自噬介导的雷公藤甲素提高西妥昔单抗对SW480细胞治疗效果的实验研究

目的观察雷公藤甲素联合表皮生长因子受体单克隆抗体西妥昔单抗对人结直肠癌细胞SW480细胞株生物学行为的影响。方法雷公藤甲素、西妥昔单抗单独或联合作用于SW480细胞株, MTT法、细胞克隆实验检测细胞增殖抑制率;细胞划痕实验检测细胞的迁移能力;Western blot检测雷公藤甲素诱导细胞自噬性凋亡标记物p62、LC3-Ⅱ;雷公藤甲素及西妥昔单抗单独和联合作用细胞后,Western blot分析上皮间质转化(EMT)标记物E-cadherin、Vimentin以及mTOR、Snail、Twist蛋白水平。结果雷公藤甲素及西妥昔单抗对SW480细胞的增殖抑制作用均呈剂量依赖性,两药联用可明显抑制SW480细胞增殖;雷公藤甲素诱导SW480细胞p62蛋白下调,LC3-Ⅱ上调,发生自噬性凋亡;雷公藤甲素单药组、联合用药组均能明显抑制SW480细胞发生EMT,其标记分子E-cadherin蛋白上调,Vimentin蛋白下调,关键分子Snail、Twist表达下调。结论雷公藤甲素联合西妥昔单抗抑制SW480细胞增殖和转移具有明显协同作用,雷公藤甲素通过抑制mTOR通路,可以诱导细胞自噬性凋亡,自噬介导的雷公藤甲素抑制肿瘤细胞EMT可能是逆转西妥昔单抗耐药的机制。

Autophagy induced synergistic inhibitory effect of cetuximab in combination with triptolide on proliferation and metastasis of colorectal SW480 cells

Aim To investigate the effect of triptolide combined with epidermal growth factor receptor monoclonal antibody cetuximab on the biological behavior of human colorectal cancer cell line SW480.Methods MTT assay was used for estimating the survival rates of SW480 cells exposed to different concentrations and different durations of triptolid and cetuximab.Colony formation assay was used for showing the proliferation and wound healing assay was performed to assess the effects of drugs on cell migration,Western blot was used for testing the expression of LC3-Ⅱ,p62,E-cadherin,Vimentin,mTOR,Snail,and Twist.Results The SW480 growth of cells was inhibited by triptolide in a dose-and time-dependent manner and cetuximab was only in a dose-dependent manner.The combination regimen of cetuximab and triptolide exerted a synergistic effect.Triptolide could decrease expression of p62 increase expression of LC3-Ⅱand induce autophagic apoptosis.Triptolide monotherapy group and combination group could significantly inhibit EMT in SW480 cells,and the E-cadherin protein was up-regulated,Vimentin protein was down-regulated,and key molecules Snail and Twist were down-regulated.Conclusion Triptolide combined with cetuximab has a synergistic effect on the inhibition of proliferation and metastasis of SW480 cells.Triptolide induces autophagic apoptosis by inhibiting the mTOR pathway.Autophagy mediated by triptolide can inhibit EMT of SW480 cells and may be a mechanism to reverse the resistance of cetuximab.