氧化应激介导达氟沙星诱导LLC-PK1细胞周期阻滞  

Oxidative stress inducting danofloxacin induces cell cycle arrest in LLC-PK1

作　　者：李玉娟[1] 代佳娣 张大生[1] 孙志良[1] 刘兆颖[1] 

机构地区：[1]湖南农业大学动物医学院,湖南省兽药工程技术研究中心,长沙410128

出　　处：《中国抗生素杂志》2016年第5期388-395,共8页Chinese Journal of Antibiotics

基　　金：国家自然科学基金项目(No.31101859)

摘　　要：目的探讨达氟沙星诱导LLC-PK1细胞氧化应激后细胞损伤的分子机制。方法不同浓度的达氟沙星处理LLCPK1细胞后,通过WST-1法检测细胞增殖抑制情况,DHE、Amplex Ultra Red法分别检测超氧阴离子自由基(O2·-)、H2O2含量,Rhodamine123法检测线粒体膜电位,流式细胞仪检测分析细胞周期及RT-PCR法检测细胞周期相关基因表达含量的变化。结果达氟沙星呈浓度依赖性的抑制LLC-PK1细胞的增殖、升高细胞内O2·-、H2O2含量、诱导细胞周期阻滞,但线粒体膜电位没有显著性下降。100μmol/L浓度达氟沙星处理LLC-PK1细胞后细胞出现G2期周期阻滞,ROS升高含量在自身抗氧化系统修复范围内,细胞并没有出现明显损伤;而浓度达到400μmol/L时细胞发生氧化应激出现可逆性的G1期细胞阻滞,此时细胞周期素抑制蛋白p53、p21、p27m RNA表达显著升高,LLC-PK1细胞对氧化应激造成的损伤进行修复。结论 400μmol/L浓度范围内的达氟沙星处理LLC-PK1细胞24h所致的氧化应激诱导细胞周期阻滞,细胞损伤在自身修复范围之内并未导致细胞凋亡。Objective To evaluate the underlying molecular mechanisms of cell damage and oxidative stress induced by danofloxacin in LLC-PK1 cells. Methods The growth inhibitory effect, the change of O2^·- , H2O2 was determined by WST-1, DHE and Amplex UltraRed method, respectively. Mitochondrial membrane potential was examined by rhodamine 123. The cell cycle and the expression change of related gene were measured by flow cytometry and RT-PCR, respectively. Results Danofloxacin induced a concentration-dependent suppress in the proliferation of LLC-PK1. The 02＂, H202 production were also increased and caused cell cycle arrest, but the mitochondrial membrane potential of cells was no significantly decline. When 100μmol/L danofloxacin treated with LLC-PK1 cells, cycle was arrested in G2 phase. These results indicated that cell didn＇t significant damage and the increased of ROS was in the range of self-repair. However, 400μmol/L danofloxacin could induce a reversible G1 phase cell cycle arrest. The cyclin inhibition protein p53, p21, p27 mRNA expression were significantly increased. The oxidative stress induced by danofloxacin in LLC-PK1 cells could aslo be repaired. Conclusion Danofloxacin in the concentration range of 400μmol/L could induce cell cycle arrest in LLC-PK1 cells within 24h, but did not induce cell apoptosis because cellular damage could be repaired.

关 键 词：达氟沙星 LLC—PK1 氧化应激 肾毒性 细胞周期 

分 类 号：R978.1[医药卫生—药品] S859.796[医药卫生—药学]