[1]周 巧,李佳会,庞金龙,等.吉非替尼抑制糖酵解诱导非小细胞肺癌的程序性死亡[J].南方医科大学学报,2020,(06):884-892.[doi:10.12122/j.issn.1673-4254.2020.06.17]
 Gefitinib inhibits glycolysis and induces programmed cell death in non-small cell lungcancer cells[J].Journal of Southern Medical University,2020,(06):884-892.[doi:10.12122/j.issn.1673-4254.2020.06.17]
点击复制

吉非替尼抑制糖酵解诱导非小细胞肺癌的程序性死亡()
分享到:

《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2020年06期
页码:
884-892
栏目:
出版日期:
2020-06-17

文章信息/Info

Title:
Gefitinib inhibits glycolysis and induces programmed cell death in non-small cell lung cancer cells
作者:
周 巧李佳会庞金龙范方田李姗姗刘 浩
关键词:
吉非替尼非小细胞肺癌糖酵解死亡形式
Keywords:
gefitinib non-small cell lung cancer glycolysis programmed cell death
DOI:
10.12122/j.issn.1673-4254.2020.06.17
文献标志码:
A
摘要:
目的 探究吉非替尼对非小细胞肺癌A549和H1975细胞死亡形式的影响,并从糖酵解方面探讨其可能机制。方法 A549细胞加入浓度分别为0、20、30、40 µmol/L的吉非替尼,H1975细胞加入浓度分别为0、20、40、80 µmol/L的吉非替尼。采用MTT法检测吉非替尼对非小细胞肺癌细胞的增殖抑制作用。用乳酸试剂盒检测细胞内乳酸的变化,Western blot法检测细胞内糖酵解相关蛋白(PKM2、HK2)和PI3K-Akt-mTOR信号通路中蛋白的表达水平;2-NBDG检测细胞葡萄糖摄取能力,ATP试剂盒检测胞内ATP水平;JC-1试剂盒检测细胞线粒体膜电位,Annexin V-FITC/PI双染法检测细胞凋亡,Western blot法检测凋亡蛋白(Bax、Bcl-2)以及自噬标志蛋白LC3B的相对表达水平。结果 MTT结果显示吉非替尼可呈时间剂量依赖性的抑制A549和H1975细胞增殖(P<0.05),A549细胞24、48、72 h的IC50值分别为48.6、28.6和19.7 µmol/L,H1975细胞24、48、72 h的IC50值分别为321.6、49.1和14.6 µmol/L。乳酸检测结果显示,吉非替尼抑制胞内乳酸水平(P<0.05)。Western blot结果显示,糖酵解相关蛋白PKM2、HK2表达下调(P<0.05),PI3K-Akt-mTOR信号通路中相关蛋白表达下调(P<0.05)。吉非替尼也可以抑制A549和H1975细胞葡萄糖摄取、ATP水平(P<0.05)。JC-1试剂盒和Annexin V-FITC/PI双染法检测出吉非替尼能够诱导A549和H1975细胞发生凋亡,A549细胞中0、20、30、40 µmol/L的凋亡率分别为(10.77±1.0)%、(14.5±0.4)%、(17.4±0.2)%、(32.1±0.6)%,差异有统计学意义(P<0.05);而H1975细胞0、20、40、80 µmol/L的凋亡率分别为(10.5±0.6)%、(13.2±0.92)%、(18.9±0.98)%、(35.1±1.4)%,差异有统计学意义(P<0.05)。促凋亡蛋白Bax表达增加和抑凋亡蛋白Bcl-2表达下调(P<0.05)。同时LC3B表达增加证明吉非替尼能够诱导A549和H1975细胞自噬增加(P<0.05)。结论 吉非替尼对A549和H1975细胞具有增殖抑制、诱导凋亡和增加自噬作用,凋亡机制可能为吉非替尼影响A549和H1975细胞糖酵解功能和PI3K-Akt-mTOR信号通路。
Abstract:
Objective To observe the cell death pattern induced by gefitinib in non-small cell lung cancer A549 and H1975 cells and explore the possible mechanism in light of glycolysis. Methods The inhibitory effects of gefitinib at 20, 30, or 40 µmol/L in A549 cells and at 20, 40, or 80 µmol/L in H1975 cells were examined using MTT assay. The changes of lactic acid level in the cells were determined with a lactic acid kit, and the expression levels of glycolysis-related proteins (PKM2 and HK2) and the proteins in PI3K-Akt-mTOR signaling pathway were detected using Western blotting. 2-NBDG was used for detecting glucose uptake capacity of the cells, and ATP kit was used to detect the intracellular ATP level. The mitochondrial membrane potential of the cells was examined with the JC-1 kit, and cell apoptosis was analyzed with Annexin V-FITC/PI double staining. The relative expression levels of the apoptotic proteins Bax and Bcl-2 and the autophagy marker protein LC3B were detected with Western blotting. Results MTT assay showed that gefitinib inhibited the proliferation of A549 and H1975 cells in a time- and dose-dependent manner (P<0.05). The IC50 of gefitinib at 24, 48 and 72 h was 48.6, 28.6 and 19.7 µmol/L in A549 cells and was 321.6, 49.1 and 14.6 µmol/L in H1975 cells, respectively. Gefitinib significantly lowered intracellular lactic acid level of the cells (P<0.05) and down-regulated the expressions of PKM2 and HK2 proteins (P<0.05) and PI3K-Akt-mTOR signaling pathway-associated proteins (P<0.05). Gefitinib obviously inhibited glucose uptake and ATP levels in both A549 and H1975 cells (P<0.05). Treatment with gefitinib induced obviously enhanced apoptosis in the cells, resulting in apoptosis rates of (10.77± 1.0)%, (14.5±0.4)%, (17.4±0.2)% and (32.1±0.6)% at 0, 20, 30 and 40 µmol/L in A549 cells (P<0.05) and of (10.5±0.6)%, (13.2± 0.92)%, (18.9±0.98)% and (35.1±1.4)% at 0, 20, 40 and 80 µmol/L in H1975 cells, respectively (P<0.05). The protein expression of Bax increased and that of Bcl-2 decreased following gefitinib treatment in the cells (P<0.05). Gefitinib significantly increased autophagy in A549 and H1975 cells as shown by increased LC3B expressions following the treatment (P<0.05). Conclusion Gefitinib can inhibit the proliferation, induce apoptosis and increase autophagy in A549 and H1975 cells. Gefitinib induces apoptosis of the cells possibly by affecting glycolysis and PI3K-Akt-mTOR signaling pathway.

相似文献/References:

[1]何本夫,孙爱民,黄碧燕,等.吉非替尼对鼻咽癌细胞株CNE2放疗增敏作用[J].南方医科大学学报,2011,(06):991.
[2]龙顺钦,廖桂雅,河文峰,等.生活质量量表评价参附注射液对肺癌化疗患者生活质量的影响[J].南方医科大学学报,2011,(12):2090.
[3]邱幸生,陈龙华,陈永清.非小细胞肺癌脑转移低分割放疗疗效分析[J].南方医科大学学报,2005,(11):1375.
 QIU Xing-sheng,CHEN Long-hua,CHEN Yong-qing.Three-dimensional conformal hypofractionated radiotherapy for brain metastases of non-small-cell lung carcinoma:implications for whole brain irradiation[J].Journal of Southern Medical University,2005,(06):1375.
[4]翁毅敏,谷力加,钟文昭,等.肺癌全肺切除术后严重心肺并发症相关因素分析[J].南方医科大学学报,2006,(11):1671.
[5]吴璇,李建雄.克唑替尼治疗EML4-ALK重排阳性患者的临床疗效[J].南方医科大学学报,2015,(05):753.
[6]谢亚琳,梁继珍,苏宁.吉非替尼与厄洛替尼在EGFR基因敏感突变晚期NSCLC患者一线治疗中的疗效比较[J].南方医科大学学报,2015,(03):446.
[7]胡婷华,姚煜,于硕,等.非小细胞肺癌组织中CXCR4和Nrf2的表达及其临床意义[J].南方医科大学学报,2014,(02):153.
[8]郭倩倩,刘志燕,姜丽丽,等.营养缺乏对人肺癌细胞自噬的诱导作用[J].南方医科大学学报,2014,(05):627.
[9]吴伟东,刘 丹,侯文进,等.RNA干扰抑制CD59对非小细胞肺癌GLC-P细胞株增殖的影响[J].南方医科大学学报,2015,(06):903.
[10]洪斌,沈学远,陈江勇.联合亚甲蓝和99mTc-硫胶体核素示踪法在早期非小细胞肺癌前哨淋巴结定位中的应用[J].南方医科大学学报,2014,(07):1053.

更新日期/Last Update: 2020-06-17