国际肿瘤学杂志

国际肿瘤学杂志 ›› 2016, Vol. 43 ›› Issue (5): 321-325.doi: 10.3760/cma.j.issn.1673-422X.2016.05.001

• 论著 •    下一篇

P2X7R激动剂BzATP对非小细胞肺癌A549细胞生长和凋亡的影响

曾康华,茹琴,熊琪,艾永循   

  1. 430056 江汉大学武汉生物医学研究院（曾康华、茹琴、熊琪），医学院（艾永循）
  • 出版日期:2016-05-08 发布日期:2016-04-07
  • 通讯作者: 艾永循 E-mail:iyx548@aliyun.com
  • 基金资助:

    国家自然科学基金（81302203）

Effect of P2X7R agonist BzATP on cell growth and apoptosis in non-small cell lung cancer A549 cells

Zeng Kanghua, Ru Qin, Xiong Qi, Ai Yongxun   

  1. Wuhan Institute of Biomedical Sciences, Jianghan University, Wuhan 430056, China
  • Online:2016-05-08 Published:2016-04-07
  • Contact: Ai Yongxun E-mail:iyx548@aliyun.com
  • Supported by:

    National Natural Science Foundation of China (81302203)

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摘要： 目的研究配体门控离子通道P2X7受体（P2X7R）在非小细胞肺癌A549细胞中的表达，观察P2X7R激动剂2′3′O（4苯甲酰苯甲酰）腺苷三磷酸三乙烷胺盐（BzATP）对A549细胞生长及凋亡的影响，并探究相关作用机制。方法采用免疫荧光法检测P2X7R在A549细胞中的表达。用不同浓度的BzATP（150、300、600 μmol/L）处理，未用BzATP干预的细胞作为对照组。采用四甲基偶氮唑蓝（MTT）和Hoest33342染色法分别检测细胞存活率与凋亡情况，酶联免疫吸附试验检测上清液中肿瘤坏死因子-α（TNF-α）的浓度，Western blotting检测核转录因子-κB（NF-κB） p65、NF-κB抑制因子α（IκBα）及磷酸化NF-κB抑制因子α（phospho-IκBα）蛋白的表达。结果P2X7R在A549细胞膜上表达。在300、600 μmol/L BzATP作用下A549细胞存活率分别为(67.87±8.98)%、(44.73±6.92)%，较对照组(98.60±1.44)%明显下降，差异均具有统计学意义（t=4.481，P=0.027；t=3.920，P=0.038）。BzATP可促进细胞凋亡，并且300、600 μmol/L BzATP可上调细胞培养上清中TNF-α浓度，分别为(57.35±6.41)pg/ml、(78.63±11.33)pg/ml，与对照组(42.56±0.37)pg/ml比较差异具有统计学意义（t=6.410，P=0.035；t=11.330,P=0.005）。此外，BzATP可下调NF-κB p65的表达，上调IκBα的表达，对phospho-IκBα的表达无明显作用。结论P2X7R表达于A549细胞膜，BzATP能够抑制细胞增殖，促进细胞凋亡，其作用机制可能与促进细胞中TNF-α的释放，抑制NF-κB通路有关。

关键词: 癌, 非小细胞肺, 细胞凋亡, P2X7受体, 2&prime, 3&prime, O(4-苯甲酰-苯甲酰)腺苷三磷酸三乙烷胺盐

Abstract: ObjectiveTo investigate the expression of P2X7 receptor (P2X7R) and the effect of P2X7R agonist 2′3′O(4benzoylbenzoyl) ethane adenosine triphosphate three amine salt (BzATP) on cell growth and apoptosis in nonsmall cell lung cancer A549 cells, and to explore the related mechanism. MethodsThe expression of P2X7R in A549 cells was detected by immunofluorescence. Cells were treated with different concentrations (150, 300, 600 μmol/L) of BzATP. Cells untreated with BzATP were used as control group. 3(4,5dimethyl2thiazoly)2,5diphenyl2Htetrazolium bromide (MTT) assay and Hoest33342 staining were respectively used to detect cell viability and apoptosis. Enzymelinked immunosorbent assay (ELISA) was uesd to detect the concentration of tumor necrosis factorα (TNFα) of cell culture supernatants. The expressions of nuclear factorκB (NFκB) p65, inhibitor of α of NFκB (IκBα) and the phosphorylation of inhibitor of α of NFκB (phosphoIκBα) were detected by Western blotting. ResultsP2X7R was expressed on the cell membrane of A549 cells. Survival rate of A549 cell was significantly decreased with the concentrations of BzATP at 300 and 600 μmol/L ［(67.87±8.98)%, (44.73±6.92)%］, compared with the control group (98.60±1.44)%, the differences were statistically significant (t=4.481, P=0.027; t=3.920, P=0.038). BzATP promoted apoptosis, and increased the concentration of TNFα of supernatant at 300 and 600 μmol/L ［(57.35±6.41)pg/ml, (78.63±11.33)pg/ml］, compared with the control group (42.56±0.37)pg/ml, the differences were statistically significant (t=6.410, P=0.035; t=11.330, P=0.005). In addation, the expressions of NFκB p65 and IκBα were respectively downregulated and upregulated by BzATP, while the expression of phosphoIκBα was not significantly altered. ConclusionP2X7R is expressed on A549 cell membrane. BzATP can inhibit cell proliferation and induce the apoptosis of A549 cells, and the mechanism of action may be related to promoting the release of TNFα and inhibition of NFκB pathway.

Key words: Carcinoma, nonsmallcell lung, Apoptosis, P2X7 receptor, 2′3′O(4benzoylbenzoyl) ethane adenosine triphosphate three amine salt