抗氧化剂吡咯烷二硫代氨基甲酸有助于提高卡铂对宫颈癌SiHa细胞化疗的敏感性

Pyrrolidine dithiocarbamate helps improve the chemosensitivity of cervical cancer cell SiHa to carboplatin

目的探讨抗氧化剂吡咯烷二硫代氨基甲酸(PDTC)提高人宫颈癌SiHa细胞对卡铂化疗的敏感性及其可能的机制。方法体外培养SiHa细胞,分为空白组、PDTC组、卡铂组和PDTC+卡铂联合组。MTT检测细胞生长抑制率、流式细胞术检测细胞凋亡率和细胞周期、免疫细胞化学方法观察了NF-κBp65在细胞质和细胞核中的表达变化。结果 PDTC能够有效地抑制宫颈癌SiHa细胞的增殖,呈时间和剂量依赖关系;小剂量PDTC(12.5μmol/L)和卡铂联合应用与单用卡铂相比,能明显增加细胞生长抑制率(P<0.01)。流式细胞仪检测示,各药物组与对照组比较及PDTC+卡铂联用组与单用卡铂组比较宫颈癌SiHa细胞凋亡率的差异有显著性(P<0.01)。SiHa细胞经PDTC作用后G0/G1期比例较空白组明显增加,卡铂单用组使细胞周期阻滞于G2/M期,PDTC+卡铂组使细胞周期进一步阻滞于G2/M期,同时降低S期比例。免疫细胞化学显示,NF-κBp65在宫颈癌SiHa中主要表达于细胞质中,卡铂诱导24 h后细胞质中的NF-κBp65转移至细胞核,其活性增强,PDTC能够抑制此作用。结论卡铂能够诱导NF-κBp65的活化,小剂量PDTC(12.5μmol/L)和卡铂联合应用与单用卡铂相比,能明显增加细胞生长抑制率和细胞凋亡率,提高宫颈癌SiHa细胞对卡铂的敏感性。

Objective To study the effect of antioxidant pyrrolidine dithiocarbamate（PDTC） in promoting the chemosensitivity of human cervical cancer cell line SiHa to carboplatin and the possible mechanisms.Methods Human cervical carcinoma SiHa cell lines were cultured in vitro.They were treated with different concentrations of PDTC,carboplatin or PDTC combined with carboplatin for different durations.Then the inhibition of SiHa cells growth was detected by MTT assay;flow cytometry was used to analyze the apoptosis and cell cycle.The cellular distribution and protein level of NF-κBp65 were detected by immunocytochemical staining.Results PDTC could significantly inhibit the proliferation of SiHa cells in time-dependent and dose-dependent manners（P〈0.01）.Compared with carboplatin alone,PDTC（12.5 μmol/L） combined with carboplatin could significantly inhibit the proliferation of SiHa cells（P〈0.01）.The apoptosis rate of SiHa cell differed significantly between the drug group and the control group（P〈0.01）;the difference between PDTC ＋ carboplatin combination group and carboplatin alone group was significant（P0.01）.PDTC promoted G0/G1 phase of cell cycle arrest while carboplatin promoted G2/M phase of cell cycle arrest.Interestingly,G2/M phase was further arrested when carboplatin was added.The cells of S phase were also decreased.NF-κBp65 was mainly located in the cytoplasm of SiHa cell in the control group.The expression of NF-κBp65 in SiHa cell was transferred from cytoplasm to nucleus after 24-hour treatment with carboplatin,which increased the activity,whereas PDTC could inhibit the activity.Conclusion The activity of NF-κB p65 in SiHa cell was increased when treated with carboplatin.Even at a lower concentration,carboplatin combined with PDTC（12.5 μmol/L） resulted in a more significant growth inhibition and apoptosis of SiHa cell than carboplatin alone.PDTC can increase the chemosensitivity of cervical cancer SiHa cell to carboplatin.