香烟烟雾提取物对大鼠股四头肌细胞氧化损伤的研究

目的 研究经香烟烟雾提取物（CSE）刺激后大鼠股四头肌细胞8-羟基脱氧鸟苷（8-OHdG）含量的变化以及细胞内活性氧（ROS）水平和8-羟基鸟嘌呤DNA糖苷酶1（OGG1）表达对8-OHdG含量变化的影响.方法 用不同浓度CSE分别刺激大鼠股四头肌细胞24 h、48 h和72 h后,采用高效液相色谱-电化学法（HPLC-ECD）检测大鼠股四头肌细胞8-OHdG的含量,流式细胞术检测大鼠股四头肌细胞ROS水平,实时定量PCR（Realtime PCR）检测OGG1 mRNA水平,用Western blot检测OGG1蛋白水平.结果 经相同CSE浓度分别刺激大鼠股四头肌细胞24 h、48 h和72 h后,大鼠股四头肌细胞中8-OHdG含量、ROS水平及OGG1 mRNA和蛋白水平差异均无统计学意义.但经不同浓度CSE刺激相同时间后,10.00％ CSE和20.00％CSE刺激组大鼠股四头肌细胞8-OHdG含量明显高于对照组和5.00％ CSE刺激组（P＜0.05）;大鼠股四头肌细胞内ROS水平、OGG1 mRNA和蛋白水平较对照组均明显升高（P＜0.05）;大鼠股四头肌细胞8-OHdG含量与其ROS水平呈明显正相关（r=0.746,P＝0.000）;经CSE刺激后,大鼠股四头肌细胞OGG1 mRNA和蛋白水平均高于对照组（P＜0.05）,但大鼠股四头肌细胞8-OHdG含量与其OGG1 mRNA和蛋白水平无明显相关（r=0.392,P＝0.536;r=0.297,P=0.426）.结论 CSE刺激可引起大鼠股四头肌细胞8-OHdG升高,8-OHdG含量与ROS水平有关,但与其修复酶OGG1表达水平无明显相关.

Study on oxidative damage of rat quadriceps femoris muscle induced by cigarette smoke extract

Objective To study the effect of cigarette smoke extract （CSE） on the quadriceps femoris muscle cells of rat in 8-hydroxydeoxyguanine （8-OHdG） content and the relationship between 8-OHdG content and level of reactive oxygen species （ROS） and 8-oxoguanine DNA glycosylase-1 （OGG1）.Methods The quadriceps femoris muscle cells of rats were cultured,which were stimulated by CSE according to different time and concentration.8-OHdG was detected by high press liquid chromatography with electrochemical detection （HPLC-ECD）.The level of ROS was determined by fluorescent probe of 2′,7′-dichlorodihydrofluorescein diacetate （DCFH-DA）.The expression of OGG1 mRNA was tested by Realtime PCR.The expression of OGG1 protein was tested by Western blot.Results There was non-significant difference between the 8-OHdG,ROS,OGG1 mRNA and protein level in the different time.However,the 8-OHdG content of the quadriceps femoris muscle cells induced by 10.00% CSE and 20.00% CSE was significantly increased in the quadriceps femoris muscle cells induced by 5.00% CSE and control group （P 〈0.05）.There were no significant differences in 8-OHdG content between 5.00% CSE and control group （P 〉0.05）.Compared with control,intracellular ROS level were dramatically increased in CSE-treated groups （P 〈 0.05）,and the 8-OHdG content was increased according to the level of ROS （r =0.746,P =0.000）.Compared with control,the expression of OGG1 mRNA and protein were dramatically increased in CSE-treated groups （P 〈0.05）,however,the 8-OHdG content had no correlation with the expression of OGG1 mRNA and protein （r =0.392,P =0.536; r =0.297,P =0.426）.Conclusions CSE could increase the level of 8-OHdG in the quadriceps femoris muscle cells of rats.The 8-OHdG content had correlation with ROS level,not with the expression of OGG1.