Rev-erb激动剂SR9009通过降低自噬抑制人结肠癌细胞系HCT116增殖

基础医学与临床 ›› 2020, Vol. 40 ›› Issue (11): 1494-1499.

Rev-erb激动剂SR9009通过降低自噬抑制人结肠癌细胞系HCT116增殖

石光¹, 宁娜¹, 侯晓鸿¹, 孙聪¹, 原媛², 王丽¹, 王晓晖^1*

1.山西医科大学基础医学院病理教研室;
2.山西医科大学基础医学院形态学实验室, 山西太原 030001

收稿日期:2019-11-07 修回日期:2020-01-20 出版日期:2020-11-05 发布日期:2020-10-30
通讯作者: ^* 163.wangxh@163.com

Rev-erb agonist SR9009 inhibits the proliferation of human colon cancer cell line HCT116 through reducing autophagy

SHI Guang¹, NING Na¹, HOU Xiao-hong¹, SUN Cong¹, YUAN Yuan², WANG Li¹, WANG Xiao-hui^1*

1. Department of Pathology;
2. Laboratory of Morphology, School of Basic Medicine, Shanxi Medical University, Taiyuan 030001, China

Received:2019-11-07 Revised:2020-01-20 Online:2020-11-05 Published:2020-10-30
Contact: ^* 163.wangxh@163.com

摘要/Abstract

摘要： 目的探讨血红素调节核受体(Rev-erb)激动剂SR9009对结肠癌细胞系HCT116增殖活性的作用及其机制。方法培养HCT116细胞,向培养液中加入Rev-erb激动剂SR9009,倒置相差显微镜观察细胞的增殖;选择SR9009的适宜浓度与作用时间,采用CCK-8法检测细胞活性;Real-time PCR检测Rev-erb α、Rev-erb β以及自噬基因Beclin1、LC3、p62的转录;Western blot检测Rev-erb α、Rev-erb β、Beclin1、LC3和p62的蛋白表达。结果 1)与对照组相比,SR9009显著抑制HCT116细胞的增殖(P＜0.05),SR9009的有效作用浓度为20 μmol/L,作用时间为24 h。 2)SR9009显著提高HCT116细胞Rev-erb α和Rev-erb β的转录(P＜0.01)以及蛋白表达水平(P＜0.05)。3)SR9009显著降低HCT116细胞Beclin1和LC3的表达水平(P＜0.05),引起自噬底物p62累积。4)在SR9009处理基础上,用3-甲基腺嘌呤(3-MA)抑制自噬,HCT116细胞活性降低(P＜0.05);用雷帕霉素提高自噬,HCT116细胞活性回升(P＜0.05)。结论 Rev-erb激动剂SR9009可以通过降低自噬来抑制结肠癌细胞系HCT116的增殖活性。

关键词: Rev-erb, SR9009, 结肠癌, 自噬

Abstract: Objective To investigate the effect of hemeregulated nuclear receptor(Rev-erb) agonist SR9009 on the proliferation and potential mechanism of human colon cancer cell line HCT116. Methods HCT116 cells were cultured and treated with Rev-erb agonist SR9009 with proper dosagee and treatment time. The cell proliferation was observed with a phase contrast microscopve. The cell viability was tested using a CCK-8 kit. The mRNA transcription of Rev-erb α, Rev-erb β and autophagy genes Beclin1, LC3 and p62 were detected by real-time PCR. The protein expression of Rev-erb α, Rev-erb β, Beclin1, LC3 and p62 were detected by Western blot. Results 1)Rev-erb agonist SR9009 inhibited the HCT116 cell proliferation as compared with control group (P＜0.05), and the right dose of SR9009 was 20 μmol/L and the treatment time was 24 hours. 2)SR9009 significantly increased Rev-erb α and Rev-erb β mRNA transcription (P＜0.01) and protein expression (P＜0.05) in HCT116 cells. 3)R9009 significantly reduced the expression of autophagy genes Beclin1 and LC3 in HCT116 cells (P＜0.05), causing autophagy protein p62 accumulation. 4)HCT116 cell viability was further decreased with an autophagy inhibitor 3-methyladenine (3-MA) after SR9009 treatment (P＜0.05), the cell viability was restored by an autophagy activator rapamycin (P＜0.05). Conclusions Rev-erb agonist SR9009 inhibits proliferation and cell viability of colon cancer cell line HCT116 by reducing autophagy.

Key words: Rev-erb, SR9009, colon cancer, autophagy

中图分类号:

石光, 宁娜, 侯晓鸿, 孙聪, 原媛, 王丽, 王晓晖. Rev-erb激动剂SR9009通过降低自噬抑制人结肠癌细胞系HCT116增殖[J]. 基础医学与临床, 2020, 40(11): 1494-1499.

SHI Guang, NING Na, HOU Xiao-hong, SUN Cong, YUAN Yuan, WANG Li, WANG Xiao-hui. Rev-erb agonist SR9009 inhibits the proliferation of human colon cancer cell line HCT116 through reducing autophagy[J]. Basic & Clinical Medicine, 2020, 40(11): 1494-1499.

参考文献

[1] Bray F, Ferlay J, Soerjomataram I, et al. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2018, 68:394-424.
[2] Wu Y, Yao J, Xie J, et al. The role of autophagy in colitis-associated colorectal cancer[J]. Signal Transduct Target Ther, 2018, 3:1-11.
[3] Ma D, Li S, Molusky MM, et al. Circadian autophagy rhythm: a link between clock and metabolism?[J]. Trends Endocrinol Metab, 2012, 23:319-325.
[4] Sulli G, Rommel A, Wang X, et al. Pharmacological activation of REV-ERBs is lethal in cancer and oncogene-induced senescence[J]. Nature, 2018, 553:351-355.
[5] Yu T, Guo F, Yu Y, et al. Fusobacterium nucleatum promotes chemoresistance to colorectal cancer by modulating autophagy[J]. Cell, 2017, 170:548-563.
[6] Sachdeva UM, Thompson CB. Diurnal rhythms of autophagy: implications for cell biology and human disease[J]. Autophagy, 2008, 4:581-589.
[7] Huang RC. The discoveries of molecular mechanisms for the circadian rhythm: the 2017 Nobel prize in physiology or medicine[J]. Biomed J, 2018, 41:5-8.
[8] Ikeda R, Tsuchiya Y, Koike N, et al. REV-ERBα and REV-ERBβ function as key factors regulating mammalian circadian output[J]. Sci Rep, 2019, 9:1-9.
[9] Solt LA, Wang Y, Banerjee S. Regulation of circadian behaviour and metabolism by synthetic REV-ERB agonists[J]. Nature, 2012, 485:62-68.
[10] Ercolani L, Ferrari A, De Mei C,et al. Circadian clock: time for novel anticancer strategies?[J]. Pharmacol Res, 2015, 100:288-295.
[11] Wang Y, Kojetin D, Burris TP. Anti-proliferative actions of a synthetic REV-ERBα/β agonist in breast cancer cells[J]. Biochem Pharmacol, 2015, 96:315-322.
[12] Dong Z, Zhang G, Qu M, et al. Targeting glioblastoma stem cells through disruption of the circadian clock[J]. Cancer Discov, 2019, 9:1325-1343.
[13] Huang G, Zhang F, Ye Q, et al. The circadian clock regulates autophagy directly through the nuclear hormone receptor Nr1d1/Rev-erbα and indirectly via Cebpb/(C/ebpβ) in zebrafish[J]. Autophagy, 2016, 12:1292-1309.
[14] Grimaldi B. Lysosomotropic REV-ERB antagonism: A metabolic connection between circadian rhythm and autophagy may tell cancer cells “it's time to die”[J]. Mol Cell Oncol, 2014, 2:e965626.doi: 10.4161/23723548.2014.965626.
[15] Bauvy C, Gane P, Arico S, et al. Autophagy delays sulindac sulfide-induced apoptosis in the human intestinal colon cancer cell line HT-29[J]. Exp Cell Res, 2001, 268:139-149.