2,2′,4,4′-四溴联苯醚对人乳腺癌细胞MCF-7细胞生物学行为影响
|
摘要
目的乳腺癌是一种激素依赖性肿瘤,生殖模式的改变和环境雌激素暴露增加导致女性乳腺癌发病率不断增高。本研究旨在探讨环境中广泛存在的内分泌干扰物2,2′,4,4′-四溴联苯醚(2,2′,4,4′-tetrabromodiphenyl ethers,BDE-47)对乳腺癌细胞MCF-7增殖、迁移和侵袭等细胞生物学行为的影响及可能机制。方法体外培养MCF-7细胞,以梯度浓度BDE-47分别处理细胞,采用MTT法和Transwell实验分别检测细胞增殖及迁移侵袭能力,并通过蛋白质印迹法检测间质上皮转化(epithelial-to-mesenchymal transition,EMT)相关标志物和β-catenin的蛋白表达水平。结果与无BDE-47暴露亲代细胞株比较,MTT实验结果显示,BDE-47暴露<72h对细胞无明显促增殖作用,F=2.002,P=0.233。迁移试验结果显示,2.5、5和10μmol/L浓度组与对照组比较,迁移细胞数差异均有统计学意义,t值分别为-12.901、-5.250和-12.799,P值分别为<0.001、0.006和0.001。侵袭实验结果显示,2.5、5和10μmol/L浓度BDE-47处理组与对照组比较,侵袭细胞数差异均有统计学意义,t值分别为-4.281、-10.056和-12.331,P值分别为0.013、0.001和<0.001;且均呈浓度依赖性。蛋白质印迹法结果表明,不同浓度BDE-47处理48h后,MCF-7细胞E-cadherin蛋白表达均下降,N-cadherin、Vimentin和β-catenin表达上调。结论 BDE-47暴露对乳腺癌细胞MCF-7细胞增殖无明显影响,可能通过β-catenin信号通路诱导细胞EMT促进乳腺癌细胞MCF-7的迁移和侵袭。
OBJECTIVE Breast cancer is a hormone-dependent tumor.Changes in reproductive patterns and increase of environmental estrogen exposure cause the continued increasing incidence of breast cancer.The relationship between breast cancer and 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47),an ubiquitous endocrine disruptor in environment,remains unclear.The present study was to explore the effects of BDE-47 on the cell biological behavior of breast cancer cells MCF-7 including cell proliferation,invasion and migration in vitro,and the underlying mechanisms as well.METHODS MCF-7 cells were treated with serial dilution of BDE-47.MTT assay and transwell assays were used to evaluate the proliferation,invasion and migration ability.Meanwhile,the protein levels of biomarkers of epithelial-to-mesenchymal transition(EMT)andβ-catenin were assessed by Western blot.RESUITS BDE-47 treatment showed no obvious affects on cell proliferation of MCF-7 in 72 h(F=2.002,P=0.233).Compared with the parental cells,BDE-47-treated MCF-7 cells showed an significant increase in the number of migrated cells among gradient concentration groups(2.5,5,10μmol/L;t=-12.901,P<0.001;t=-5.250,P=0.006;t=-12.799,P=0.001)as well as in invaded cell number among serial doses of BDE-47(t=-4.281,P=0.013;t=-10.056,P=0.001;t=-12.331,P<0.001).BDE-47 promoted the migration and invasion of MCF-7 cells in a concentration-dependent manner.The results of Western blot revealed that BDE-47 treatment decreased E-cadherin expression but increased protein levels of N-cadherin,Vimentin andβ-catenin.CONCLUSION BDE-47 has no effects on cell proliferation of MCF-7 cells while promotes the cell migration and invasion by EMT mechanisms viaβ-catenin signaling pathway.
OBJECTIVE Breast cancer is a hormone-dependent tumor.Changes in reproductive patterns and increase of environmental estrogen exposure cause the continued increasing incidence of breast cancer.The relationship between breast cancer and 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47),an ubiquitous endocrine disruptor in environment,remains unclear.The present study was to explore the effects of BDE-47 on the cell biological behavior of breast cancer cells MCF-7 including cell proliferation,invasion and migration in vitro,and the underlying mechanisms as well.METHODS MCF-7 cells were treated with serial dilution of BDE-47.MTT assay and transwell assays were used to evaluate the proliferation,invasion and migration ability.Meanwhile,the protein levels of biomarkers of epithelial-to-mesenchymal transition(EMT)andβ-catenin were assessed by Western blot.RESUITS BDE-47 treatment showed no obvious affects on cell proliferation of MCF-7 in 72 h(F=2.002,P=0.233).Compared with the parental cells,BDE-47-treated MCF-7 cells showed an significant increase in the number of migrated cells among gradient concentration groups(2.5,5,10μmol/L;t=-12.901,P<0.001;t=-5.250,P=0.006;t=-12.799,P=0.001)as well as in invaded cell number among serial doses of BDE-47(t=-4.281,P=0.013;t=-10.056,P=0.001;t=-12.331,P<0.001).BDE-47 promoted the migration and invasion of MCF-7 cells in a concentration-dependent manner.The results of Western blot revealed that BDE-47 treatment decreased E-cadherin expression but increased protein levels of N-cadherin,Vimentin andβ-catenin.CONCLUSION BDE-47 has no effects on cell proliferation of MCF-7 cells while promotes the cell migration and invasion by EMT mechanisms viaβ-catenin signaling pathway.
引文
[1]Sweeney MF,Hasan N,Soto AM,et al.Environmental endocrine disruptors:Effects on the human male reproductive system[J].Rev Endocr Metab Disord,2015,16(4):341-357.
[2]Zhang J,Chen L,Xiao L et al.Polybrominated diphenyl ether concentrations in human breast milk specimens worldwide[J].Epidemiology,2017,28(Suppl 1):S89-S97.
[3]Sjodin A,Patterson DG,Jr.Bergman A.A review on human exposure to brominated flame retardants-particularly polybrominated diphenyl ethers[J].Environ Int,2003,29(6):829-839.
[4]Barber JL,Walsh MJ,Hewitt R,et al.Low-dose treatment with polybrominated diphenyl ethers(PBDEs)induce altered characteristics in MCF-7cells[J].Mutagenesis,2006,21(5):351-360.
[5]Sjodin A,Jones RS,Caudill SP,et al.Polybrominated diphenyl ethers,polychlorinated biphenyls,and persistent pesticides in serum from the national health and nutrition examination survey:2003-2008[J].Environ Sci Technol,2014,48(1):753-760.
[6]Bergman A,Heindel JJ,Kasten T,et al.The impact of endocrine disruption:a consensus statement on the state of the science[J].Environ Health Perspect,2013,121(4):A104-A106.
[7]翟金霞.多溴联苯醚的健康效应研究进展[J].中华预防医学杂志,2016,50(6):559-562.
[8]Huang Y,Zhu G,Peng L,et al.Effect of 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47)on sexual behaviors and reproductive function in male zebrafish(Danio rerio)[J].Ecotoxicol Environ Saf,2015,111:102-108.
[9]Hurley S,Reynolds P,Goldberg D,et al.Adipose levels of polybrominated diphenyl ethers and risk of breast cancer[J].Breast Cancer Res Treat,2011,129(2):505-511.
[10]Holmes AK,Koller KR,Kieszak SM,et al.Case-control study of breast cancer and exposure to synthetic environmental chemicals among Alaska Native women[J].Int J Circumpolar Health,2014,73:25760.
[11]Karpeta A,Maniecka A,Gregoraszczuk EL.Different mechanisms of action of 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47)and its metabolites(5-OH-BDE-47and 6-OH-BDE-47)on cell proliferation in OVCAR-3ovarian cancer cells and MCF-7breast cancer cells[J].JAppl Toxicol,2016,36(12):1558-1567.
[12]Kwiecinska P,Wrobel A,Gregoraszczuk EL.Combinatory effects of PBDEs and 17beta-estradiol on MCF-7cell proliferation and apoptosis[J].Pharmacol Rep,2011,63(1):189-194.
[13]Tian PC,Wang HL,Chen GH,et al.2,2′,4,4′-Tetrabromodiphenyl ether promotes human neuroblastoma SH-SY5Ycells migration via the GPER/PI3K/Akt signal pathway[J].Hum Exp Toxicol,2016,35(2):124-134.
[14]Felipe Lima J,Nofech-Mozes S,Bayani J,et al.EMT in Breast Carcinoma-A Review[J].J Clin Med,2016,5(7).doi:10.3390/jcm5070065.
[15]Schramm HM.Should EMT of cancer cells be understood as epithelial-myeloid transition[J].J Cancer,2014,5(2):125-132.
[16]Anastas JN,Moon RT.WNT signalling pathways as therapeutic targets in cancer[J].Nat Rev Cancer,2013,13(1):11-26.
[17]Yang X,Li L,Huang Q,et al.Wnt signaling through Snail1and Zeb1regulates bone metastasis in lung cancer[J].Am J Cancer Res,2015,5(2):748-755.
[18]Novellasdemunt L,Antas P,Li VS.Targeting Wnt signaling in colorectal cancer.A review in the Theme:Cell signaling:proteins,pathways and mechanisms[J].Am J Physiol Cell Physiol,2015,309(8):C511-521.
[19]赵鹏,刘冬,张卉,等.RBMS3通过Wnt/β-catenin信号通路抑制胃癌细胞侵袭及其上皮-间质转化[J].肿瘤,2017,37(10):1032-1040.
[2]Zhang J,Chen L,Xiao L et al.Polybrominated diphenyl ether concentrations in human breast milk specimens worldwide[J].Epidemiology,2017,28(Suppl 1):S89-S97.
[3]Sjodin A,Patterson DG,Jr.Bergman A.A review on human exposure to brominated flame retardants-particularly polybrominated diphenyl ethers[J].Environ Int,2003,29(6):829-839.
[4]Barber JL,Walsh MJ,Hewitt R,et al.Low-dose treatment with polybrominated diphenyl ethers(PBDEs)induce altered characteristics in MCF-7cells[J].Mutagenesis,2006,21(5):351-360.
[5]Sjodin A,Jones RS,Caudill SP,et al.Polybrominated diphenyl ethers,polychlorinated biphenyls,and persistent pesticides in serum from the national health and nutrition examination survey:2003-2008[J].Environ Sci Technol,2014,48(1):753-760.
[6]Bergman A,Heindel JJ,Kasten T,et al.The impact of endocrine disruption:a consensus statement on the state of the science[J].Environ Health Perspect,2013,121(4):A104-A106.
[7]翟金霞.多溴联苯醚的健康效应研究进展[J].中华预防医学杂志,2016,50(6):559-562.
[8]Huang Y,Zhu G,Peng L,et al.Effect of 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47)on sexual behaviors and reproductive function in male zebrafish(Danio rerio)[J].Ecotoxicol Environ Saf,2015,111:102-108.
[9]Hurley S,Reynolds P,Goldberg D,et al.Adipose levels of polybrominated diphenyl ethers and risk of breast cancer[J].Breast Cancer Res Treat,2011,129(2):505-511.
[10]Holmes AK,Koller KR,Kieszak SM,et al.Case-control study of breast cancer and exposure to synthetic environmental chemicals among Alaska Native women[J].Int J Circumpolar Health,2014,73:25760.
[11]Karpeta A,Maniecka A,Gregoraszczuk EL.Different mechanisms of action of 2,2′,4,4′-tetrabromodiphenyl ether(BDE-47)and its metabolites(5-OH-BDE-47and 6-OH-BDE-47)on cell proliferation in OVCAR-3ovarian cancer cells and MCF-7breast cancer cells[J].JAppl Toxicol,2016,36(12):1558-1567.
[12]Kwiecinska P,Wrobel A,Gregoraszczuk EL.Combinatory effects of PBDEs and 17beta-estradiol on MCF-7cell proliferation and apoptosis[J].Pharmacol Rep,2011,63(1):189-194.
[13]Tian PC,Wang HL,Chen GH,et al.2,2′,4,4′-Tetrabromodiphenyl ether promotes human neuroblastoma SH-SY5Ycells migration via the GPER/PI3K/Akt signal pathway[J].Hum Exp Toxicol,2016,35(2):124-134.
[14]Felipe Lima J,Nofech-Mozes S,Bayani J,et al.EMT in Breast Carcinoma-A Review[J].J Clin Med,2016,5(7).doi:10.3390/jcm5070065.
[15]Schramm HM.Should EMT of cancer cells be understood as epithelial-myeloid transition[J].J Cancer,2014,5(2):125-132.
[16]Anastas JN,Moon RT.WNT signalling pathways as therapeutic targets in cancer[J].Nat Rev Cancer,2013,13(1):11-26.
[17]Yang X,Li L,Huang Q,et al.Wnt signaling through Snail1and Zeb1regulates bone metastasis in lung cancer[J].Am J Cancer Res,2015,5(2):748-755.
[18]Novellasdemunt L,Antas P,Li VS.Targeting Wnt signaling in colorectal cancer.A review in the Theme:Cell signaling:proteins,pathways and mechanisms[J].Am J Physiol Cell Physiol,2015,309(8):C511-521.
[19]赵鹏,刘冬,张卉,等.RBMS3通过Wnt/β-catenin信号通路抑制胃癌细胞侵袭及其上皮-间质转化[J].肿瘤,2017,37(10):1032-1040.