人参皂苷Rh_2对结肠癌耐药细胞HCT116/L-OHP侵袭迁移能力的影响
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摘要
目的:观察人参皂苷Rh_2(ginsenoside Rh_2,GRh_2)对结肠癌耐药细胞HCT116/L-OHP迁移、侵袭的影响,并探讨其作用机制。方法:细胞计数试剂盒(cell counting kit-8,CCK-8)法检测不同质量浓度GRh_2(0,2. 5,5,10,20,40 mg·L~(-1))对HCT116/L-OHP细胞增殖抑制作用;划痕实验,Transwell实验及黏附实验分别检测GRh_2(0,2. 5,5,10 mg·L~(-1))对细胞迁移、侵袭及黏附能力的影响;蛋白免疫印迹法(Western blot)检测GRh_2(0,5,10,20,30 mg·L~(-1))对HCT116/L-OHP细胞上皮型钙黏蛋白(E-cadherin)和基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)蛋白表达水平的影响。结果:与空白组比较,GRh_2(5,10,20,40 mg·L~(-1))可显著抑制HCT116/L-OHP耐药细胞的增殖能力(P <0. 05,P <0. 01),并且呈浓度依赖性;与空白组比较,GRh_2组(5,10 mg·L~(-1))划痕愈合率明显减小(P <0. 05,P <0. 01),GRh_2组穿过小室的细胞数量明显减少(P <0. 05,P <0. 01),细胞的迁移和侵袭能力受到显著抑制,并且呈浓度依赖性; GRh_2组黏附细胞数量显著减少(P <0. 05,P <0. 01),细胞的黏附能力受到显著抑制,并且呈浓度依赖性;与空白组比较,GRh_2(10,20,30 mg·L~(-1))促进了E-cadherin的蛋白表达(P <0. 05,P <0. 01),同时抑制了MMP-9蛋白表达(P <0. 01),呈一定的浓度依赖性。结论:GRh_2可显著抑制结肠癌耐药细胞HCT116/L-OHP的侵袭和迁移能力,其潜在机制可能与促进E-cadherin并抑制MMP-9的表达有关。
Objective: To observe effect of ginsenoside Rh_2( GRh_2) on the invasion and migration of colon cancer resistant cells HCT116/L-OHP and its specific mechanism. Method: Cell counting kit-8( CCK-8)assay was used to detect the inhibitory effect of different concentrations of GRh_2( 0,2. 5,5,10,20,40 mg·L~(-1)) on HCT116/L-OHP cell proliferation,scratch assay,Transwell assay and adhesion assay were used to detect the effects of GRh_2( 0,2. 5,5,10 mg·L~(-1)) on cell migration,invasion and adhesion. The protein expression levels of E-cadherin and matrix metalloproteinase-9( MMP-9) were examined by Western blot. Result:Compared with control group,GRh_2( 5,10,20,40 mg·L~(-1)) significantly inhibited the proliferation of HCT116/L-OHP cells in a dose-dependent manner( P < 0. 05,P < 0. 01); Compared with the control group,the scratch healing rate of GRh_2 group( 5,10 mg·L~(-1)) was significantly decreased( P < 0. 05,P < 0. 01),the number of cells passing through the chamber of GRh_2 group was significantly decreased( P < 0. 05,P < 0. 01),cell migration and invasive ability were significantly inhibited in a dose-dependent manner. The number of adherent cells in GRh_2 group was significantly reduced( P < 0. 05,P < 0. 01),and the cell adhesion ability was significantly inhibited.Compared with the control group,GRh_2( 10,20,30 mg·L~(-1)) promoted E-cadherin protein expression( P <0. 05,P < 0. 01),while protein expression of MMP-9 was inhibited( P < 0. 01). Conclusion: GRh_2 can significantly inhibit the invasion and migration of HCT116/L-OHP in colon cancer cells, and its potential mechanism may be related to the promotion of E-cadherin and the inhibition of MMP-9 expression in a dosedependent manner.
Objective: To observe effect of ginsenoside Rh_2( GRh_2) on the invasion and migration of colon cancer resistant cells HCT116/L-OHP and its specific mechanism. Method: Cell counting kit-8( CCK-8)assay was used to detect the inhibitory effect of different concentrations of GRh_2( 0,2. 5,5,10,20,40 mg·L~(-1)) on HCT116/L-OHP cell proliferation,scratch assay,Transwell assay and adhesion assay were used to detect the effects of GRh_2( 0,2. 5,5,10 mg·L~(-1)) on cell migration,invasion and adhesion. The protein expression levels of E-cadherin and matrix metalloproteinase-9( MMP-9) were examined by Western blot. Result:Compared with control group,GRh_2( 5,10,20,40 mg·L~(-1)) significantly inhibited the proliferation of HCT116/L-OHP cells in a dose-dependent manner( P < 0. 05,P < 0. 01); Compared with the control group,the scratch healing rate of GRh_2 group( 5,10 mg·L~(-1)) was significantly decreased( P < 0. 05,P < 0. 01),the number of cells passing through the chamber of GRh_2 group was significantly decreased( P < 0. 05,P < 0. 01),cell migration and invasive ability were significantly inhibited in a dose-dependent manner. The number of adherent cells in GRh_2 group was significantly reduced( P < 0. 05,P < 0. 01),and the cell adhesion ability was significantly inhibited.Compared with the control group,GRh_2( 10,20,30 mg·L~(-1)) promoted E-cadherin protein expression( P <0. 05,P < 0. 01),while protein expression of MMP-9 was inhibited( P < 0. 01). Conclusion: GRh_2 can significantly inhibit the invasion and migration of HCT116/L-OHP in colon cancer cells, and its potential mechanism may be related to the promotion of E-cadherin and the inhibition of MMP-9 expression in a dosedependent manner.
引文
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[14]李丹,金凤,陶丽,等.金龙胶囊对胃癌细胞MGC-803和BGC-823侵袭转移能力的影响[J].中国实验方剂学杂志,2018,24(19):117-123.
[15]孔德帅,杨哲. E-cadherin和Ki-67在结直肠癌发展中的研究进展[J].临床与病理杂志,2015,35(12):2195-2199.
[16]常琳琳,朱虹,郑琳,等. E-cadherin在肿瘤治疗中的研究进展[J].药学进展,2015,39(10):754-760.
[17]王珊珊,赵新鲜,郭立中.益肺解毒汤对人肺腺癌A549细胞侵袭迁移的影响及相关机制[J].中国实验方剂学杂志,2017,23(22):108-113.
[18] Shirahata A,Sakata M,Sakuraba K,et al. Vimentin methylation as a marker for advanced colorectal carcinoma[J]. Anticancer Res,2009,29(1):279-281.
[19] Salem N,Kama I,Al-Maghrabi J,et al. High expression of matrix metalloproteinases:MMP-2 and MMP-9predicts poor survival outcome in colorectal carcinoma[J]. Future Oncol,2016,12(3):323-331.
[2] CHEN W Q,ZHENG R S,Baade P D,et al. Cancer Statistics in China,2015[J]. CA Cancer J Clin,2016,66(2):115-132.
[3] Rokavec M,Oner M G,LI H,et al. IL-6R/STAT3/miR-34a feedback loop promotes EMT-mediated colorectal cancer invasion and metastasis[J]. J Clin Invest,2014,124(4):1853-1867.
[4] De Krijger I,Mekenkamp L J, Punt C J, et al.MicroRNAs in colorectal cancer metastasis[J]. J Pathol,2011,224(4):438-447.
[5]石晓丽. SIRT1的抑制剂EX527对耐药细胞侵袭转移的影响及机制研究[D].郑州:郑州大学,2018.
[6] Kim J H,Kim M,Yun S M,et al. Ginsenoside Rh2induces apoptosis and inhibits epithelial-mesenchymal transition in HEC1A and Ishikawa endometrial cancer cells[J]. Biome Pharmacother,2017,doi:10. 1016/j.biopha. 2017. 09. 033.
[7] YANG J J,YUAN D H,XING T C,et al. Ginsenoside Rh2inhibiting HCT116 colon cancer cell proliferation through blocking PDZ-binding kinase/T-LAK celloriginated protein kinase[J]. J Ginseng Res,2016,40(4):400-408.
[8]胡双双,闫克敏,王娇娇,等.人参皂苷Rh2对胃癌细胞SGC7901/ADR耐药敏感性的影响[J].中草药,2018,49(17):4113-4117.
[9] SHI Q,LI J,FENG Z,et al. Effect of ginsenoside Rh2on the migratory ability of Hep G2 liver carcinoma cells:Recruiting histone deacetylase and inhibiting activator protein 1 transcription factors[J]. Mol Med Rep,2014,10(4):1779-1785.
[10]冯子强,左国伟,石庆强,等.人参皂苷Rh2抑制肝癌Hep G2细胞迁移的实验研究[J].中国免疫学杂志,2015,31(1):61-65.
[11] TANG X P,TANG G D,FANG C Y,et al. Effects of ginsenoside Rh2on growth and migration of pancreatic cancer cells[J]. World J Gastroenterol,2013,19(10):1582-1592.
[12] Kim J H,Choi J S. Effect of ginsenoside Rh2via activation of Caspase-3 and Bcl-2-insensitive pathway in ovarian cancer cells[J]. Physiol Res,2016,65(6):1031-1037.
[13]苑家鑫,邹澄,田华,等.人参皂苷Rh2对B16黑色素瘤细胞侵袭、转移能力的影响[J].中国医药导报,2016,13(12):135-138.
[14]李丹,金凤,陶丽,等.金龙胶囊对胃癌细胞MGC-803和BGC-823侵袭转移能力的影响[J].中国实验方剂学杂志,2018,24(19):117-123.
[15]孔德帅,杨哲. E-cadherin和Ki-67在结直肠癌发展中的研究进展[J].临床与病理杂志,2015,35(12):2195-2199.
[16]常琳琳,朱虹,郑琳,等. E-cadherin在肿瘤治疗中的研究进展[J].药学进展,2015,39(10):754-760.
[17]王珊珊,赵新鲜,郭立中.益肺解毒汤对人肺腺癌A549细胞侵袭迁移的影响及相关机制[J].中国实验方剂学杂志,2017,23(22):108-113.
[18] Shirahata A,Sakata M,Sakuraba K,et al. Vimentin methylation as a marker for advanced colorectal carcinoma[J]. Anticancer Res,2009,29(1):279-281.
[19] Salem N,Kama I,Al-Maghrabi J,et al. High expression of matrix metalloproteinases:MMP-2 and MMP-9predicts poor survival outcome in colorectal carcinoma[J]. Future Oncol,2016,12(3):323-331.