红霉素联合顺钳对A549细胞的细胞周期和凋亡的影响  

Study of erythromycin and cisplatin on cell cycle distributions and apoptosis in A549 cells exposed to cigarette smoke extract

作　　者：甘罗曼 黄斯明[2] 李文涛[3] 柳广南[3] Gan Luonian;Huang Siming;Li Wentao;Liu Guannan(Medical College of Qiunghai University,Qinghai 810016,China;Department of Respiratory Medicine,the First Affiliated Hospital of Guangxi Medical University,Nanning 530021,China;Department of Respiratory Medicine,the Second Affiliated Hospital of Guangxi Medical University,Nanning 530007,China)

机构地区：[1]青海大学医学院,810016 [2]广西医科大学第一附属医院呼吸内科,南宁530021 [3]广西医科大学第二附属医院呼吸内科,南宁530007

出　　处：《国际呼吸杂志》2019年第4期241-248,共8页International Journal of Respiration

基　　金：国家自然科学基金(81360013);广西高校中青年教师基础能力提升项目(2018KY0135).

摘　　要：目的研究红霉素(EM)对顺钳(CDDP)诱导A549细胞周期的细胞凋亡作用的影响。方法以A549细胞为研究对象.配制CDDP浓度为0.25、0.5、1、2,4、8 mg/1。用不同浓度CDDP刺激A549细胞48 h.采用Cell Counting Kit-8法检测不同浓度CDDP处理A549细胞后的细胞活性,并分别计算CDDP各自的半数抑制浓度(IC50)。将试验分组为空白对照组(只加10%灭活胎牛血清的RPMI-1640细胞培养液〉、10 mg/L EM组、10 mg/1.EM+IC50-CDDP组、IC50-CDDP组.各组细胞培养48 h后,流式细胞仪检测各组细胞的凋亡、细胞周期百分比。采用实时荧光定量聚合酶链反应检测HDAC2,Caspase^9、Caspask3、cyclinDl mRNA的表达;采用蛋白质印迹法检测HDAC2、Caspase-9,Caspase-3,cyclinDl蛋白的表达。结果不同浓度CDDP处理A549细胞48h,细胞生长受到不同程度抑制,并且呈浓度抑制依赖性.浓度越高.对细胞生长抑制作用越大(P<0.05)。IC50-CDDP为3.419 mg/L。流式细胞仪细胞周期结果显示:与空白对照组比较.其它3组细胞周期期比列均增加.差异均有统计学意义(P值均<0.05),细胞凋亡结果显示:与空白对照组比较.10 mg/L EM组的细胞凋亡有增加,但差异无统计学意义(P>0.05),而10 mg/L EM+IC50-CDDP组及IC50-CDDP组的细胞凋亡明显增加(P值均<0.05).与空白对照组比较,10 mg I.EM组及10 mg/L EM+IC50-CDDP组HDAC2、Caspase-9>Caspasr3 mRNA及蛋白表达均升高,cyclinDl mRNA及蛋白的表达均下降,差异均有统计学意义(P值均<0.05).结论EM可使A549细胞的cyclinDl的活性下降.增强CDDP对A549细胞的细胞阻滞作用。EM还可促进凋亡相关蛋白Caspase-9,Caspase3表达增加,增强CDDP对A549细胞的凋亡作用.其机制可能与EM提高A549细胞的HDAC2表达有关。因此.可考虑将EM联合CDDP作为治疗肺腺癌患者的新型药物组合。Objective To study the apoptosis of lung adenocarcinoma cell(A549)treated with erythromycin(EM)and cisplatin(CDDP).Methods Taking A549 cells as research object,differential concentrations of CDDP were prepared as 0.25,0.5,1,2.4,8 mg/L.respectively,Stimulated activity of A549 cells treated with different concentrations of CDDP for 48 h.The half inhibition concentration of CDDP(IC50)calculated by Cell Counting Kii-8 method.The research was divided as the blank control group(only 10%inactivation of fetal boxine serum RPMI-1640 cell culture medium),the 10 mg/L EM group,the 10 mg/L EM+IC50-CDDP group and the IC50-CDDP group.They were cultured for 48 h,then their apoptosis and cell cycle percentage were detected by flow cytometry.Real time qualitative PCR and Western blot were used to detect the expression of HDAC2,Caspase-9,Caspase-3,cyclinDl mRNA and proteins.Results A549 cells growth was inhibited by differential concentrations of CDDP in 48 h,and the inhibitory effect was concentration-dependent,the higher the concentration,the greater the inhibitory effect of cells growth(P<C0.05).The concentration of IC50-CDDP was 3.449 mg/L.Compared with the blank control group,the flow cytometry showed that the cells G(G[phase of the cell cycle in other three groups increased significantly(all P V0.05).And the results showed that the cells apoptosis of A549 were incresed in lOmg/L EM group compared with the blank control group,but the difference was not statistically significant(P>0.05).The cells apoptosis of A549 were obviously incresed in 10 mg/L EM+IC50-CDDP group and IC50-CDDP gruop(both P<C 0.05).Compared with the blank control group,the expression levels of HDAC2,Caspase-9,Caspase-3,cyclinDl mRNA and proteins were increasd in 10 mg/L EM group and 10 mg/L EM+IC50-CDDP group,but the expression level of cyclinDl mRNA and protein were decreased,the differences were statistically significant(all P<C0.05).Conclusions EM can decrease cyclinDl activity and enhance the blocking effect of CDDP on A549 cells.It also increases the ex

关 键 词：红霉素 顺钳 细胞周期 细胞凋亡 

分 类 号：R734.2[医药卫生—肿瘤]