二氢杨梅素对绒毛膜癌细胞增殖和迁移能力的影响
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摘要
目的探讨二氢杨梅素(DMY)对绒毛膜癌(绒癌)JEG-3及JAR细胞增殖和迁移能力的影响。方法 MTT法检测不同浓度的二氢杨梅素(0 mg/L, 20 mg/L, 40 mg/L, 60 mg/L, 80 mg/L)作用一定时间后,对绒癌JEG-3和JAR细胞增殖能力的影响;细胞划痕实验和Transwell法检测不同浓度二氢杨梅素(0 mg/L, 40 mg/L, 60 mg/L, 80 mg/L)分别作用绒癌JEG-3细胞及JAR细胞一定时间后,对其迁移能力的影响;Real-time PCR和Western blotting方法分别检测不同浓度二氢杨梅素(0 mg/L, 40 mg/L, 60 mg/L, 80 mg/L)作用绒癌JEG-3及JAR细胞后,基质金属蛋白酶2(MMP-2)mRNA和蛋白表达水平的影响。结果不同浓度二氢杨梅素作用绒癌JEG-3和JAR细胞24 h和48 h后,随着二氢杨梅素浓度增加,对JEG-3和JAR细胞增殖抑制作用增强(P<0.05)。二氢杨梅素作用绒癌JEG-3及JAR细胞后,显著抑制细胞迁移能力,且具有浓度依赖性(P<0.05)。不同浓度二氢杨梅素作用JEG-3和JAR细胞后,MMP-2的mRNA和蛋白表达水平明显下降(P<0.05)。结论二氢杨梅素能够抑制JEG-3及JAR细胞的增殖能力且具有浓度依赖性,同时二氢杨梅素可能通过下调绒癌JEG-3及JAR细胞中MMP-2 mRNA和蛋白的表达,抑制绒癌细胞的侵袭迁移。
Objective To investigate the effect of dihydromyricetin(DMY)on proliferation and migration in choriocarcinoma JEG-3 and JAR cells. Methods JEG-3 and JAR cells were treated with different concentrations(0 mg/L, 20 mg/L, 40 mg/L, 60 mg/L, 80 mg/L) of DMY for 24 hours and 48 hours, and the proliferation was analyzed by methylthio tetrazole(MTT) assay. The effect of DMY on migration was detected by wound healing(after 24 hours) and Transwell assay(after treated JEG-3 for 36 hours and JAR for 24 hours). The expression of matrix metalloproteinase 2(MMP-2) at mRNA and protein levels with different concentrations of DMY(0 mg/L, 40 mg/L, 60 mg/L, 80 mg/L) were detected by Real-time PCR(after 12 hours,24 hours,36 hours,48 hours)and Western blotting(after treated 36 hours) respectively. Results The proliferation of JEG-3 and JAR cells was inhibited significantly(P<0.05), after DMY treatment for 24 hours and 48 hours.DMY inhibited the migration of JEG-3 and JAR cells significantly(P<0.05) with a dose-dependent manner. After JEG-3 and JAR Cells treated with DMY for 36 hours and 48 hours, the expression of MMP-2 mRNA decreased significantly(P<0.05),there were no significantly changes in DMY treatment with 12 hours and 24 hours. The expression level of MMP-2 protein was inhibited significantly after treatment with DMY for 36 hours(P<0.05). Conclusion DMY might inhibit the proliferation in choriocarcinoma JEG-3 and JAR cells with a dose-dependent manner. The invasion and migration were inhibited by DMY through downregulation of MMP-2.
Objective To investigate the effect of dihydromyricetin(DMY)on proliferation and migration in choriocarcinoma JEG-3 and JAR cells. Methods JEG-3 and JAR cells were treated with different concentrations(0 mg/L, 20 mg/L, 40 mg/L, 60 mg/L, 80 mg/L) of DMY for 24 hours and 48 hours, and the proliferation was analyzed by methylthio tetrazole(MTT) assay. The effect of DMY on migration was detected by wound healing(after 24 hours) and Transwell assay(after treated JEG-3 for 36 hours and JAR for 24 hours). The expression of matrix metalloproteinase 2(MMP-2) at mRNA and protein levels with different concentrations of DMY(0 mg/L, 40 mg/L, 60 mg/L, 80 mg/L) were detected by Real-time PCR(after 12 hours,24 hours,36 hours,48 hours)and Western blotting(after treated 36 hours) respectively. Results The proliferation of JEG-3 and JAR cells was inhibited significantly(P<0.05), after DMY treatment for 24 hours and 48 hours.DMY inhibited the migration of JEG-3 and JAR cells significantly(P<0.05) with a dose-dependent manner. After JEG-3 and JAR Cells treated with DMY for 36 hours and 48 hours, the expression of MMP-2 mRNA decreased significantly(P<0.05),there were no significantly changes in DMY treatment with 12 hours and 24 hours. The expression level of MMP-2 protein was inhibited significantly after treatment with DMY for 36 hours(P<0.05). Conclusion DMY might inhibit the proliferation in choriocarcinoma JEG-3 and JAR cells with a dose-dependent manner. The invasion and migration were inhibited by DMY through downregulation of MMP-2.
引文
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[2] Xia J,Guo S,Fang T,et al.Dihydromyricetin induces autophagy in HepG2 cells involved in inhibition of mTOR and regulating its upstream pathways[J].Food Chemi Toxicol,2014,66:7-13.
[3] Zhou DZ,Sun HY,Yue JQ,et al.Dihydromyricetin induces apoptosis and cytorotective autophagy through ROS-NF-κB signalling in human melanoma cells[J].Free Radi Res,2017,51(5):517-528.
[4] Zhou Y,Shu F,Liang X,et al.Ampelosin induces cell growth inhibition and apoptosis in breast cancer cells through ROS generation and endoplasmic reticulum stress pathway[J].PLoS One,2014,9(2):e89021.
[5] Jiang L,Zhang Q,Ren H,et al.Dihydromyricetin enhances the chemo-sensitivity of nedalatin via regulation of the p53/Bcl-2 pathway in heatocellular carcinoma cells[J].PLoS One,2015,10(4):e0124994.
[6] Liu B,Zhou W,Chen X,et al.Dihydromyricetin induces mouse hepatoma Hepal6 cell apoptosis via the transforming growth factor β pathway[J].Mol Med Rep,2015,11(3):1609-1614.
[7] Liu J,Shu Y,Zhang Q,et al.Dihydromyricetin induces apoptosis and inhibits proliferation in hepatocellular carcinoma cells [J].Oncol Lett,2014,8(4):1645-1651.
[8] Jacob A,rekeris R.The regulation of MMP targeting to invadopodia during cancer metastasis[J].Front Cell Dev Biol,2015,3(4):1-4.
[9] Kessenbrock K,Plaks Ⅴ,Werb Z.Matrix metalloproteinases:regulators of the tumor microenvironment [J].Cell,2010,141(1):52-67.
[10] Roach DM,Fitridge RA,Laws PE,et al.Up-regulation of MMP-2 and MMP-9 leads to degradation of type Ⅳ collagen during skeletal muscle reperfusion injury;protection by the MMP inhibitor,doxycycline[J].Eur J Vasc Endovasc Surg,2002,23(3):260-269.
[11] Zhang YSh,Yang WL,Cui Ch.Chemical constituents from ampelopsis grossedentata [J].Chinese Traditional and Herbal Drugs,2003,34(5):402-403.(in Chinese)张友胜,杨伟丽,崔春.显齿蛇葡萄化学成分的研究[J].中草药,2003,34(5):402-403.
[12] Qiu P,Dong Y,Li B,et al.Dihydromyricetin modulates p62 and autohagy crosstalk with the Kea-1/Nrf2 pathway to alleviate ethanol-induced hepatic injury[J].Toxicol Lett,2017,274:31-41.
[13] Qiu ZhD,Luo FL,Shu Y,et al.Inhibitory effect of dihydromyricetin on adhesion,invasion and migration in hepatocellular carcinoma cells and the mechanism[J].Chinese Journal of General Surgery,2015,24(9):1263-1268.(in Chinese)邱志东,罗芳兰,舒洋,等.二氢杨梅素对肝癌细胞黏附、侵袭及迁移的抑制作用及机制[J].中国普通外科杂志,2015,24(9):1263-1268.
[14] Zhang QY,Li R,Zeng GF,et al.Dihydromyricetin inhibits migration and invasion of hepatoma cells through regulation of MMP-9 expression[J].World J Gastroenterol,2014,20(29):10082-10093.
[15] Zhou FZh,Zhang XF.Suppression of distant pulmonary metastasis of 4T1 mice breast carcinoma by dihydromyricetin administration [J].Chinese Journal of Clinicians(Electronic Edition),2014,8(9):1674-1678.(in Chinese)周防震,张新芳.二氢杨梅素对4T1小鼠乳腺癌肺转移的抑制作用[J].中华临床医师杂志(电子版),2014,8(9):1674-1678.
[16] Zuo YZh,Sun DY,Bi HD,et al.Apoptosis-induced and antitumor effect of Dihydromyricetin[J].Chinese Traditional Patent Medicine,2015,37(8):1849-1852.(in Chinese)左彦珍,孙大永,毕红东,等.二氢杨梅素诱导凋亡抗肿瘤作用[J].中成药,2015,37(8):1849-1852.
[17] Sun DY,Zuo YZh,Liang HH,et al.Effects of dihydromyricetin on apoptosis of lung adenocarcinoma A549 cells[J].Acta Anatomica Sinica,2015,46(3):359-362.(in Chinese)孙大永,左彦珍,梁鸿鹄,等.二氢杨梅素对肺癌A549细胞凋亡的作用[J].解剖学报,2015,46(3):359-362.
[18] Liu L,Yin XL,Wang X,et al.Determination of dihydromyricetin in rat plasma by LC-MS/MS and its application to a pharmacokinetic study[J].Pharm Biol,2016,55(1):657-662.
[19] Ding YCh,Liu ZL,Xu XY,et al.Study on synthesis,characterization and anti-tumor activity of quercetin-3-sulphate sodium[J].Journal of Gannan Medical University,2012,32(1):16-18.(in Chinese)丁冶春,刘宗亮,徐仙赟,等.槲皮素-3-硫酸酯钠的合成、表征及抗肿瘤活性研究[J].赣南医学院学报,2012,32(1):16-18.