GSDME通过调控细胞焦亡影响乳腺癌MCF-7细胞对紫杉醇的敏感性
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摘要
目的:探讨GSDME是否通过调控细胞焦亡影响乳腺癌MCF-7细胞对化疗药物紫杉醇(paclitaxel,PTX)的敏感性。方法:利用RNA干扰技术敲降GSDME在MCF-7细胞中的表达,采用CCK-8法、流式细胞术、乳酸脱氢酶(lactate dehydrogenase,LDH)释放实验及Wb技术分别检测GSDME低表达前后,PTX对细胞增殖、焦亡、LDH释放、GSDME-N端蛋白及cleaved-caspase-3蛋白表达水平的变化情况。结果:与对照组比较,PTX处理组细胞的焦亡率、LDH释放量、GSDME-N端蛋白及cleaved-caspase-3蛋白的表达水平均显著升高(均P<0.01);与si-NC组比较,敲低GSDME可使si-GSDME组细胞对PTX的敏感性降低,其细胞焦亡率、LDH释放量及GSDME-N端蛋白表达水平均显著下降(均P<0.01)。结论:敲减MCF-7细胞中GSDME的表达量可显著抑制细胞焦亡并降低细胞对PTX的敏感性。
Objective: To investigate whether GSDME affects the sensitivity of breast cancer MCF-7 cells to paclitaxel(PTX) by regulating cell pyroptosis. Methods: GSDME was knocked-down in MCF-7 cells by RNA interference technique. CCK-8 assay, flow cytometry,lactate dehydrogenase(LDH) release method and Wb were respectively used to detect cell proliferation, pyroptotic rate, LDH release, GSDME-N-terminal protein and cleaved-caspase-3 protein levels in PTX-treated MCF-7 cells before and after GSDME knockdown. Results: Compared with the control group, the pyroptotic rate, LDH release, GSDME-N-terminal protein and cleaved-caspase-3 protein levels in the PTX-treatment group significantly increased(all P<0.01). Compared with the si-NC group, the PTX-sensitivity of si-GSDME group decreased, and the pyroptotic rate, LDH release and GSDME-N-terminal protein all significantly decreased(all P<0.01). Conclusion: Knock-down of GSDME in MCF-7 cells significantly inhibited cell pyroptosis and reduced drug sensitivity of MCF-7 cells to PTX.
Objective: To investigate whether GSDME affects the sensitivity of breast cancer MCF-7 cells to paclitaxel(PTX) by regulating cell pyroptosis. Methods: GSDME was knocked-down in MCF-7 cells by RNA interference technique. CCK-8 assay, flow cytometry,lactate dehydrogenase(LDH) release method and Wb were respectively used to detect cell proliferation, pyroptotic rate, LDH release, GSDME-N-terminal protein and cleaved-caspase-3 protein levels in PTX-treated MCF-7 cells before and after GSDME knockdown. Results: Compared with the control group, the pyroptotic rate, LDH release, GSDME-N-terminal protein and cleaved-caspase-3 protein levels in the PTX-treatment group significantly increased(all P<0.01). Compared with the si-NC group, the PTX-sensitivity of si-GSDME group decreased, and the pyroptotic rate, LDH release and GSDME-N-terminal protein all significantly decreased(all P<0.01). Conclusion: Knock-down of GSDME in MCF-7 cells significantly inhibited cell pyroptosis and reduced drug sensitivity of MCF-7 cells to PTX.
引文
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[3]LIN Y,LIU Z,LI Y,et al.Short-term hyperthermia promotes the sensitivity of MCF-7 human breast cancer cells to paclitaxel[J].Biol Pharm Bull,2013,36(3):376-383.DOI:10.1248/bpb.b12-00774.
[4]KOMATSU M,WHEELER H E,CHUNG S,et al.Pharmacoethnicity in paclitaxel-induced sensory peripheral neuropathy[J].Clin Cancer Res,2015,21(19):4337-4346.DOI:10.1158/1078-0432.CCR-15-0133.
[5]MOHAMMADI A,MANSOORI B,AGHAPOUR M,et al.The urtica dioica extract enhances sensitivity of paclitaxel drug to MDA-MB-468 breast cancer cells[J/OL].Biomed Pharmacother,2016,83:835-842[2018-09-08].http://linkinghub.elsevier.com/retrieve/pii/s0753-3322(16)30840-x.DOI:10.1016/j.biopha.2016.07.056.
[6]GALLUZZI L,VITALE I,AQRONSON S A,et al.Molecular mechanisms of cell death:recommendations of the nomenclature committee on cell death 2018[J].Cell Death Differ,2018,25(3):486-541.DOI:10.1038/s41418-017-0012-4.
[7]WANG Y,GAO W,SHI X,et al.Chemotherapy drugs induce pyroptosis through caspase-3 cleavage of a gasdermin[J].Nature,2017,547(7661):99-103.DOI:10.1038/nature22393.
[8]ROGERS C,FERNANDES-ALBERMRI T,MAYES L,et al.Cleavage of DFNA5 by caspase-3 during apoptosis mediates progression to secondary necrotic/pyroptotic cell death[J/OL].Nat Commun,2017,8:14128[2018-09-08].https://www.nature.com/articles/ncomms14128.DOI:10.1038/ncomms14128.
[9]LU H,ZHANG S,WU J,et al.Molecular targeted therapies elicit concurrent apoptotic and GSDME-dependent pyroptotic tumor cell death[J].Clin Cancer Res,2018,24(23):1478.DOI:10.1158/1078-0432.CCR-18-1478.
[10]WANG Y,YIN B,LI D,et al.GSDME mediates caspase-3-dependent pyroptosis in gastric cancer[J].Biochem Biophys Res Commun,2018,495(1):1418-1425.DOI:10.1016/j.bbrc.2017.11.156.
[11]LAGE H,HELMBACH H,GROTTKE C,et al.DFNA5(ICERE-1)contributes to acquired etoposide resistance in melanoma cells[J].FEBS Lett,2001,494(1/2):54-59.DOI:10.1016/S0014-5793(01)02304-3.
[12]GUO Q,WU Y,HOU Y,et al.Cytokine secretion and pyroptosis of thyroid follicular cells mediated by enhanced NLRP3,NLRP1,NL-RC4,and AIM2 inflammasomes are associated with autoimmune thyroiditis[J].Front Immunol,2018,4(9):1197-1212.DOI:10.3389/fimmu.2018.01197.
[13]TANG M,LI X,LIU P,et al.Bradykinin B2 receptors play a neuroprotective role in Hypoxia/reoxygenation injury related to pyroptosis pathway[J].Curr Neurovasc Res,2018,15(2):138-144.DOI:10.2174/1567202615661805807341.
[14]路璐,王云凤,吕以东,等.miR-129-5p通过HMGB1调控乳腺癌MCF-7细胞对紫杉醇的敏感性[J].中国肿瘤生物治疗杂志,2018,25(1):62-67.DOI:10.3872/j.issn.1007-385x.2018.01.011.
[15]HE Y,AMER A O.Microbial modulation of host apoptosis and pyroptosis[J/OL].Front Cell Infect Microbiol,2014,4:83-87[2018-09-08].https://wwww.ncbi.nlm.nih.gov/pmc/articles/PMC4062964.DOI:10.3389/fcimb.2014.00083.
[16]SHI J,GAO W,SHAO F.Pyroptosis:gasdermin-mediated programmed necrotic cell death[J].Trends Biochem Sci,2017,42(4):245-254.DOI:10.1016/j.tibs.2016.1 0.004.
[17]KOVACS S B,MIAO E A.Gasdermins:effectors of pyroptosis[J].Trends Cell Biol,2017,27(9):673-684.DOI:10.1016/j.tcb.2017.05.005.
[18]YOSHIKO M,MANABU F,HIROKI K,et al.The potential role of DFNA5,a hearing impairment gene,in p53-mediated cellular response to DNA damage[J].J Hum Genet,2006,51(8):652-664.DOI:10.1007/s10038-006-0004-6.
[19]WNG C,TANG L,SHEN D W,et al.The expression and regulation of DFNA5 in human hepatocellular carcinoma DFNA5 in hepatocellular carcinoma[J].Mol Biol Rep,2013,40(12):6525-6531.DOI:10.1007/s11033-013-2581-8.