摘要目的:探讨骨髓间充质干细胞来源的外泌体(BMMSC-exos)对活化肝星状细胞(HSC)的作用及其机制。方法:应用脂多糖(LPS)诱导HSC形成激活体外模型。按照实验要求分为对照组(Con)、活化组(LPS)、LPS＋BMMSCs(L＋B)组、LPS＋BMMSC-exo(L＋B-exo)组、L＋B-exo＋小干扰RNA阴性对照组(si-NC)及L＋B-exo＋敲降NCOA4组(si-NCOA4)。通过细胞增殖测定细胞活力；实时定量聚合酶链反应(RT-qPCR)检测平滑肌激动蛋白(α-SMA)、核受体共激活因子4(NCOA4)和铁蛋白(FTH1)mRNA；蛋白质免疫印迹法检测NCOA4、FTH1、波形蛋白(Vimentin)、α-SMA、谷胱甘肽过氧化物酶4(GPX4)、p62、环氧合酶-2(COX-2)、苄氯素1(Beclin 1)及微管相关蛋白(LC3)蛋白表达水平；免疫荧光化学检测NCOA4、FTH1及Vimentin表达；检测丙二醛(MDA)、还原型谷胱甘肽(GSH)及脂质活性氧(Lipid ROS)含量变化，多组间的数据比较采用单因素方差分析。结果:BMMSC-exos对活化的HSC-T6细胞的影响：L＋B-exo组纤维化标志物α-SMA和Vimentin表达量(0.58±0.04、0.25±0.05)低于LPS组(1.26±0.04、1.06±0.03)和L＋B组(0.76±0.04、0.75±0.15， F＝50.607、35.916， P<0.05)。BMMSC-exos促进活化的HSC-T6细胞铁死亡和自噬：L＋B-exo组MDA与Lipid ROS的表达量[(4.27±1.06) nmol/(mg·prot)、8 950.49±114.34]高于LPS组[(2.27±0.73) nmol/(mg·prot)、5 492.73±25.69]和L＋B组[(3.27±0.90) nmol/(mg·prot)、7 198.05±116.63， F＝1.263、1 372.009， P<0.05]；L＋B-exo组GSH水平[(10.47±0.31) μg/10 6 cells]低于LPS组[(15.94±0.11) μg/10 6 cells]和L＋B组[(12.60±0.15) μg/10 6 cells， F＝646.842， P<0.05]；L＋B-exo组NCOA4蛋白表达量[(1.19±0.06)]高于LPS组[(0.73±0.12)]和L＋B组[(0.95±0.13)， F＝44.456， P<0.05]；L＋B-exo组FTH1蛋白表达量(0.25±0.05)低于LPS组(1.06±0.03)和L＋B组(0.75±0.15， F＝35.916， P<0.05)。NCOA4介导的铁蛋白自噬参与BMMSC-exos对HSC-T6细胞的作用：si-NCOA4组的MDA[(0.83±0.11) nmol/(mg·prot)]低于si-NC组[(1.17±0.09) nmol/(mg·prot)， F＝17.217， P<0.05]。 结论:BMMSC-exos通过NCOA4介导的铁蛋白自噬诱导活化的HSC铁死亡，从而抑制HSC活化，发挥抗肝纤维化作用。

abstractsObjective:To investigate the effects of bone marrow mesenchymal stem cell-derived exosomes (BMMSC-exos) on the activation of hepatic stellate cells (HSC) and its mechanism.Methods:Lipopolysaccharide (LPS) was used to induce the formation of an in vitro model of HSC activation. According to the experimental requirements, they were divided into control (Con), LPS, LPS+ BMMSCs (L+ B), LPS+ BMMSC-exo (L+ B-exo), L+ B-exo+ small interfering RNA-negative control (si-NC), and L+ B-exo+ knockdown of NCOA4 (si-NCOA4) groups. Cell viability was determined by cell proliferation. The mRNA expression levels of smooth muscle agonist protein (α-SMA), nuclear receptor coactivator 4 (NCOA4), and ferritin (FTH1) were detected by real-time quantitative polymerase chain reaction (RT-qPCR). The protein immunoblotting was performed for the detection of NCOA4, FTH1, Vimentin, α-SMA, glutathione peroxidase NCOA4, FTH1, Vimentin, α-SMA, glutathione peroxidase 4 (GPX4), p62, benzyl chloride 1 (Beclin 1), and microtubule-associated protein (LC3). The immunofluorescence chemistry was used to detect the expression of NCOA4, FTH1 and Vimentin. The changes in the content of malondialdehyde (MDA), reduced glutathione (GSH), and lipid ROS were detected. One-way analysis of variance (ANOVA) was used to compare the data among multiple groups. Results:Effects of BMMSC-exos on activated HSC-T6 cells: the expression of fibrosis markers α-SMA and Vimentin in the L+ B-exo group (0.58±0.04, 0.25±0.05) was lower than that in the LPS group (1.26±0.04, 1.06±0.03) and the L+ B group (0.76±0.04, 1.06±0.03, F=50.607, 35.916, P<0.05). BMMSC-exos promoted iron death and autophagy in activated HSC-T6 cells: the expression of MDA and Lipid ROS in the L+ B-exo group [(4.27±1.06) nmol/(mg·prot), 8 950.49±114.34] was higher than that in the LPS group [(2.27±0.73) nmol/(mg·prot), 5 492.73±25.69] and L+ B group [(3.27±0.90) nmol/(mg·prot), 7 198.05±116.63, F=1.263, 1 372.009, P<0.05]; GSH level in the L+ B-exo group [(10.47±0.31) μg/10 6 cells] was lower than that in the LPS group [(15.94±0.11) μg/10 6 cells] and L+ B group [(12.60±0.15) μg/10 6 cells, F=646.842, P<0.05]; and the expression of NCOA4 in the L+ B-exo group [(1.19±0.06)] was higher than that in the LPS group [(0.73±0.12)] and L+ B group [(0.95±0.13), F=44.456, P<0.05]; FTH1 expression in the L+ B-exo group (0.25±0.05) was lower than that in the LPS group (1.06±0.03) and the L+ B group (0.75±0.15, F=35.916, P<0.05). NCOA4-mediated ferritin autophagy was involved in the effect of BMMSC-exos on HSC-T6 cells: the MDA in the si-NCOA4 group [(0.83±0.11) nmol/(mg·prot)] was lower than that in the si-NC group [(1.17±0.09) nmol/(mg·prot), F=17.217, P<0.05]. Conclusion:BMMSC-exos induced iron death of activated HSC through NCOA4-mediated ferritin autophagy, thereby inhibiting HSC activation and exerting anti-hepatic fibrosis effects.