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

doi:10.3760/cma.j.issn.1673-422X.2016.05.001

Abstract

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

Zeng Kanghua, Ru Qin, Xiong Qi, Ai Yongxun. Effect of P2X7R agonist BzATP on cell growth and apoptosis in non-small cell lung cancer A549 cells[J]. Journal of International Oncology, 2016, 43(5): 321-325.

References

［1］ Teixeira JM, de OliveiraFusaro MC, Parada CA, et al. Peripheral P2X7 receptorinduced mechanical hyperalgesia is mediated by bradykinin［J］. Neuroscience, 2014, 277: 163-173. DOI: 10.1016/j.neuroscience.2014.06.057. ［2］ Jubb AM, Miller KD, Rugo HS, et al. Impact of exploratory biomarkers on the treatment effect of bevacizumab in metastatic breast cancer［J］. Clin Cancer Res, 2011, 17(2): 372-381. DOI: 10.1158/10780432.CCR101791. ［3］ Qiu Y, Li WH, Zhang HQ, et al. P2X7 mediates ATPdriven invasiveness in prostate cancer cells［J］. PLoS One, 2014, 9(12): e114371. DOI: 10.1371/ journal.pone.0114371. ［4］ Mello Pde A, FilippiChiela EC, Nascimento J, et al. Adenosine uptake is the major effector of extracellular ATP toxicity in human cervical cancer cells［J］. Mol Biol Cell, 2014, 25(19): 2905-2918. DOI: 10.1091/mbc.E14010042. ［5］ Künzli BM, Bernlochner MI, Rath S, et al. Impact of CD39 and purinergic signalling on the growth and metastasis of coloretal cancer［J］. Purinergic Signal, 2011, 7(2): 231-241. DOI: 10.1007/s1130201192289. ［6］ Gehring MP, Pereira TC, Zanin RF, et al. P2X7 receptor activation leads to increased cell death in a radiosensitive human glioma cell line［J］. Purinergic Signal, 2012, 8(4): 729-739. DOI: 10.1007/s1130201293192. ［7］ Gehring MP, Kipper F, Nicoletti NF, et al. P2X7 receptor as predictor gene for glioma radiosensitivity and median survival［J］. Int J Biochem Cell Biol, 2015, 68: 92-100. DOI: 10.1016/j.biocel.2015.09.001. ［8］ Xia J, Yu X, Tang L, et al. P2X7 receptor stimulates breast cancer invasion and migration via the AKT pathway［J］. Oncol Rep, 2015, 34(1): 103-110. DOI: 10.3892/or.2015.3979. ［9］ Takai E, Tsukimoto M, Harada H, et al. Autocrine regulation of TGFβ1induced cell migration by exocytosis of ATP and activation of P2 receptors in human lung cancer cells［J］. J Cell Sci, 2012, 125(Pt 21): 5051-5060. DOI: 10.1242/jcs.104976. ［10］韦星, 李朝敢, 李韬, 等. 白英生物碱通过抑制NFκB信号通路诱导人肺腺癌A549细胞凋亡［J］. 中药材, 2013, 36(5): 787-790. DOI: 10.13863/j.issn10014454.2013.05.029. ［11］ Zhang X, Meng L, He B, et al. The role of P2X7 receptor in ATPmediated human leukemia cell death: calcium influxindependent［J］. Acta Biochim Biophys Sin (Shanghai), 2009, 41(5): 362-369. ［12］ Nalluri S, GhoshalGupta S, Kutiyanawalla A, et al. TIMP1 inhibits apoptosis in lung adenocarcinoma cells via interaction with Bcl2［J］. PLoS One, 2015, 10(9): e0137673. DOI: 10.1371/journal.pone.0137673. ［13］ Li H, Ma G, Gui D, et al. Characterization of the porcine p65 subunit of NFκB and its association with virus antibody levels［J］. Mol Immunol, 2011, 48(6-7): 914-923. DOI: 10.1016/j.molimm.2010.12.015. ［14］Scheinman R. NFκB and rheumatoid arthritis: will understanding genetic risk lead to a therapeuti reward?［J］. For Immunopathol Dis Therap, 2013, 4(2): 93-110.

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