迟鹏 郝庆 姚艳春 孙玉兰 许德颖 王嘉怡

摘要 目的：研究胡黃连苷Ⅱ联合曲美替尼通过miR-4319对结直肠癌细胞HT-29凋亡和上皮细胞间质转化（EMT）的影响。方法：将结直肠癌细胞HT-29分成6组，分别为对照组、曲美替尼组、胡黄连苷Ⅱ组、联合组、miR-NC抑制组、miR-4319抑制组，n=9。采用CCK-8实验检测细胞增殖，流式细胞术检测细胞凋亡，Western Blotting法检测Vimentin、E-cadherin、Bax、C-Caspase-3蛋白表达，qRT-PCR检测miR-4319表达。比较6个组的检测结果。结果：与对照组比较，曲美替尼组、胡黄连苷Ⅱ组、联合组的细胞成活率和Vimentin的表达均减少（均P<0.05），而凋亡率和Bax、C-Caspase-3、E-cadherin的表达均增加（均P<0.05）;与miR-NC抑制组比较，miR-4319抑制组的细胞成活率、Vimentin的表达均显著增加（均P<0.05），细胞凋亡率和Bax、C-Caspase-3、E-cadherin蛋白表达却显著减少（均P<0.05）;qRT-PCR结果显示，与对照组比较，曲美替尼组、胡黄连苷Ⅱ组细胞中miR-4319的表达增多（均P<0.05）。结论：胡黄连苷Ⅱ联合曲美替尼通过上调miR-4319诱导结直肠癌细胞HT-29凋亡，并抑制细胞EMT。

关键词 胡黄连苷Ⅱ;曲美替尼;结直肠癌;miR-4319;增殖;凋亡;上皮间质转化;HT-29细胞

Effects of Coprininoside Ⅱ Combined with Trametinib in HT-29 Cells Apoptosis and EMT

CHI Peng1，HAO Qing2，YAO Yanchun1，SUN Yulan1，XU Deying3，WANG Jiayi4

（1 Department of Gastroenterology，Anshan Central Hospital Liaoning Province，Anshan 114001，China; 2 Department of Digestive Medicine，Shengjing Hospita Affiliated to China Medical University，Shenyang 110000，China; 3 Radiotherapy Department of Panjin Central Hospital of Liaoning Province，Panjin 124010，China; 4 Xiangnan University，Chenzhou 423000，China）

Abstract Objective：To study the effects of cucurbitacin Ⅱ combined with trametinib on the colorectal cancer cells HT-29 apoptosis and epithelial-mesenchymal transition（EMT） through miR-4319.Methods：Colorectal cancer cells HT-29 were divided into 6 groups，namely the a control group，a trametinib group，a coprininoside II group，a combination group，an miR-NC inhibition group，and an miR-4319 inhibition group（n=9）.In 6 groups，CCK-8 experiment was used to detect cell proliferation，flow cytometry was used to detect cell apoptosis，Western blot was used to detect the protein expression of Vimentin，E-cadherin，Bax，and C-Caspase-3，and the expression of miR-4319 was detected by qRT-PCR.Compare the experimental results of the 6 groups.Results：Compared with the control group，the cell survival rate，and the expression of Vimentin in the trametinib group，the berberine Ⅱ group and the combination group were reduced（P<0.05），while the apoptosis rate and the expression of Bax，C-Caspase-3，E-cadherin both increased（P<0.05）; compared with the miR-NC inhibition group，the cell survival rate and Vimentin expression in the miR-4319 inhibition group were significantly increased（P<0.05）.However，the apoptosis rate，the expression of Bax，C-Caspase-3 and E-cadherin were significantly reduced（P<0.05）; qRT-PCR results showed that compared with the control group，the expression of miR-4319 in the cells of the trametinib group and berberineⅡgroup increased（P<0.05）.Conclusion：The combination of cucurbitacin Ⅱ and trametinib induces the apoptosis and inhibits cell EMT of colorectal cancer cells HT-29 by up-regulating miR-4319.

Keywords Cuberitin II; Trametinib; Colorectal cancer; miR-4319; Proliferation; Apoptosis; EMT; HT-29 cells

中图分类号：R271.8文献标识码：Adoi：10.3969/j.issn.1673-7202.2021.17.012

结直肠癌发病率和致死率极高，是世界范围的公共卫生问题[1-2]。曲美替尼是口服MEK1/2抑制剂，具有抑制肿瘤细胞增殖的作用。曲美替尼可体外诱导结直肠癌细胞凋亡并抑制细胞增殖[3]。胡黄连苷Ⅱ提取自玄参植物胡黄连根茎，具有抗氧化、预防肝损伤、改善脑损伤等功效[4]。有研究发现，胡黄连苷Ⅱ可诱导肾癌细胞凋亡，抑制乳腺癌细胞转移，胡黄连苷Ⅱ可能是抗肿瘤药物[5]。目前胡黄连苷Ⅱ联合曲美替尼对结直肠癌细胞的作用鲜有研究。miRNA在人体内普遍表达，参与肿瘤等疾病进展[6]。抗肿瘤药物抑制肿瘤恶性生长的机制与miRNA表达改变有关。我们前期的预实验发现，胡黄连苷Ⅱ联合曲美替尼可提高结直肠癌细胞中miR-4319表达水平，胡黄连苷Ⅱ联合曲美替尼作用机制可能与miR-4319有关。既往文献[7]显示，miR-4319在结直肠癌中发挥类似抑癌基因的作用。本研究探讨胡黄连……