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目的探讨双氢青蒿素体外抗肝纤维化作用及其可能机制。方法双氢青蒿素设50、25、12.5、6.25、3.125、1.5625、0.78125、0.390125μg/ml共8个浓度进行细胞毒性试验,选择最大无细胞毒作用的双氢青蒿素浓度进行以下实验。体外培养肝星状细胞(HSC),分为正常对照组、铁沉积模型组[25μmol/L柠檬酸铁胺(FAC)]、去铁铵组(50μmol/L去铁铵)、双氢青蒿素组(双氢青蒿素)、铁沉积+双氢青蒿素组(25μmol/L FAC+双氢青蒿素)和去铁铵+双氢青蒿素组(50μmol/L去铁铵+双氢青蒿素),培养24 h。免疫组化法检测各组细胞中α平滑肌肌动蛋白(α-SMA)的表达,PCR法检测HSC铁代谢及细胞活化相关因子[包括铁调素、转铁蛋白受体1(TfR1)、Ⅰ型胶原及转化生长因子β1(TGF-β1)]mRNA表达。结果筛选出6.25μg/ml浓度的双氢青蒿素进行试验。铁沉积模型组细胞中α-SMA大量表达,各给药组细胞中α-SMA表达较正常对照组和铁沉积模型组明显减少。铁沉积模型组铁调素mRNA表达明显降低,而经去铁铵和(或)双氢青蒿素干预后,铁调素mRNA表达均显著升高(P<0.05)。双氢青蒿素组、铁沉积+双氢青蒿素组和去铁铵+双氢青蒿素组能明显降低TfR1、Ⅰ型胶原、TGF-β1 mRNA表达(P<0.05)。结论双氢青蒿素具有一定的体外抗肝纤维化作用,其机制与调节细胞铁的代谢、抑制肝星状细胞活化有关。
Objective To investigate the anti-hepatic fibrosis effect of dihydroartemisinin in vitro and its possible mechanism. Methods Dihydroartemisinin was set at a concentration of 50,25,12.5,6.25,3.125,1.5625,0.78125,0.390125μg / ml in a total of eight concentrations of cytotoxicity test, select the maximum concentration of cytotoxic effects of dihydroartemisinin the following experiment . Hepatic stellate cells (HSCs) were cultured in vitro and divided into normal control group, iron deposition model group (25μmol / L ferric ammonium citrate (FAC)], desferrioxamine group (50μmol / L desferrioxamine) (Dihydroartemisinin), iron deposition + dihydroartemisinin group (25μmol / L FAC + dihydroartemisinin), and deferoxamine + dihydroartemisinin group (50μmol / L desferrioxamine + bis Artemisinin), cultured for 24 h. The expression of α-smooth muscle actin (α-SMA) in each group was detected by immunohistochemistry. The expressions of iron metabolism and cell activation related factors [including hepcidin, transferrin receptor 1 (TfR1) Type collagen and transforming growth factor β1 (TGF-β1)] mRNA expression. As a result, dihydroartemisinin at a concentration of 6.25 μg / ml was screened. The expression of α-SMA in the iron deposition model group was abundant, and the expression of α-SMA in each group was significantly reduced compared with the normal control group and the iron deposition model group. Hepcidin mRNA expression in the iron deposition model group was significantly reduced, while hepcidin mRNA expression was significantly increased (P <0.05) after intervention with desferrioxamine and / or dihydroartemisinin. Dihydroartemisinin group, iron deposition + dihydroartemisinin group and deferoxamine + dihydroartemisinin group could significantly reduce the expression of TfR1, type Ⅰ collagen and TGF-β1 mRNA (P <0.05). Conclusion Dihydroartemisinin has certain anti-hepatic fibrosis effect in vitro, and its mechanism is related to regulation of iron metabolism and inhibition of hepatic stellate cell activation.