CIK分泌的细胞因子对逆转耐顺铂肺腺癌细胞系A549/DDP耐药性的研究

熊艳娟; 杜春娟; 于津浦; 曹水

doi:10.3969/j.issn.1000-8179.2013.12.001

CIK分泌的细胞因子对逆转耐顺铂肺腺癌细胞系A549/DDP耐药性的研究

Cytokines secreted by cytokine-induced killer cells modulate resistance to cisplatin in the A549/DDP cell line

摘要

摘要:
目的探讨细胞因子诱导的杀伤细胞(cytokine induced killer, CIK) 分泌的细胞因子在逆转耐顺铂(DDP) 人肺腺癌细胞系A549/DDP顺铂耐药性中的可能作用。
方法采用transwell非接触共培养CIK与A549/DDP细胞, 收集不同时间点的共培养上清, 酶联免疫试剂方法(ELISA) 检测IFN-γ、TNF-α、IL-2的分泌情况, 四氮甲唑兰比色法(MTT法) 检测DDP耐药性的变化, 实时定量PCR检测GST-π基因表达水平的变化。
结果分析发现A549/DDP细胞的耐药逆转与IFN-γ的分泌水平显著相关, 与TNF-α、IL-2的分泌量无显著相关。给予anti-IFN-γ中和抗体后, 共培养20h A549/DDP的DDP耐药性显著上升(P < 0.05), GST-π基因水平的表达量较未加中和抗体组也显著增加(P < 0.05)。而加入anti-TNF-α、anti-IL-2中和抗体组, A549/DDP细胞的DDP耐药性及GST-π基因水平的表达量较未加入中和抗体组无明显变化(P > 0.05)。
结论 CIK通过分泌IFN-γ下调A549/DDP细胞中GST-γ的表达来实现其逆转DDP耐药的作用。

Abstract:
Objective This study aimed to explore the possible roles of cytokines secreted by cytokine-induced killer (CIK) cells in reversing the DDP resistance of A549/DDP.
Methods Supernatants of co-cultured systems were collected to detect the levels of IFN-γ, TNF-α, and IL-2 using enzyme-linked immunosorbent assay (ELISA).The correlations between cytokines and resistance reversal indices were analyzed at different co-cultured time points.Specific neutralizing antibodies against IFN-γ, TNF-α, and IL-2 were added to the co-cultured system to evaluate the effects of the aforementioned cytokines on reversing DDP resistance in A549/DDP, as well as on the expression of GST-π gene.
Results ELISA and correlation analyses showed that IFN-γ was related to the reversal of DDP resistance of A549/DDP.After neutralization by anti-IFN-γ antibody, the DDP resistance of A549/DDP co-cultured with CIK for 20 h significantly recovered (P < 0.05) and the expression of GST-π at mRNA level significantly recovered compared with A549/DDP co-cultured without anti-IFN-γ antibody (P < 0.05).
Conclusion IFN-γ secreted by CIK might decrease the expression of GST-γ in A549/DDP to achieve reversal of DDP resistance.

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