微摩尔浓度铜离子通过刺激活性氧产生促进人角质-第三军医大学学报.docVIP

微摩尔浓度铜离子通过刺激活性氧产生促进人角质形成细胞增殖400038 重庆，第三军医大学西南医院全军烧伤研究所，创伤、SKLZZ201226) ［通信作者］ 彭代智， E-mail: dzpengpub@126.com [摘要] 目的 观察不同浓度Cu2+对人角质形成细胞株（HaCaT）增殖活性及细胞内活性氧（ROS）水平的影响，用活性氧清除剂N-乙酰半胱氨酸（NAC）验证两者之间的关系。方法 Cu2+（0、10-8 、10-7 、10-6、10-510-4、10-或（和）5 m NAC处理HaCaT细胞后，采用CCK-8法和DCFH-DA分别检测细胞的增殖活性和细胞内ROS产生结果 与阴性对照相Cu2+)比，10-6、10-5 Cu2+处理48 h后HaCaT细胞相对增殖率升高116.19%（P=0.027）、113.37%（P=0.036），，而其细胞内ROS产生在30 min达到127.47%（P=0.001）、124.28%（P=0.031）60 min进一步上升至140.47%（P=0.002）、134.29%（P=0.007）。5 m NAC能拮抗微摩尔浓度Cu2+的上述作用。10-8、10- Cu2+在处理60 min也刺激细胞内ROS产生明显升高至110.99%（P=0.016）及117.34%（P=0.016）细胞（P值均大于0.05）结论 浓度Cu2+能促进人角质形成细胞增殖，这种促增殖效应可能与细胞内ROS水平的适度上调有关。 关键词铜离子；人角质形成细胞；细胞增殖；活性氧；N-乙酰半胱氨酸icromolar concentration of copper (ii) ions enhance the proliferation of human skin keratinocyte through appropriate generation of reactive oxygen species Duan Xiaodong, Peng Daizhi, Zhang Yilan, Chen Zhiyong, Huang Yalan (Institute of Burn Research, Southwest Hospital, State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical University, Chongqing, 40038, China) [Abstract] Objective To investigate the effects of micromolar copper (ii) ions (Cu2+) on the proliferation and reactive oxygen species (ROS) generation of human skin keratinocyte line HaCaT cells， and verify the role of ROS in human keratinocyte proliferation. Methods HaCaT cells were treated with different concentrations of copper (ii) ions (10-8 , 10-7 , 10-6 , 10-5 , 10-4 and 10-3 mol/L) or (and) 5 m mol/L N-Acetyl-L-cysteine (NAC), the proliferation rates and ROS generation of HaCaT cells were measured by cell counting kit-8 (CCK-8) and fluorescent probe DCFH-DA, respectively. Both intracellular?ROS?levels and mitochondria?localization were labeled by coculture with DCFH-DA and MitoTracker probes, and visualized under laser confocal microscopy. Results After cultured with 10-6 mol/L and 10-5 mol/L Cu2+ for 48h, the cell relative proliferation rates reached 116.19% (P=0.027) and 113.37% (P=0.036) respectively and were signi