敲除TBRG4对肺癌H1299细胞增殖的影响及其机制研究

doi:10.3760/cma.j.cn101202-20210927-00273

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摘要目的探讨敲除转化生长因子β调节因子4（TBRG4）对肺癌H1299细胞增殖的影响及其机制。方法培养肺癌H1299细胞、TBRG4敲除阴性对照及TBRG4敲除H1299细胞,分别命名为对照组、阴性对照组、TBRG4敲除组。采用细胞计数（CCK）-8试剂盒检测各组细胞的增殖情况。采用还原型谷胱甘肽（GSH）检测试剂盒检测各组细胞GSH的含量。采用实时荧光定量聚合酶链反应（qRT-PCR）检测各组细胞TBRG4、沉默信息调节因子2相关酶4（SIRT4）、c-Myc和谷氨酰胺酶1（GLS1）mRNA的表达。采用Western blot检测各组细胞TBRG4、SIRT4、c-Myc和GLS1蛋白的表达水平。结果 CCK-8和GSH含量检测结果显示,对照组、阴性对照组、TBRG4敲除组H1299细胞的吸光度值分别为1.025±0.021、1.032±0.019、0.726±0.018,GSH的含量分别为（20.836±0.367）、（21.167±0.460）、（15.091±0.241） μmol/L,与对照组和阴性对照组相比,TBRG4敲除组的吸光度值、GSH含量均降低,差异均有统计学意义（P值均<0.001）。与对照组和阴性对照组相比,TBRG4敲除组TBRG4、c-Myc、GLS1基因mRNA和蛋白的相对表达水平均减少,SIRT4基因mRNA和蛋白的相对表达水平均增加,差异均有统计学意义（P值均<0.001）。结论敲除TBRG4可抑制肺癌H1299细胞增殖,其作用机制可能与促进SIRT4的表达、抑制c-Myc和GLS1的表达、阻断谷氨酰胺代谢有关。

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	王安生
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	桑海威
	李超

关键词 ：肺肿瘤, H1299细胞, 转化生长因子&#x003b2, 调节因子4, 细胞增殖, 谷氨酰胺代谢

Abstract：Objective To investigate the effect of transforming growth factor β regulator 4 (TBRG4) knockdown on glutamine metabolism and lung cancer H1299 cell proliferation and to elucidate the underlying mechanism. Methods H1299 cells, TBRG4 knockout negative control, and TBRG4 knockout H1299 cells were cultured and set as the control group, the negative control group, and the TBRG4 knockout group, respectively. The proliferation of cells was measured using cell counting kit (CCK)-8 assay, and the content of glutathione (GSH) was evaluated using GSH assay. The mRNA expression levels of TBRG4, silencing information regulator 2-related enzyme 4 (SIRT4), c-Myc, and glutaminase 1 (GLS1) genes were detected using quantitative real-time polymerase chain reaction. The protein expression levels of TBRG4, SIRT4, c-Myc, and GLS1 were measured using western blot. Results CCK-8 and GSH assays showed that the optical density values of H1299 cells in the control group, negative control group, and TBRG4 knockout group were 1.025 ± 0.021, 1.032 ± 0.019, and 0.726±0.018, respectively, and the GSH contents were (20.836±0.367), (21.167±0.46), and (15.091±0.241) μmol/L, respectively. Compared with the those of the control group and the negative control group, the OD value and GSH content of the TBRG4 knockout group significantly decreased (all P values < 0.001). Compared with the control group and the negative control group, the TBRG4 knockout group showed significantly lower mRNA and protein expression levels of TBRG4, c-myc, and GLS1 and significantly higher mRNA and protein expression levels of SIRT4 (all P values < 0.001). Conclusion TBRG4 knockout inhibits the proliferation of lung cancer H1299 cells possibly by promoting the expression of SIRT4 and inhibiting the expression of c-Myc and GLS1 to block glutamine metabolism.

Key words： Lung neoplasms H1299 cell Transforming growth factor &#x003b2 regulator 4 Cell proliferation Glutamine metabolism

收稿日期: 2021-09-27

基金资助:安徽高校自然科学研究重点项目（KJ2019A0311）

通讯作者: 王安生,Email：807435169@qq.com

引用本文:

王安生, 朱浩楠, 洪海宁, 李建, 陈娜娜, 桑海威, 李超. 敲除TBRG4对肺癌H1299细胞增殖的影响及其机制研究[J]. 中华解剖与临床杂志, 2022, 27(7): 516-520.
Wang Ansheng, Zhu Haonan, Hong Haining, Li Jian, Chen Nana, Sang Haiwei, Li Chao. Inhibitory effect of transfected transforming growth factor β regulator 4 knockdown on the proliferation of lung cancer H1299 cells. Chinese Journal of Anatomy and Clinics, 2022, 27(7): 516-520.

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http://www.cjac.com.cn/CN/10.3760/cma.j.cn101202-20210927-00273 或 http://www.cjac.com.cn/CN/Y2022/V27/I7/516

[1]	Sung H, Ferlay J, Siegel RL, et al.Global Cancer Statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2021,71(3):209-249. DOI: 10.3322/caac.21660.
[2]	Schirrmacher V.Mitochondria at work: new insights into regulation and dysregulation of cellular energy supply and metabolism[J]. Biomedicines, 2020,8(11):526. DOI: 10.3390/biomedicines8110526.
[3]	霍宏宇, 胡苏琼, 孙洋,等. 肿瘤微环境及其能量代谢的重编程[J]. 生命科学, 2017, 29(1):70-76.DOI:10.13376/j.cbls/2017010.Huo HY, Hu SQ, Sun Y, et al.Tumor microenvironment and energy metabolic reprogramming[J]. Chinese Bulletin of Life Sciences, 2017, 29(1):70-76. DOI:10.13376/j.cbls/2017010.
[4]	Aminzadeh S, Vidali S, Sperl W, et al.Energy metabolism in neuroblastoma and Wilms tumor[J]. Transl Pediatr, 2015,4(1):20-32. DOI: 10.3978/j.issn.2224-4336.2015.01.04.
[5]	Netea-Maier RT, Smit J, Netea MG.Metabolic changes in tumor cells and tumor-associated macrophages: a mutual relationship[J]. Cancer Lett, 2018,413:102-109. DOI: 10.1016/j.canlet.2017.10.037.
[6]	Sun L, Suo C, Li ST, et al.Metabolic reprogramming for cancer cells and their microenvironment: beyond the Warburg effect[J]. Biochim Biophys Acta Rev Cancer, 2018,1870(1):51-66. DOI: 10.1016/j.bbcan.2018.06.005.
[7]	DeBerardinis RJ, Mancuso A, Daikhin E, et al. Beyond aerobic glycolysis: transformed cells can engage in glutamine metabolism that exceeds the requirement for protein and nucleotide synthesis[J]. Proc Natl Acad Sci U S A, 2007,104(49):19345-19350. DOI: 10.1073/pnas.0709747104.
[8]	van Geldermalsen M, Wang Q, Nagarajah R, et al. ASCT2/SLC1A5 controls glutamine uptake and tumour growth in triple-negative basal-like breast cancer[J]. Oncogene, 2016,35(24):3201-3208. DOI: 10.1038/onc.2015.381.
[9]	Craze ML, Cheung H, Jewa N, et al.MYC regulation of glutamine-proline regulatory axis is key in luminal B breast cancer[J]. Br J Cancer, 2018,118(2):258-265. DOI: 10.1038/bjc.2017.387.
[10]	Wang Y, He J, Liao M, et al.An overview of Sirtuins as potential therapeutic target: Structure, function and modulators[J]. Eur J Med Chem, 2019,161:48-77. DOI: 10.1016/j.ejmech.2018.10.028.
[11]	Huang F, Zhou P, Wang Z, et al.Knockdown of TBRG4 suppresses proliferation, invasion and promotes apoptosis of osteosarcoma cells by downregulating TGF-‍β1 expression and PI3K/AKT signaling pathway[J]. Arch Biochem Biophys, 2020,686:108351. DOI: 10.1016/j.abb.2020.108351.
[12]	Wolf AR, Mootha VK.Functional genomic analysis of human mitochondrial RNA processing[J]. Cell Rep, 2014,7(3):918-931. DOI: 10.1016/j.celrep.2014.03.035.
[13]	Cruz-Bermúdez A, Laza-Briviesca R, Vicente-Blanco RJ, et al.Cancer-associated fibroblasts modify lung cancer metabolism involving ROS and TGF-‍β signaling[J]. Free Radic Biol Med, 2019,130:163-173. DOI: 10.1016/j.freeradbiomed.2018.10.450.
[14]	Wang A, Zhao C, Liu X, et al.Knockdown of TBRG4 affects tumorigenesis in human H1299 lung cancer cells by regulating DDIT3, CAV1 and RRM2[J]. Oncol Lett, 2018,15(1):121-128. DOI: 10.3892/ol.2017.7328.
[15]	Miess H, Dankworth B, Gouw AM, et al.The glutathione redox system is essential to prevent ferroptosis caused by impaired lipid metabolism in clear cell renal cell carcinoma[J]. Oncogene, 2018,37(40):5435-5450. DOI: 10.1038/s41388-018-0315-z.
[16]	Traverso N, Ricciarelli R, Nitti M, et al.Role of glutathione in cancer progression and chemoresistance[J]. Oxid Med Cell Longev, 2013,2013:972913. DOI: 10.1155/2013/972913.
[17]	Jeong SM, Xiao C, Finley LW, et al.SIRT4 has tumor-suppressiveactivity and regulates the cellular metabolic response to DNA damage by inhibiting mitochondrial glutamine metabolism[J]. Cancer Cell, 2013, 23(4):450-463. DOI: 10.1016/j.ccr.2013.02.024.
[18]	Du L, Liu X, Ren Y, et al.Loss of SIRT4 promotes the self-renewal of breast cancer stem cells[J]. Theranostics, 2020,10(21):9458-9476. DOI: 10.7150/thno.44688.
[19]	Kodama M, Oshikawa K, Shimizu H, et al.A shift in glutamine nitrogen metabolism contributes to the malignant progression of cancer[J]. Nat Commun, 2020,11(1):1320. DOI: 10.1038/s41467-020-15136-9.
[20]	Ala M.Target c-Myc to treat pancreatic cancer[J]. Cancer Biol Ther, 2022,23(1):34-50. DOI: 10.1080/15384047.2021.2017223.
[21]	Vanhove K, Derveaux E, Graulus GJ, et al.Glutamine addiction and therapeutic strategies in lung cancer[J]. Int J Mol Sci, 2019,20(2):252. DOI: 10.3390/ijms20020252.
[22]	Shaw E, Talwadekar M, Rashida Z, et al.Anabolic SIRT4 exerts retrograde control over TORC1 signaling by glutamine sparing in the mitochondria[J].Mol Cell Biol,2020,40(2):e00212-19. DOI: 10.1128/MCB.00212-19.