS：Cultured LX-2 cells were treated with 5 μg/L TGF-β1 and different concentrations（ 0,10,25 and 50 mg/L） of Pycnogenol.The viability of the LX-2 cells under the conditions with or without autophagy inhibitor 3-MA and ERK inhibitor PD98059 was determined by MTT assay.The protein levels of α-SMA,Col Ⅰ,TIMP-1,LC3-Ⅱ/Ⅰ,beclin 1,p-ERK1/2 and ERK1/2 were detected by Western blot.RESULTS：Compared with control group,5 μg/L TGF-β1 treatment elevated the cell viability,and increased the protein levels of α-SMA,ColⅠ,TIMP-1,LC3-Ⅱ/Ⅰ,beclin 1,p-ERK1/2,and ERK1/2 in the LX-2 cells（ P〈0.05）.However,these effects were reversed by Pycnogenol pretreatment in a dose-dependent manner and the inhibitory effect of 50 mg/L Pycnogenol was the most significant in the LX-2 cells（ P〈0.05）.Furthermore,compared with TGF-β1 group,pretreatment with 50 mg/L Pycnogenol,5 mmol/L 3-MA or 20 μmol/L PD98059 downregulated TGF-β1-induced cell viability and the protein levels of α-SMA and LC3-Ⅱ/Ⅰ in the LX-2 cells（ P〈0.05）.CONCLUSION：Pycnogenol suppresses TGF-β1-induced hepatic stellate cell activation via p-ERK and autophagy inhibition.