口腔医学研究

口腔医学研究 ›› 2019, Vol. 35 ›› Issue (6): 573-576.DOI: 10.13701/j.cnki.kqyxyj.2019.06.015

• 口腔肿瘤学研究 • 上一篇    下一篇

白藜芦醇通过调控雷帕霉素靶蛋白/M2型丙酮酸激酶轴抑制口腔鳞状细胞癌细胞的糖酵解作用

熊璟,何元春*,刘杰,张英,李永强,邱勇棋,庄瑞   

  1. 汉中市中心医院口腔科 陕西 汉中 723000
  • 收稿日期:2018-11-07 出版日期:2019-06-28 发布日期:2019-06-27
  • 通讯作者: 何元春,E-mail:heyc_g109@163.com
  • 作者简介:熊璟(1985~ ),女,陕西汉中人,硕士,主要从事口腔疾病防治工作。

Resveratrol Inhibits Glycolysis of Oral Squamous Cell Carcinoma by Regulating mTOR/ PKM2 Axis

XIONG Jing, HE Yuan-chun*, LIU Jie, ZHANG Ying, LI Yong-qiang, QIU Yong-qi, ZHUANG Rui   

  1. Department of Stomatology, Hanzhong Central Hospital, Hanzhong 723000, China
  • Received:2018-11-07 Online:2019-06-28 Published:2019-06-27

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摘要： 目的:通过雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)/M2型丙酮酸激酶(M2 isoform pyruvate kinase,PKM2)轴探讨白藜芦醇对口腔鳞状细胞癌(oral squamous cell carcinoma,OSCC)细胞的糖酵解的影响。方法:培养CAL-27和SCC-15细胞,采用不同浓度白藜芦醇(0、50、100、200、400、800 μmol/L)分别处理细胞24 h,噻唑蓝(methyl thiazolyl tetrazolium,MTT)法检测其对细胞增殖的影响并计算半数抑制浓度(half maximal inhibitory concentration,IC50)值。选用250 μmol/L白藜芦醇或10 nmol/L mTOR抑制剂Rapamycin干预细胞24 h,采用2-脱氧葡萄糖类似物[(2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]-D-glucose,2-NBDG]法检测细胞葡萄糖摄取率;比色法检测细胞上清中乳酸含量、细胞内糖酵解关键酶己糖激酶(hexokinase,HK)和丙酮酸激酶(pyruvate kinase,PK)活性;Western blot检测细胞中己糖激酶2(hexokinase2,HK2)、PKM2、mTOR和磷酸化mTOR(phosphorylation mTOR,p-mTOR)等蛋白表达情况。结果:白藜芦醇对两株OSCC细胞的增殖有明显的抑制作用(P＜0.05),并呈现浓度依赖性。白藜芦醇可显著抑制两株OSCC细胞的葡萄糖摄取能力以及降低乳酸的产生量(P＜0.05),并可抑制两株细胞PK活性(P＜0.05)、下调PKM2蛋白的表达(P＜0.05),但对HK活性及HK2蛋白的表达无显著影响(P＞0.05)。同时,白藜芦醇干预还可显著降低细胞内p-mTOR水平(P＜0.05)。另外,Rapamycin干预也可抑制两株细胞的葡萄糖摄取能力、降低细胞上清中乳酸的产生量和细胞内p-mTOR水平,同时也可下调PKM2蛋白的表达。结论:白藜芦醇可抑制OSCC细胞的糖酵解作用,其机制可能与抑制mTOR/PKM2轴通路相关。

关键词: 白藜芦醇, 口腔鳞状细胞癌, 糖酵解, 雷帕霉素靶蛋白/M2型丙酮酸激酶轴

Abstract: Objective: To investigate the effect resveratrol on glycolysis of oral squamous cell carcinoma (OSCC) by regulating mTOR/PKM2 axis. Methods: OSCC cell lines CAL-27 and SCC-15 were cultured and treated with different concentrations of resveratrol (0 μmol/L, 50 μmol/L, 100 μmol/L, 200 μmol/L, 400 μmol/L, and 800 μmol/L) for 24 h. Cell proliferation was evaluated by MTT assay, and the value of IC50 was calculated. OSCC cells were treated with 250 μmol/L resveratrol or 10 nmol/L rapamycin (mTOR inhibitor) for 24 h, and then the cellular glucose uptake was assessed by 2-NBDG staining. Lactate production in the cell supernatant and the activity of hexokinase and pyruvate kinase in the cells were detected using colorimetric method. The proteins expression of hexokinase-2 (HK2), M2 isoform pyruvate kinase (PKM2), mTOR, and p-mTOR in the cells were analyzed by Western blotting. Results: Resveratrol had an obvious inhibitory effect on the proliferation of CAL-27 and SCC-15 cells in a concentration dependent manner (P＜0.05). Resveratrol intervention significantly inhibited the glucose uptake and decreased the production of lactate in both OSCC cells (P<0.05). It also significantly inhibited the activity of pyruvate kinase and decreased the protein expression of PKM2 in CAL-27 and SCC-15 cells (P<0.05). However, it had no significant effect on the activity of hexokinase and the protein expression of HK-2 (P>0.05). At the same time, resveratrol intervention also significantly decreased the phosphorylation level of mTOR (P<0.05). In addition, rapamycin intervention could also inhibit glucose uptake, decrease the production of lac- tate and the phosphorylation level of mTOR, and down-regulate the protein expression of PKM2 in CAL-27 and SCC-15 cells. Conclusion: Resveratrol inhibits the glycolysis of OSCC cells, and its mechanism might relate to the inhibition of the mTOR/PKM2 axis pathway.

Key words: Resveratrol, Oral squamous cell carcinoma, Glycolysis mTOR/PKM2 axis