香烟烟雾提取物对鼠成肌细胞分化的影响的研究

Explore the Effect of Cigarette Smoke Extract on Mouse Myoblast Cell Line Differentiation

目的:为了探讨COPD患者周围骨骼肌萎缩的可能的分子机制,用香烟烟雾的提取物(CSE)对鼠成肌细胞系(C2C12)进行了体外实验研究。方法:把C2C12细胞按150个/ml的浓度接种在12孔的平底塑料盘里,分为对照组和实验组,24小时后,在实验组的细胞中加入不同浓度(1%,2.5%,3.5%,5%)的CSE.我们把加入CSE的当天记录为第0天(day 0),第二天记录为第1天(day1),依次类推。CSE作用24小时后,用温的杜氏磷酸缓冲液(PBS)洗涤两次后换为10%马血清细胞分化生长液使细胞分化,而对照组更换新鲜的细胞生长液即可。并观察细胞的变化,并在加入CSE后的第0、3、6天,分别检测各组的肌酸激酶(CK)活性,并用激光扫描共焦荧光显微镜技术,观察各组的肌球蛋白重链(MHC)的生成情况。结果:重复相同实验4次后,对各组数值进行单因素方差分析的统计学处理,结果发现1%,2.5%组与对照组比较无显著差异(P>0.05);3.5%实验组与对照组比较day3有显著差异(P<0.05),day6无显著差异(P>0.05);5%实验组与对照组比较,day3,day6均有显著差异(P<0.05);并且激光扫描共焦荧光显徽镜技术观察对照组和实验组(3.5%)的MHC的照片显示CSE对MHC的形成有影响。结论:体外研究显示,CSE对C2C12的分化有影响。

Objective： In order to explore the possible molecule mechanisms of the peripheral skeletal muscle wasting and dysfunction in the patients with chronic obstruct pulmonary disease（COPD） ;we observed the effect of cigarette smoke extract （CSE） on mouse myoblast cell line （C2C12） differentiation in vitro. Methods： Seeded 150 cell/ml cells in 12 wells plate with grow media, divided in two groups： control group and treatment group. After 24 hours, added different concentration CSE （1% ,2.5% ,3.5% ,5% ）in treatment group,and added grow media in control group, we say today as day0 ,next was dayl and so on. After 24 hours,for the treatment group,threw away the old media,use warm PBS washed twice then changed 10% horse serum differentiation media;for the control group,threw away old media, then changed new grow media. Tested the croatine kinase （CK） value at day0 day3 day6 ,and use eonfocal microscopy oberved the Myosin Heavy Chain （MHC）. Resuits： After repeated the same experiment 4 times, All values are expressed as mean ＋ standard error of the mean （ SEM） Comparisons of ex- perimental values for CSE-treated cells to untreated controls were done by analysis of variance （ ANOVA ）. Compared with control group, there was no significant in 1% ,2.5 % groups （ P 〉 0. 05 ） ;3.5 % group has significant （ P 〈 0. 05 ） at day3 and no significant （ P 〉 0.05 ） at day6 ;5 % group had significant at day3 and day6 compared with control group. There was effect of CSE on MHC with confocal microscopy compared with two groups. Conclusions： There ws effect of CSE on C2C12 differentiation.