三阴性乳腺癌细胞系MDA-MB-231与其脑转移细胞系中microRNA差异表达分析

doi:10.3969/j.issn.1004-616x.2021.03.011

癌变·畸变·突变 ›› 2021, Vol. 33 ›› Issue (3): 218-224.doi: 10.3969/j.issn.1004-616x.2021.03.011

三阴性乳腺癌细胞系MDA-MB-231与其脑转移细胞系中microRNA差异表达分析

罗娇^1,3, 张坤驰^2,4, 周莉^1,3

1. 贵州医科大学临床医学院, 贵州贵阳 550025;
2. 上海健康医学院上海市分子影像学重点实验室, 上海 201318;
3. 上海健康医学院附属周浦医院肿瘤血液科, 上海 201318;
4. 贵州工程职业学院, 贵州铜仁 554300

收稿日期:2020-08-27 修回日期:2021-02-24 出版日期:2021-05-30 发布日期:2021-06-09
通讯作者: 周莉，E-mail：lzhou_sh@hotmail.com E-mail:lzhou_sh@hotmail.com
作者简介:罗娇，E-mail：l287923@163.com。
基金资助:
浦东新区学科带头人培养项目（PWRd2017-02）；上海市自然科学基金面上项目（18ZR1417800）；国家自然科学基金（81801833）；贵州省科技计划项目（黔科合基础[2019]1129号）

Analysis of differentially expressed microRNAs between the triple negative breast cancer cell line MDA-MB-231 and its brain metastasis cell line

LUO Jiao^1,3, ZHANG Kunchi^2,4, ZHOU Li^1,3

1. Clinical Medicine, Guizhou Medical University, Guiyang 550025, Guizhou;
2. Shanghai Key Laboratory for Molecular Imaging, Shanghai University of Medicine & Health Sciences, Shanghai 201318;
3. Department of Oncology and Hematology, Shanghai University of Medicine & Health Sciences Affiliated Zhoupu Hospital, Shanghai 201318;
4. Guizhou Engineering Vocational College, Tongren 554300, Guizhou, China

Received:2020-08-27 Revised:2021-02-24 Online:2021-05-30 Published:2021-06-09

摘要/Abstract

摘要： 目的：分析三阴性乳腺癌（TNBC）细胞系MDA-MB-231（MB-231）与其脑转移细胞系MDA-MB-231Brm （MB-231Brm）中microRNA （miRNA）的表达差异，寻找有价值的三阴性乳腺癌脑转移相关的miRNA。方法：Trizol法提取MB-231和MB-231Brm细胞总RNA，用Illumina高通量测序技术分离并检测miRNA表达情况，将其与已知的miRNA数据库比对，采用miRDeep2软件进行新的miRNA预测；使用差异基因检测法筛选差异表达的miRNA，构建差异表达谱。应用生物信息学分析进行miRNA靶基因的功能分析，并用实时荧光定量PCR （qPCR）进行验证。结果：MB-231和MB-231Brm细胞中miRNA的种类和表达存在差异，两者共有的miRNA为961种，前者自有164种，后者自有196种；与MB-231相比，MB-231Brm细胞的miRNA中，有145个表达显著上调，54个表达显著下调，1 122个表达未见明显差异；差异表达分析显示，与MB-231相比，MB-231Brm中的miR-199a-3p和miR-103a-3p等表达显著增加，而miR-1246和miR-4787-5p等表达显著下降；基因本体分析（GO）发现差异表达的miRNA所调控的靶基因与细胞组分、分子功能及生物过程均相关；qPCR验证发现，与MB-231相比，miR-1246在MB-231Brm中表达显著降低且差异有统计学意义（P < 0.05）。结论：MB-231和MB-231Brm中存在差异表达的miRNA，且在TNBC脑转移过程中其调控的靶基因通过参与细胞组分形成、调节分子功能及调控生物过程等发挥作用；并发现miR-1246在脑转移细胞系中的表达下降，其可能通过某种途径参与TNBC脑转移的发生、发展过程。

关键词: 三阴性乳腺癌, 脑转移, 差异表达, 微小RNA, miR-1246

Abstract: OBJECTIVE： To analyze differentially expressed miRNAs between the triple negative breast cancer cell line MDA-MB-231 (MB-231) and its brain metastasis cell line MDA-MB-231Brm (MB-231Brm)， and to screen biomarkers associated with the brain metastases. METHODS： The Trizol method was used to extract miRNA from MB-231 and MB-231Brm，and PCR was used to construct RNA libraries. After filtering the data and comparing to known miRNA databases，the miRDeep2 method was used to make new miRNA predictions for unknown miRNAs. Then， a differential gene detection method was used to screen the differentially expressed miRNAs. Finally，bioinformatics analysis tools was used to perform functional analysis of the target genes which could be regulated by these miRNAs. Quantitative PCR (qPCR) analyses were used to validate the microarray data. RESULTS： There were differences in the types and expressions of miRNAs between the MB-231 and the MB-231Brm cells. Briefly， 961 kinds of miRNA were shared by both， 164 species of miRNAs only existed in MB-231， and 196 species of miRNAs only existed in MB-231Brm. Compared with MB-231，145 expressions were significantly up-regulated in MB-231Brm，and 64 expressions were significantly down-regulated，and there were no significant differences in 1 122 expressions. Results from differential expression analyses demonstrate that expressions of miR-199a-3p were significantly upregulated in MB-231Brm compared with MB-231 while miR-1246 and miR-4787-5p were down-regulated significantly. GO analyses reveal that genes which were regulated by differentially expressed miRNAs were related to cellular components，molecular functions and biological processes. Results from qPCR analyses confirmed that expressions of miR-1246 were down- regulated in MB-231Brm， compared with MB-231 (P < 0.05). CONCLUSION： There were differentially expressed miRNAs in MB-231 and MB-231Brm cells， and genes that were targeted by these miRNAs were associated with cellular components， molecular functions and biological processes during TNBC brain metastasis. Moreover， expressions of miR-1246 were down-regulated in MB-231Brm，compared with MB-231. Therefore， miR-1246 may participate in the occurrence and development of TNBC brain metastasis.

Key words: triple negative breast cancer, brain metastasis, differentially expression, microRNA, miR-1246

中图分类号:

R737.9

罗娇, 张坤驰, 周莉. 三阴性乳腺癌细胞系MDA-MB-231与其脑转移细胞系中microRNA差异表达分析[J]. 癌变·畸变·突变, 2021, 33(3): 218-224.

LUO Jiao, ZHANG Kunchi, ZHOU Li. Analysis of differentially expressed microRNAs between the triple negative breast cancer cell line MDA-MB-231 and its brain metastasis cell line[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2021, 33(3): 218-224.

参考文献

[1] CHEN W, ZHENG R, BAADE P D, et al. Cancer statistics in China, 2015[J]. CA Cancer J Clin, 2016, 66(2): 115-132.
[2] BIANCHINI G, BALKO J M, MAYER I A, et al. Triplenegative breast cancer: challenges and opportunities of a heterogeneous disease[J]. Nat Rev Clin Oncol, 2016, 13(11): 674-690.
[3] LEE A, DJAMGOZ M B A. Triple negative breast cancer: Emerging therapeutic modalities and novel combination therapies[J]. Cancer Treat Rev, 2018, 62: 110-122.
[4] BAUER K R, BROWN M, CRESS R D, et al. Descriptive analysis of estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and HER2-negative invasive breast cancer, the so-called triple-negative phenotype: a populationbased study from the California Cancer Registry[J]. Cancer, 2007, 109(9): 1721-1728.
[5] KWAN M L, KUSHI L H, WELTZIEN E, et al. Epidemiology of breast cancer subtypes in two prospective cohort studies of breast cancer survivors[J]. Breast Cancer Res, 2009, 11(3): R31.
[6] LIN N U, CLAUS E, SOHL J, et al. Sites of distant recurrence and clinical outcomes in patients with metastatic triplenegative breast cancer: high incidence of central nervous system metastases[J]. Cancer, 2008, 113(10): 2638-2645.
[7] DAWOOD S, LEI X D, LITTON J K, et al. Incidence of brain metastases as a first site of recurrence among women with triple receptor-negative breast cancer[J]. Cancer, 2012, 118(19): 4652-4659.
[8] MARTIN A M, CAGNEY D N, CATALANO P J, et al. Brain metastases in newly diagnosed breast cancer: a populationbased study[J]. JAMA Oncol, 2017, 3(8): 1069-1077.
[9] ACHROL A S, RENNERT R C, ANDERS C, et al. Brain metastases[J]. Nat Rev Dis Primers, 2019, 5(1): 1-26.
[10] TSENG L M, HSU N C, CHEN S C, et al. Distant metastasis in triple-negative breast cancer[J]. Neoplasma, 2013, 60(3): 290-294.
[11] MORIKAWA A, WANG R, PATIL S, et al. Characteristics and prognostic factors for patients with HER2-overexpressing breast cancer and brain metastases in the era of HER2-targeted therapy: an argument for earlier detection[J]. Clin Breast Cancer, 2018, 18(5): 353-361.
[12] SALVATI M, TROPEANO M P, MAIOLA V, et al. Multiple brain metastases: a surgical series and neurosurgical perspective [J]. Neurol Sci, 2018, 39(4): 671-677.
[13] BOWMAN K M, KUMTHEKAR P. Medical management of brain metastases and leptomeningeal disease in patients with breast carcinoma[J]. Future Oncol, 2018, 14(4): 391-407.
[14] PIASECKA D, BRAUN M, KORDEK R, et al. MicroRNAs in regulation of triple-negative breast cancer progression[J]. J Cancer Res Clin Oncol, 2018, 144(8): 1401-1411.
[15] KONG P, CHEN L, YU M X, et al. miR-3178 inhibits cell proliferation and metastasis by targeting Notch1 in triplenegative breast cancer[J]. Cell Death Dis, 2018, 9(11): 1059.
[16] CHU J H, LI Y F, FAN X M, et al. MiR-4319 suppress the malignancy of triple-negative breast cancer by regulating selfrenewal and tumorigenesis of stem cells[J]. Cell Physiol Biochem, 2018, 48(2): 593-604.
[17] HONG Z P, HONG C Y, MA B, et al. MicroRNA-126-3p inhibits the proliferation, migration, invasion, and angiogenesis of triple-negative breast cancer cells by targeting RGS3[J]. Oncol Rep, 2019, 42(4): 1569-1579.
[18] LEGENDRE M, AUDIC S, POIROT O, et al. mRNA deep sequencing reveals 75 new genes and a complex transcriptional landscape in Mimivirus[J]. Genome Res, 2010, 20(5): 664-674.
[19] VAN'T VEER L J, DAI H Y, VAN DE VIJVER M J, et al. Gene expression profiling predicts clinical outcome of breast cancer[J]. Nature, 2002, 415(6871): 530-536.
[20] MA X J, DAHIYA S, RICHARDSON E, et al. Gene expression profiling of the tumor microenvironment during breast cancer progression[J]. Breast Cancer Res, 2009, 11(1): R7.
[21] REIS-FILHO J S, PUSZTAI L. Gene expression profiling in breast cancer: classification, prognostication, and prediction[J]. Lancet, 2011, 378(9805): 1812-1823.
[22] ZOHRABIAN V M, NANDU H, GULATI N, et al. Gene expression profiling of metastatic brain cancer[J]. Oncol Rep, 2007, 18(2): 321-328.
[23] ZHOU L, GAO H F, LIU D S, et al. Gene expression profiling of brain metastatic cell from triple negative breast cancer: Understanding the molecular events[J]. Gene, 2018, 640: 21-27.
[24] ZHANG Y, LIAO J M, ZENG S X, et al. p53 downregulates Down syndrome-associated DYRK1A through miR-1246[J]. EMBO Rep, 2011, 12(8): 811-817.
[25] LIAO J M, ZHOU X, ZHANG Y, et al. MiR-1246: a new link of the p53 family with cancer and Down syndrome[J]. Cell Cycle, 2012, 11(14): 2624-2630.
[26] MALLA R R, KUMARI S, GAVARA M M, et al. A perspective on the diagnostics, prognostics, and therapeutics of microRNAs of triple-negative breast cancer[J]. Biophys Rev, 2019, 11(2): 227-234.
[27] TAKESHITA N, HOSHINO I, MORI M, et al. Serum microRNA expression profile: miR-1246 as a novel diagnostic and prognostic biomarker for oesophageal squamous cell carcinoma[J]. Br J Cancer, 2013, 108(3): 644-652.
[28] SUN Z, MENG C T, WANG S H, et al. MicroRNA-1246 enhances migration and invasion through CADM1 in hepatocellular carcinoma[J]. BMC Cancer, 2014, 14: 616.
[29] HUANG W H, LI H F, LUO R C. The microRNA-1246 promotes metastasis in non-small cell lung cancer by targeting cytoplasmic polyadenylation element-binding protein 4[J]. Diagn Pathol, 2015, 10: 127.
[30] YUAN D X, XU J P, WANG J, et al. Extracellular miR-1246 promotes lung cancer cell proliferation and enhances radioresistance by directly targeting DR5[J]. Oncotarget, 2016, 7(22): 32707-32722.
[31] LIN S S, PENG C Y, LIAO Y W, et al. miR-1246 targets CCNG2 to enhance cancer stemness and chemoresistance in oral carcinomas[J]. Cancers (Basel), 2018, 10(8): E272.
[32] TODESCHINI P, SALVIATO E, PARACCHINI L, et al. Circulating miRNA landscape identifies miR-1246 as promising diagnostic biomarker in high-grade serous ovarian carcinoma: a validation across two independent cohorts[J]. Cancer Lett, 2017, 388: 320-327.
[33] HANNAFON B N, TRIGOSO Y D, CALLOWAY C L, et al. Plasma exosome microRNAs are indicative of breast cancer[J]. Breast Cancer Res, 2016, 18(1): 90.
[34] LI X J, REN Z J, TANG J H, et al. Exosomal microRNA MiR-1246 promotes cell proliferation, invasion and drug resistance by targeting CCNG2 in breast cancer[J]. Cell Physiol Biochem, 2017, 44(5): 1741-1748.
[35] LIAO J M, ZHOU X, ZHANG Y, et al. MiR-1246: a new link of the p53 family with cancer and Down syndrome[J]. Cell Cycle, 2012, 11(14): 2624-2630.
[36] ZHANG Q, CAO L Y, CHENG S J, et al. p53-induced microRNA-1246 inhibits the cell growth of human hepatocellular carcinoma cells by targeting NFIB[J]. Oncol Rep, 2015, 33(3): 1335-1341.
[37] SHAH S P, ROTH A, GOYA R, et al. The clonal and mutational evolution spectrum of primary triple-negative breast cancers[J]. Nature, 2012, 486(7403): 395-399.
[38] WALERYCH D, NAPOLI M, COLLAVIN L, et al. The rebel angel: mutant p53 as the driving oncogene in breast cancer[J]. Carcinogenesis, 2012, 33(11): 2007-2017.
[39] TURNER N, MORETTI E, SICLARI O, et al. Targeting triple negative breast cancer: is p53 the answer?[J]. Cancer Treat Rev, 2013, 39(5): 541-550.
[40] LIU R Z, VO T M, JAIN S, et al. NFIB promotes cell survival by directly suppressing p21 transcription in TP53-mutated triple-negative breast cancer[J]. J Pathol, 2019, 247(2): 186-198.
[41] CUSTÓDIO-SANTOS T, VIDEIRA M, BRITO M A. Brain metastasization of breast cancer[J]. Biochim Biophys Acta Rev Cancer, 2017, 1868(1): 132-147.

三阴性乳腺癌细胞系MDA-MB-231与其脑转移细胞系中microRNA差异表达分析

Analysis of differentially expressed microRNAs between the triple negative breast cancer cell line MDA-MB-231 and its brain metastasis cell line

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