白藜芦醇联合5-FU对宫颈癌HeLa细胞EMT转化的影响
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摘要
目的探索白藜芦醇联合5-FU对宫颈癌细胞系HeLa细胞EMT的影响。方法 MTT实验检测白藜芦醇、5-FU以及联合使用对HeLa细胞增殖的影响;Transwell小室迁移实验检测各组细胞迁移能力;免疫荧光实验检测各组细胞中Vimentin蛋白表达变化;Western bloting实验检测各组细胞中N-cadherin、E-cadherin、Twist1、Snail1蛋白表达变化;Elisa实验检测各组细胞上清中MMP-2、MMP-9的表达。结果与单独使用白藜芦醇或5-FU相比,联合使用后HeLa细胞的IC50明显降低,提高了HeLa细胞对5-FU的敏感性。进一步发现与二者单独使用相比,联合使用导致HeLa细胞迁移能力显著下降。在分子水平上,白藜芦醇联合5-FU诱导细胞中E-cadherin蛋白表达更高,Vimentin、N-cadherin、Twist1、Snail1、MMP-2、MMP-9蛋白表达更低。结论与单独使用白藜芦醇或5-FU相比,白藜芦醇联合5-FU对HeLa细胞迁移能力的抑制增强,二者联合增强效应的分子机制在于联合使用对HeLa细胞EMT具有更强的逆转能力。
Objective To investigate the effects of resveratrol combined with 5-FU on EMT transformation in cervical carcinoma cell line HeLa. Methods MTT assay was used to detect the effects of resveratrol, 5-FU and combination on the proliferation of HeLa cells. Transwell chamber migration assay was used to detect the migration ability of each group. Immunofluorescence assay was used to detect the expression of Vimentin protein in each group. The expressions of N-cadherin, E-cadherin, Twist1 and Snail1 in each group were detected by Western blotting(WB). The expressions of MMP-2 and MMP-9 in the supernatant of each group were detected by Elisa. Results Compared with resveratrol or 5-FU alone, the combined use of 5-FU significantly reduced the IC50 of HeLa cells and increased the sensitivity of HeLa cells to 5-FU. It was further found that the combined use resulted in a significant decrease in the migration ability of HeLa cells compared to the two alone. At the molecular level, resveratrol combined with 5-FU induced higher expression of E-cadherin protein, and Vimentin, N-cadherin, Twist1, Snail1, MMP-2, and MMP-9 proteins were lower. Conclusion The inhibition of HeLa cell migration ability is enhanced by resveratrol or 5-FU alone, and resveratrol combined with 5-FU. The molecular mechanism of the combined enhancement effect of the two is in combination with MDA-MB.-468 cell EMT transformation has a stronger reversal ability.
Objective To investigate the effects of resveratrol combined with 5-FU on EMT transformation in cervical carcinoma cell line HeLa. Methods MTT assay was used to detect the effects of resveratrol, 5-FU and combination on the proliferation of HeLa cells. Transwell chamber migration assay was used to detect the migration ability of each group. Immunofluorescence assay was used to detect the expression of Vimentin protein in each group. The expressions of N-cadherin, E-cadherin, Twist1 and Snail1 in each group were detected by Western blotting(WB). The expressions of MMP-2 and MMP-9 in the supernatant of each group were detected by Elisa. Results Compared with resveratrol or 5-FU alone, the combined use of 5-FU significantly reduced the IC50 of HeLa cells and increased the sensitivity of HeLa cells to 5-FU. It was further found that the combined use resulted in a significant decrease in the migration ability of HeLa cells compared to the two alone. At the molecular level, resveratrol combined with 5-FU induced higher expression of E-cadherin protein, and Vimentin, N-cadherin, Twist1, Snail1, MMP-2, and MMP-9 proteins were lower. Conclusion The inhibition of HeLa cell migration ability is enhanced by resveratrol or 5-FU alone, and resveratrol combined with 5-FU. The molecular mechanism of the combined enhancement effect of the two is in combination with MDA-MB.-468 cell EMT transformation has a stronger reversal ability.
引文
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[15] 李冬梅,张淑珍,车薇,等.白藜芦醇联合5-氟尿嘧啶抗肿瘤活性的研究 [J].实用药物与临床,2013,16(12):1130-1132.
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[18] 邓淑琴,张新毅.上皮细胞间质转化与肿瘤转移 [J].疾病监测与控制,2014,8(4):233-235.
[19] 马衣努尔·买提托合提.上皮间质化(EMT)在宫颈癌发生发展中诱导肿瘤干细胞的作用及其机制研究[D].华中科技大学,2013.
[20] 邢丹.E-cadherin、vimentin和ALDH1在乳腺癌中的表达及其与乳腺癌转移的相关性研究 [D].大连医科大学,2015.
[21] Tanaka K,Tokunaga E,Inoue Y,et al.Impact of expression of vimentin and axl in breast cancer [J].Clinical Breast Cancer,2016,16(6):520-526.
[22] Li M,Wu J,Zhang P.Research progress of application of E-cadherin,N-cadherin and vimentin in tumor metastasis [M].2016.
[23] Kumar S,Das A,Barai A,et al.MMP secretion rate and inter-invadopodia spacing collectively govern cancer invasiveness [J].Biophysical Journal,2018,114(3):650-662.
[24] Martin TA,Goyal A,Watkins G,et al.Expression of the transcription factors snail,slug,and twist and their clinical significance in human breast cancer [J].Annals of Surgical Oncology,2005,12(6):488-496.
[2] Sheng L,Zhong J,Liangyu,et al.Effect of resveratrol and in combination with 5-FU on murine liver cancer [J].World Journal of Gastroenterology,2004,10(20):3048-3052.
[3] 吴异兰,王晗.白藜芦醇对肺癌紫杉醇耐药细胞的增殖抑制作用及其机制 [J].癌变·畸变·突变,2018,30(3):200-203,20.
[4] 常环环,杨姝,李文帅,等.白藜芦醇与奥沙利铂联合应用抑制结直肠癌细胞增殖的机制 [J].解剖学杂志,2017,40(2):125-128.
[5] 高倩.白藜芦醇抗肿瘤作用机制研究进展 [J].中华实用诊断与治疗杂志,2016,30(9):845-847.
[6] 阿里木江·吾守,潘红芽.上皮—间充质转分化与转录因子 Snail、Slug、Twist 在口腔鳞状细胞癌发生、发展中的作用[J].中国口腔颌面外科杂志,2011,9(6):521-525.
[7] Moore MA,Baumann F,Foliaki S,et al.Cancer epidemiol-ogy in the pacific islands-past,present and future[J].Asi-an Pac J Cancer Prev,2010,11:99-106.
[8] Qiu Y,Liu Y,Liw H,et al.P2Y2 receptor promotes the migration and invasion of breast cancer cells via EMT-related genes snail and E-cadherin [J].Oncology Reports,2018,39(1):138-150.
[9] 申震,周颖,赵卫东,等.Vimentin及E-Cadherin在子宫颈癌中的表达及其与上皮细胞间质化的相关性[J].安徽医科大学学报,2013,48(5):505-508.
[10] 乔迪,宋静慧.Wnt/β-catenin信号通路激活与宫颈病变研究新进展[J].国际生殖健康/计划生育杂志,2016,35(6):510-514.
[11] Rafael D,Doktorovová S,Florindo H F,et al.EMT blockage strategies:Targeting Akt dependent mechanisms for breast cancer metastatic behaviour modulation[J].Current Gene Therapy,2015,15(3):300-312.
[12] 邱丽敏,姜爽,QIULI-MIN,等.天南星多糖联合顺铂对乳腺癌MDA-MB-231细胞增殖、凋亡及上皮间质转化的影响 [J].中药材,2016,39(3):630-633.
[13] Nezami M A,Hager S,Garner J.EMT and anti-EMT strategies in cancer [J].Journal of Cancer Therapy,2015,06(11):1013-1019.
[14] 李燕,邢克飞,李福洋,等.白藜芦醇联合5-FU对人乳腺癌细胞系MCF7的促凋亡作用 [J].医学争鸣,2006,27(7):658-661.
[15] 李冬梅,张淑珍,车薇,等.白藜芦醇联合5-氟尿嘧啶抗肿瘤活性的研究 [J].实用药物与临床,2013,16(12):1130-1132.
[16] Wang J,Liang WJ,Min GT,et al.LTBP2 promotes the migration and invasion of gastric cancer cells and predicts poor outcome of patients with gastric cancer [J].International Journal of Oncology,2018,52(6):1886-1898.
[17] 彭凯丽,陈宏辉.上皮间质细胞转化在肿瘤侵袭转移及化疗耐药中的作用 [J].实用医药杂志,2018,(1):85-87.
[18] 邓淑琴,张新毅.上皮细胞间质转化与肿瘤转移 [J].疾病监测与控制,2014,8(4):233-235.
[19] 马衣努尔·买提托合提.上皮间质化(EMT)在宫颈癌发生发展中诱导肿瘤干细胞的作用及其机制研究[D].华中科技大学,2013.
[20] 邢丹.E-cadherin、vimentin和ALDH1在乳腺癌中的表达及其与乳腺癌转移的相关性研究 [D].大连医科大学,2015.
[21] Tanaka K,Tokunaga E,Inoue Y,et al.Impact of expression of vimentin and axl in breast cancer [J].Clinical Breast Cancer,2016,16(6):520-526.
[22] Li M,Wu J,Zhang P.Research progress of application of E-cadherin,N-cadherin and vimentin in tumor metastasis [M].2016.
[23] Kumar S,Das A,Barai A,et al.MMP secretion rate and inter-invadopodia spacing collectively govern cancer invasiveness [J].Biophysical Journal,2018,114(3):650-662.
[24] Martin TA,Goyal A,Watkins G,et al.Expression of the transcription factors snail,slug,and twist and their clinical significance in human breast cancer [J].Annals of Surgical Oncology,2005,12(6):488-496.
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