【摘要】 目的 探讨药食两用物质陈皮的活性成分川陈皮素（nobiletin,NOB）对过氧化氢（hydrogen peroxide,H₂O₂）诱导的人皮肤成纤维细胞（human skin fibroblasts,HSF）衰老的保护作用及其与沉默信息调节因子1（silence information regulator 1,SIRT1）/5′腺苷酸活化蛋白激酶（AMP-activated protein kinase,AMPK）介导自噬的关系。方法 体外培养HSF，以700μM H₂O₂处理4 h建立细胞衰老模型，设置空白对照组，模型组，川陈皮素低、中、高剂量组（浓度分别为3μM、6μM、12μM）及阳性对照组（维生素C，浓度为100μM）。检测指标包括细胞衰老β-半乳糖苷酶（senescence-associated β-galactosidase,SA-β-gal）阳性率及细胞内活性氧（reactive oxygen species,ROS）水平，并通过蛋白质印迹法和免疫荧光技术分析p53、p16、p21、自噬相关蛋白Beclin-1、微管相关蛋白1轻链3（microtubule-associated protein 1 light chain 3,LC3），以及哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin,mTOR）及其磷酸化形式（p-mTOR），并分别计算磷酸化AMPK（p-AMPK）/AMPK与p-mTOR/mTOR的比值。结果 与空白对照组相比，H₂O₂刺激后显著升高了模型组SA-β-gal阳性率和ROS的表达水平，上调p53、p16、p21表达，并抑制Beclin-1和LC3-II表达，降低SIRT1和p-AMPK/AMPK比值，升高p-mTOR/mTOR比值；与模型组相比，川陈皮素能够以剂量依赖的方式，显著逆转上述病理变化。结论 川陈皮素可减轻H₂O₂诱导的HSF衰老表型，其机制可能与激活SIRT1/AMPK相关自噬并抑制mTOR信号通路有关。更多还原

【Abstract】 Objective To investigate the protective effect of nobiletin（NOB）, an active ingredient of the food-medicine dual-use herb Chenpi（Citri Reticulatae Pericarpium）, against hydrogen peroxide（H₂O₂）-induced senescence in human skin fibroblasts（HSF）, and its relationship with autophagy mediated by silent information regulator 1（SIRT1）/AMP-activated protein kinase（AMPK）. Methods HSF were cultured in vitro, and a cellular senescence model was established by treatment with 700μM H₂ O₂ for 4 h. The experimental groups included a blank control group, a model group, NOB intervention groups（at concentrations of 3, 6, and 12 μM, respectively）, and a positive control group（vitamin C at 100 μM）. The senescenceassociated β-galactosidase（SA-β-gal） positivity rate and intracellular reactive oxygen species（ROS） levels were assessed.Western blotting and immunofluorescence were employed to analyze the expression of p53, p16, p21, autophagy-related proteins Beclin-1 and microtubule-associated protein 1 light chain 3（LC3）, as well as mammalian target of rapamycin（mTOR）and its phosphorylated form（p-mTOR）. The ratios of phosphorylated AMPK（p-AMPK） to AMPK and p-mTOR to m TOR were calculated. Results Compared with the blank control group, H₂ O₂ stimulation significantly increased the SA-β-gal positivity rate and ROS levels in the model group, upregulated the expression of p53, p16, and p21, inhibited the expression of Beclin-1 and LC3-II, reduced SIRT1 level and the p-AMPK/AMPK ratio, and elevated the p-mTOR/mTOR ratio. Compared with the model group, NOB dose-dependently reversed these pathological changes. Conclusion NOB alleviates the H₂O₂-induced senescent phenotype in HSF, and its mechanism may be associated with activating SIRT1/AMPK-related autophagy and inhibiting the m TOR signaling pathway.更多还原