抑制核转录因子活化逆转胃癌细胞株耐药性研究

Reversing chemoresistance to adriamycin through inhibiting activity of nuclear factor kappa B in human gastric cancer cell line

目的探讨核转录因子(NF-κB)的活化在胃癌细胞株耐药性机制中的作用。方法用阿霉素(ADM)诱导培养胃癌细胞耐药亚株SGC7901/ADM,MTT法检测ADM对胃癌细胞株的细胞毒作用,流式细胞仪检测胃癌细胞株凋亡率的变化,免疫细胞化学染色法检测胃癌细胞株NF-κB的活化情况。吡咯烷二硫代氨基甲酸脂(PTDC)抑制NF-κB活化,并检测ADM对胃癌细胞株的细胞毒作用。结果 SGC7901/ADM的IC50为2.63μg/ml,SGC7901的IC50为0.29μg/ml,SGC7901/ADM相对耐药度较亲本细胞提高了8.9倍。48h内SGC7901和SGC7901/ADM胃癌细胞株发生的凋亡率随ADM浓度增加而升高,随着作用时间延长而升高;10μg/mlADM分别作用48h时SGC7901和SGC7901/ADM胃癌细胞株发生的凋亡率为(48.53±1.02)%和(17.53±1.02)%。免疫细胞化学染色显示ADM作用SGC7901/ADM胃癌细胞9h后可检测到NF-κB核移位,最高达到65%;ADM作用SGC7901胃癌细胞株18h后可检测到少量NF-κB核移位;所有的细胞株NF-κB核移位均于24h后减弱;PTDC抑制核转录因子活化后ADM细胞毒效应增强(P<0.05)。结论 NF-κB活化所导致的胃癌细胞株凋亡率下降在胃癌细胞株耐药性机制中发挥一定作用,抑制NF-κB活化后可以逆转胃癌细胞株对ADM耐药性。

Objective To investigate the relationship between activity of nuclear factor kappa B（NF-κB） and chemoresistance in human gastric cancer cell line,and search for a new target of reversing chemoresistance in gastric cancer.Methods Human gastric cancer cell line resistant to adriamycin（SGC7901/ADM） was acquired by stepwise-increasing concentrations of adriamycin in vitro.The cytotoxicity of ADM on SGC7901/ADM and SGC7901 gastric cancer cell line was tested by MTT assay.The level of NF-κB activity was measured by immunohistochemical staining.The effects of the inhibition of inducible NF-κB activivity on chemoresistance against adriamycin by pyrrolidine dithiocarbamate（PDTC） were determined by MTT assay.Results IC50of SGC7901/ADM was 2.63 μg/ml and SGC7901＇s was 0.29μg/ml.Relative degree of resistance to adriamycin in SGC7901/ADM was improved 8.9 times than that in parental cells.In first 48 hours,the apoptosis rate of gastric cell line increased with adriamycin concentration increasing and time.The apoptosis rate of SGC7901 and SGC7901/ADM gastric cancer cell line induced by 10μg/ml adriamycin for 48 hours was（48.53±1.02）%and（17.53±1.02）% respectively.The activity of NF-κB in SGC7901/ADM gastric cancer cell line was detected 9 hours after adriamycin processing,the level of activation reached to 65%,which was significantly higher than that in SGC7901 gastric cancer cell line.After inhibiting inducible NF-κB activity by PDTC,the cytotoxicity of ADM on SGC7901/ADM gastric cancer cell line was increased significantly（P0.05）.Conclusion The change of apoptosis rate induced by NF-κB activity maybe play an important role in chemoresistance.Inhibiting activity of NF-κB can reduce chemoresistance to adriamycin in human gastric cancer cell line.NF-κB could be a new target of reversing chemoresistanc in gastric cancer.