国际口腔医学杂志

国际口腔医学杂志 ›› 2018, Vol. 45 ›› Issue (6): 628-634.doi: 10.7518/gjkq.2018.06.002

• RNA专栏 • 上一篇    下一篇

长链非编码RNA lnc-p26090对口腔鳞状细胞癌细胞糖酵解及增殖的影响

李媛媛,程斌,王韵()   

  1. 中山大学光华口腔医学院·附属口腔医院·广东省口腔医学重点实验室 广州 510055
  • 收稿日期:2018-04-02 修回日期:2018-08-10 出版日期:2018-11-01 发布日期:2018-11-15
  • 通讯作者: 王韵
  • 作者简介:李媛媛,学士,Email： liyuany6@mail2.sysu.edu.cn
  • 基金资助:
    国家自然科学基金(81502356);广州市科技计划项目(201804010040)

Effects of long non-coding RNA lnc-p26090 on the glycolysis and proliferation in oral squamous cell carcinoma

Yuanyuan Li,Bin Cheng,Yun Wang()   

  1. Guanghua School of Stomatology, Hospital of Stomatology, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China
  • Received:2018-04-02 Revised:2018-08-10 Online:2018-11-01 Published:2018-11-15
  • Contact: Yun Wang
  • Supported by:
    This study was supported by National Natural Science Foundation of China(81502356);Science and Technology Program of Guangzhou(201804010040)

RichHTML

22

PDF (PC)

280

摘要：

目的 检测长链非编码RNA lnc-p26090在口腔鳞状细胞癌（OSCC）组织及细胞系中的表达情况,探讨lnc-p26090对OSCC细胞糖酵解和生长增殖的影响。方法 利用荧光实时定量聚合酶链式反应（RT-qPCR）技术检测OSCC组织及细胞系中lnc-p26090的表达水平;利用UCSC和LNCipedia数据库网站获取lnc-p26090的染色体定位信息及蛋白质编码潜能,并在HEK 293T细胞中转染重组质粒验证其蛋白质编码能力。在OSCC细胞系HSC-3和SCC-25中转染lnc-p26090后,采用葡萄糖/乳酸测定试剂盒及RT-qPCR技术检测细胞糖酵解能力改变,同时采用细胞计数试剂盒、流式细胞术及蛋白质印迹法技术检测细胞生长增殖能力改变。结果 与正常口腔角化上皮细胞及配对的癌旁组织相比,OSCC细胞系及癌组织中lnc-p26090的表达水平显著上调（P<0.05）。通过数据库查询及重组质粒转染验证,lnc-p26090是位于人类染色体2p11.2上的长链非编码RNA。在HSC-3及SCC-25细胞中转染lnc-p26090,可显著抑制细胞葡萄糖消耗量和乳酸产生量,同时细胞中糖酵解相关基因M2型丙酮酸激酶、人磷酸果糖激酶、己糖激酶2、人葡萄糖转运蛋白1和乳酸脱氢酶A基因的表达显著降低。另外,转染lnc-p26090后细胞增殖水平也受到显著抑制,同时细胞被阻滞在G1期,细胞中周期相关蛋白G1/S-特异性周期蛋白D1和人周期素依赖性激酶4表达显著降低,p21表达显著升高。结论 lnc-p26090在OSCC组织及细胞系中表达上调,是位于人类染色体2p11.2上的长链非编码RNA,并参与调控OSCC细胞糖酵解和生长增殖过程。

关键词: 口腔鳞状细胞癌, 长链非编码RNA, 糖酵解, 细胞增殖

Abstract:

Objective To investigate the long non-coding RNA lnc-p26090 expression level in oral squamous cell carcinoma (OSCC) tissues and cell lines. To explore the effect of lnc-p26090 on OSCC cells glycolysis and proliferation.Methods Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the relative expression level of lnc-p26090 in OSCC tissues and cell lines. Then we searched the UCSC and LNCipedia websites to obtain the chromosome location and protein coding capacity of lnc-p26090. Moreover, we transfected the plasmid into HEK 293T cell to confirm the protein coding capacity of lnc-p26090. After silencing lnc-p26090 in OSCC cell lines, we detected the glucose consumption, lactic acid production and expression level of glycolysis related genes. Furthermore, cell counting kit-8 (CCK-8) assay, flow cytometry and western blot were performed to examine the effect of lnc-p26090 on proli-feration and cell cycle distribution.Results lnc-p26090 was highly expressed both in OSCC tissues and cell lines. lnc-p26090 located in chro-mosome 2p11.2, without protein coding capacity. Silencing lnc-p26090 significantly repressed the glucose consumption, lactic acid production of cells. Moreover, the expression level of glycolysis related genes: pyruvate kinase M2, phosphofructokinase M, hexokinase 2, glucose transporter 1 and lactate dehydrogenase A were decreased after lnc- p26090 knockdown in cells. Furthermore, when downregulating lnc-p26090 in cells, proliferation was inhibited, and cell-cycle was blocked in G1 phase. At the meanwhile, cyclin D1 and cyclin-dependent kinase 4 expression level was decreased and p21 was increased significantly.Conclusion lnc-p26090, which located in chromosome 2p11.2, was upregulated in OSCC tissues and cell lines. It could be involved in the glycolysis and proliferation of OSCC.

Key words: oral squamous cell carcinoma, long non-coding RNA, glycolysis, proliferation

中图分类号: 

  • R739.8

表 1

RT-qPCR引物及扩增产物信息"

基因 上游引物（5’- 3’） 下游引物（5’- 3’） 扩增产物大小/bp
lnc-p26090 TGCCCTGTGATTATCCGCAAAC CAGATGGCGGGAAGATGAAGAC 124
PKM2 CCACTTGCAATTATTTGAGGAA GTGAGCAGACCTGCCAGACT 153
PFKM GCCAGTCTAATTGCCGTTCC TACCAACTCGAACCACAGCC 174
HK2 TCAGATTGAGAGTGACTGCC TTTCTCGTATCCTGTCCACC 179
GLUT1 CCATCCTTCCTGCTATCCTAC GACATCCTTGCACTCTCATC 178
LDHA GTGGCTTGGAAGATAAGTGG CATACAGGCACACTGGAATC 167
GAPDH ACCACAGTCCATGCCATCAC TCCACCACCCTGTTGCTGTA 165

图 1

lnc-p26090的表达情况 A：在NOK细胞及OSCC细胞中的表达,*P<0.05,**P<0.01,***P<0.001;B：在14例OSCC组织及其癌旁组织中的表达。"

图 2

lnc-p26090在染色体中的定位及编码蛋白质能力A：UCSC网站显示lnc-p26090在人类染色体上的具体定位;B：LNCipedia网站分析lnc-p26090编码蛋白质潜能;C：HEK293T细胞转染3种重组质粒后GFP的表达情况。"

图 3

lnc-p26090对HSC-3、SCC-25细胞糖酵解的影响 A：转染效率;B：转染对细胞糖酵解相关基因表达的影响;C：转染后细胞的葡萄糖消耗量和乳酸产量;*P<0.05,**P<0.01,***P<0.001。"

图 4

lnc-p26090对HSC-3、SCC-25细胞生长增殖的影响 A：CCK-8法检测结果;B：流式细胞术检测结果;C：蛋白质印迹法检测结果;*P<0.05,**P<0.01,***P<0.001。"

[1] Choi S, Myers JN . Molecular pathogenesis of oral squamous cell carcinoma: implications for therapy[J]. J Dent Res, 2008,87(1):14-32.
doi: 10.1177/154405910808700104 pmid: 18096889
[2] Ayala FR, Rocha RM, Carvalho KC , et al. GLUT1 and GLUT3 as potential prognostic markers for oral squamous cell carcinoma[J]. Molecules, 2010,15(4):2374-2387.
doi: 10.3390/molecules15042374 pmid: 20428049
[3] Wang Y, Zhang X, Zhang Y , et al. Overexpression of pyruvate kinase M2 associates with aggressive clini-copathological features and unfavorable pro-gnosis in oral squamous cell carcinoma[J]. Cancer Biol Ther, 2015,16(6):839-845.
doi: 10.1080/15384047.2015.1030551 pmid: 25970228
[4] Bergmann JH, Spector DL . Long non-coding RNAs: modulators of nuclear structure and function[J]. Curr Opin Cell Biol, 2014,26:10-18.
doi: 10.1016/j.ceb.2013.08.005
[5] Cech TR, Steitz JA . The noncoding RNA revolution-trashing old rules to forge new ones[J]. Cell, 2014,157(1):77-94.
doi: 10.1016/j.cell.2014.03.008 pmid: 24679528
[6] Martens-Uzunova ES, Böttcher R, Croce CM , et al. Long noncoding RNA in prostate, bladder, and kid-ney cancer[J]. Eur Urol, 2014,65(6):1140-1151.
doi: 10.1016/j.eururo.2013.12.003 pmid: 24373479
[7] Zhang X, Li Y, Li X , et al. Long non-coding RNA P4713 contributes to the malignant phenotypes of oral squamous cell carcinoma by activating the JAK/STAT3 pathway[J]. Int J Clin Exp Pathol, 2017,10(11):10947-10958.
[8] Hanahan D, Weinberg RA . Hallmarks of cancer: the next generation[J]. Cell, 2011,144(5):646-674.
doi: 10.1016/j.cell.2011.02.013 pmid: 21376230
[9] El Mjiyad N, Caro-Maldonado A, Ramírez-Peinado S , et al. Sugar-free approaches to cancer cell killing[J]. Oncogene, 2011,30(3):253-264.
doi: 10.1038/onc.2010.466 pmid: 20972457
[10] Levine AJ, Puzio-Kuter AM . The control of the meta- bolic switch in cancers by oncogenes and tumor sup-pressor genes[J]. Science, 2010,330(6009):1340-1344.
doi: 10.1126/science.1193494
[11] Gammon L, Biddle A, Heywood HK , et al. Sub-sets of cancer stem cells differ intrinsically in their pat-terns of oxygen metabolism[J]. PLoS One, 2013,8(4):e62493.
doi: 10.1371/journal.pone.0062493 pmid: 23638097
[12] Bochenek G, Häsler R, El Mokhtari NE , et al. The large non-coding RNA ANRIL, which is associated with atherosclerosis, periodontitis and several forms of cancer, regulates ADIPOR1, VAMP3 and C11ORF10[J]. Hum Mol Genet, 2013,22(22):4516-4527.
doi: 10.1093/hmg/ddt299
[13] Goncalves MD, Cantley LC . A glycolysis outsider steps into the cancer spotlight[J]. Cell Metab, 2018,28(1):3-4.
doi: 10.1016/j.cmet.2018.06.017
[14] Coelho RG, Fortunato RS, Carvalho DP . Metabolic reprogramming in thyroid carcinoma[J]. Front Oncol, 2018,8:82.
doi: 10.3389/fonc.2018.00082
[15] Rupaimoole R, Lee J, Haemmerle M , et al. Long noncoding RNA ceruloplasmin promotes cancer growth by altering glycolysis[J]. Cell Rep, 2015,13(11):2395-2402.
doi: 10.1016/j.celrep.2015.11.047 pmid: 4691557
[16] Bian Z, Zhang J, Li M , et al. LncRNA-FEZF1-AS1 promotes tumor proliferation and metastasis in co-lorectal cancer by regulating PKM2 signaling[J]. Clin Cancer Res, 2018. doi: 10.1158/1078-0432.CCR-17- 2967.
doi: 10.1158/1078-0432.CCR-17-2967 pmid: 29914894
[17] Johnson DG, Walker CL . Cyclins and cell cycle checkpoints[J]. Annu Rev Pharmacol Toxicol, 1999,39:295-312.
doi: 10.1146/annurev.pharmtox.39.1.295 pmid: 10331086
[18] Bloom J, Cross FR . Multiple levels of cyclin speci-ficity in cell-cycle control[J]. Nat Rev Mol Cell Biol, 2007,8(2):149-160.
doi: 10.1038/nrm2105 pmid: 17245415
[19] Yao F, Zhao T, Zhong C , et al. LDHA is necessary for the tumorigenicity of esophageal squamous cell carcinoma[J]. Tumour Biol, 2013,34(1):25-31.
doi: 10.1007/s13277-012-0506-0 pmid: 22961700
[20] Xiao H, Wang J, Yan W , et al. GLUT1 regulates cell glycolysis and proliferation in prostate cancer[J]. Prostate, 2018,78(2):86-94.
doi: 10.1002/pros.23448 pmid: 29105798
[1] 周金阔,张晋弘,史晓晶,刘广顺,姜磊,刘倩峰. 长链非编码RNA小核仁RNA宿主基因22调控微小RNA-27b-3p对口腔鳞状细胞癌细胞增殖、侵袭和迁移的影响[J]. 国际口腔医学杂志, 2024, 51(1): 52-59.
[2] 李立恒,王蕊,王晓明,张智轶,张璇,安峰,王芹,张凡. 环状RNA hsa_circ_0085576调控微小RNA-498/B细胞特异性莫洛尼鼠白血病病毒整合位点1轴对口腔鳞状细胞癌细胞迁移和侵袭的影响[J]. 国际口腔医学杂志, 2024, 51(1): 60-67.
[3] 吴佳敏,夏斌,杨禾丰,许彪. 癌相关成纤维细胞在口腔鳞状细胞癌微环境中作用的研究进展[J]. 国际口腔医学杂志, 2023, 50(6): 711-717.
[4] 柳江龙, 买买提吐逊·吐尔地. 超声造影在口腔鳞状细胞癌颈部转移性淋巴结诊断中的研究进展[J]. 国际口腔医学杂志, 2023, 50(5): 514-520.
[5] 盛南宁,王珏,南欣荣. 性别决定基因盒9在口腔鳞状细胞癌作用机制和治疗中的研究进展[J]. 国际口腔医学杂志, 2023, 50(3): 314-320.
[6] 李潭,梁新华. 盘状蛋白结构域受体1在调控恶性肿瘤进展和治疗中的作用[J]. 国际口腔医学杂志, 2023, 50(2): 230-236.
[7] 赵卓平,辛鹏飞,高阳,张彩凤,张宽收,刘青梅. 光热治疗在口腔鳞状细胞癌治疗中的研究进展[J]. 国际口腔医学杂志, 2022, 49(4): 462-470.
[8] 洪娅娅,陈学鹏,姒蜜思. 非编码RNA调控牙囊干细胞成骨分化的研究进展[J]. 国际口腔医学杂志, 2022, 49(3): 263-271.
[9] 江涵,神应强,陈谦明. 毒蕈碱受体通过Yes相关蛋白信号对口腔鳞状细胞癌生物学行为的实验研究[J]. 国际口腔医学杂志, 2022, 49(2): 138-143.
[10] 蒋宇磊,夏斌,饶南荃,杨禾丰,许彪. 外泌体在口腔鳞状细胞癌恶性进展及诊疗应用的研究[J]. 国际口腔医学杂志, 2021, 48(6): 711-717.
[11] 刘娟,陈斌,闫福华. 富血小板血浆和浓缩生长因子对人牙周膜细胞增殖和成骨分化影响的研究[J]. 国际口腔医学杂志, 2021, 48(5): 520-527.
[12] 甘建国,高攀,王晓毅. 循环肿瘤细胞与口腔鳞状细胞癌相关性的研究进展[J]. 国际口腔医学杂志, 2021, 48(2): 205-212.
[13] 黄俊文,乔洁,梅子,陈茁,李杨,乔彬. 脂多糖结合蛋白在口腔鳞状细胞癌中的表达及其临床意义[J]. 国际口腔医学杂志, 2021, 48(1): 50-57.
[14] 何宇晴,但红霞,陈谦明. 光动力疗法在口腔黏膜癌变防治中的应用[J]. 国际口腔医学杂志, 2020, 47(6): 669-676.
[15] 郝福,宁毅,孙睿,郑晓旭. 口腔鳞状细胞癌中转化因子2β的表达及潜在的临床意义[J]. 国际口腔医学杂志, 2020, 47(2): 159-165.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
[1] 王昆润. 修补颌骨缺损的新型生物学相容材料[J]. 国际口腔医学杂志, 1999, 26(06): .
[2] 陆加梅. 不可复性关节盘移位患者术前张口度与关节镜术后疗效的相关性[J]. 国际口腔医学杂志, 1999, 26(06): .
[3] 王昆润. 咀嚼口香糖对牙周组织微循环的影响[J]. 国际口腔医学杂志, 1999, 26(06): .
[4] 宋红. 青少年牙周炎外周血分叶核粒细胞的趋化功能[J]. 国际口腔医学杂志, 1999, 26(06): .
[5] 高卫民,李幸红. 发达国家牙医学院口腔种植学教学现状[J]. 国际口腔医学杂志, 1999, 26(06): .
[6] 侯锐. 正畸患者釉白斑损害的纵向激光荧光研究[J]. 国际口腔医学杂志, 1999, 26(05): .
[7] 轩东英. 不同赋形剂对氢氧化钙抗菌效果的影响[J]. 国际口腔医学杂志, 1999, 26(05): .
[8] 房兵. 唇腭裂新生儿前颌骨矫正方法及对上颌骨生长发育的影响[J]. 国际口腔医学杂志, 1999, 26(05): .
[9] 杨美祥. 前牙厚度在预测上下颌牙量协调性中的作用[J]. 国际口腔医学杂志, 1999, 26(04): .
[10] 赵艳丽. 手术刀、电凝、CO_2和KTP激光对大鼠舌部创口的作用[J]. 国际口腔医学杂志, 1999, 26(04): .