LncRNA LUCAT1通过靶向miR-199a-5p调控宫颈癌细胞活性、侵袭迁移及其分子机制
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摘要
目的研究长链非编码RNA肺癌相关转录产物(LUCAT)1对宫颈癌细胞的活性、迁移和侵袭的影响,并探讨其机制。方法运用qRT-PCR法检测宫颈癌细胞Hela、正常宫颈细胞Ect1/E6E7中LUCAT1、miR-199a-5p的表达;将si-con组(转染si-con)、si-LUCAT1组(转染si-LUCAT1)、miR-199a-5p组(转染miR-199a-5p模拟物)、miR-NC组(转染miR-NC)、si-con+anti-miR-199a-5p组(共转染si-con和anti-miR-199a-5p)、si-LUCAT1+anti-miR-199a-5p组(共转染si-LUCAT1和anti-miR-199a-5p),均用脂质体法转染至Hela细胞;噻唑蓝(MTT)法检测各组细胞的活性;Transwell法检测各组细胞的迁移和侵袭;双荧光素酶报告基因检测实验检测各组细胞的荧光活性。结果与正常宫颈细胞Ect1/E6E7相比,宫颈癌细胞Hela中LUCAT1表达显著升高,miR-199a-5p表达显著降低;敲减LUCAT1、过表达miR-199a-5p均可明显抑制Hela细胞的活性、迁移和侵袭;miR-199a-5p可抑制野生型LUCAT1细胞的荧光活性,且LUCAT1可负向调控miR-199a-5p的表达;抑制miR-199a-5p可逆转敲减LUCAT1对Hela细胞的活性、迁移、侵袭的抑制作用。结论 LUCAT1可促进宫颈癌细胞的活性、迁移、侵袭,其机制可能与靶向miR-199a-5p有关,将可为宫颈癌的治疗提供靶点。
Objective To study the effects of long-chain non-coding(Lnc) RNA lung cancer-associated transcript(LUCAT)1 on the activity, migration and invasion of cervical cancer cells and explore its mechanism.Methods qRT-PCR was used to detect the expression of LUCAT1 and miR-199 a-5 p in cervical cancer Hela and normal cervical cells Ect1/E6 E7. si-con(si-con group), si-LUCAT1(si-LUCAT1 group), miR-199 a-5 p mimics(miR-199 a-5 p group), miR-NC(miR-NC group), si-con and anti-miR-199 a-5 p(si-con+anti-miR-199 a-5 p group), si-LUCAT1+anti-miR-199 a-5 p(si-LUCAT1 and anti-miR-199 a-5 p group) were transfected into Hela cells by liposome respectively. The activity of each group was detected by MTT assay, the migration and invasion of each group was detected by transwell method, the fluorescence activity of each group was detected by dual luciferase reporter gene assay.Results Compared with normal cervical cells Ect1/E6 E7, the expression of LUCAT1 in cervical cancer cells were significantly increased and the expression of miR-199 a-5 p were significantly decreased. Knockdown of LUCAT1 or overexpression of miR-199 a-5 p significantly inhibited the activity, migration and invasion of Hela cells; miR-199 a-5 p could inhibit the fluorescence activity of wild-type LUCAT1 cells and LUCAT1 could negatively regulate the expression of miR-199 a-5 p. Inhibition of miR-199 a-5 p could reverse the effect of the LUCAT1 knockdown on the inhibition of cell activity, migration and invasion of Hela cells.Conclusions Lnc RNA LUCAT1 could promote the activity, migration and invasion of cervical cancer cells. The mechanism might be related to targeting miR-199 a-5 p, which will provide a target for the treatment of cervical cancer.
Objective To study the effects of long-chain non-coding(Lnc) RNA lung cancer-associated transcript(LUCAT)1 on the activity, migration and invasion of cervical cancer cells and explore its mechanism.Methods qRT-PCR was used to detect the expression of LUCAT1 and miR-199 a-5 p in cervical cancer Hela and normal cervical cells Ect1/E6 E7. si-con(si-con group), si-LUCAT1(si-LUCAT1 group), miR-199 a-5 p mimics(miR-199 a-5 p group), miR-NC(miR-NC group), si-con and anti-miR-199 a-5 p(si-con+anti-miR-199 a-5 p group), si-LUCAT1+anti-miR-199 a-5 p(si-LUCAT1 and anti-miR-199 a-5 p group) were transfected into Hela cells by liposome respectively. The activity of each group was detected by MTT assay, the migration and invasion of each group was detected by transwell method, the fluorescence activity of each group was detected by dual luciferase reporter gene assay.Results Compared with normal cervical cells Ect1/E6 E7, the expression of LUCAT1 in cervical cancer cells were significantly increased and the expression of miR-199 a-5 p were significantly decreased. Knockdown of LUCAT1 or overexpression of miR-199 a-5 p significantly inhibited the activity, migration and invasion of Hela cells; miR-199 a-5 p could inhibit the fluorescence activity of wild-type LUCAT1 cells and LUCAT1 could negatively regulate the expression of miR-199 a-5 p. Inhibition of miR-199 a-5 p could reverse the effect of the LUCAT1 knockdown on the inhibition of cell activity, migration and invasion of Hela cells.Conclusions Lnc RNA LUCAT1 could promote the activity, migration and invasion of cervical cancer cells. The mechanism might be related to targeting miR-199 a-5 p, which will provide a target for the treatment of cervical cancer.
引文
1 Waggoner SE.Cervical cancer[J].Lancet,2003;361(9376):2217-25.
2 Brodersen J,Siersma V,Thorsen H.Consequences of screening in cervical cancer:development and dimensionality of a questionnaire[J].BMC Psychol,2018;6(1):39.
3 Kaneko N.Factors associated with cervical cancer screening among young unmarried Japanese women:results from an internet-based survey[J].BMC Womens Health,2018;18(1):132.
4 Jr SW,Bacon MA,Bajaj A,et al.Cervical cancer:a global health crisis[J].Cancer,2017;123(13):2404-12.
5 Brosnan CA,Voinnet O.The long and the short of noncoding RNAs[J].Curr Opin Cell Biol,2009;21(3):416-25.
6 Guttman M,Amit I,Garber M,et al.Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals[J].Nature,2009;458(7235):223-7.
7 沈钲杰,朱银银,沈凡含,等.lncRNA与miRNA调节关系和ceRNA关系在疾病中的研究进展[J].中国细胞生物学学报,2018;40(10):1774-80.
8 涂超峰,武明花,李桂源.LncRNA与miRNA相互调控作用及其与肿瘤的关系研究[J].中国生物化学与分子生物学报,2013;29(11):1029-34.
9 Zhan L,Jun L,Bing W.Long non-coding RNAs in ovarian cancer[J].J Exp Clin Cancer Res,2018;27(1):120.
10 Zheng ZG,Xu H,Suo SS,et al.The essential role of H19 contributing to cisplatin resistance by regulating glutathione metabolism in high-grade serous ovarian cancer[J].Sci Rep,2016;6:26093.
11 尹凌帝,孙倩,孙明,等.人类癌症中长链非编码RNA与miRNA相互作用的研究进展[J].临床肿瘤学杂志,2014;19(7):662-6.
12 Thai P,Statt S,Chen CH,et al.Characterization of a novel long noncoding RNA,SCAL1,induced by cigarette smoke and elevated in lung cancer cell lines[J].Am J Respir Cell Mol Biol,2013;49(2):204-11.
13 Yoon JH,You BH,Park CH,et al.The long noncoding RNA LUCAT1 promotes tumorigenesis by controlling ubiquitination and stability of DNA methyltransferase 1 in esophageal squamous cell carcinoma[J].Cancer Lett,2017;417:47-57.
14 Yu H,Xu Y,Zhang D,et al.Long noncoding RNA LUCAT1 promotes malignancy of ovarian cancer through regulation of miR-612/HOXA13 pathway[J].Biochem Biophys Res Commun,2018;503(3):2095-100.
15 于贵林,林杰,赵国华,等.长链非编码RNA lucat1在胃癌组织中的表达及其临床意义[J].现代肿瘤医学,2019;16(4):607-11.
16 Vishnoi A,Rani S.MiRNA biogenesis and regulation of diseases:an overview[J].Methods Mol Biol,2017;1509(1):1-10.
17 Dong ZQ,Qing ZQ,Lemei D,et al.MiR-199a-5p inhibits the growth and metastasis of colorectal cancer cells by targeting ROCK1[J].Technol Cancer Res Treat,2018;17:1533034618775509.
18 Ma S,Jia W,Ni S.miR-199a-5p inhibits the progression of papillary thyroid carcinoma by targeting SNAI1[J].Biochem Biophys Res Commun,2018;497(1):181-6.
19 Sakurai K,Furukawa C,Haraguchi T,et al.MicroRNAs miR-199a-5p and-3p target the brm subunit of SWI/SNF to generate a double-negative feedback loop in a variety of human cancers[J].Cancer Res,2011;71(5):1680-9.
20 Lee JW,Choi CH,Choi JJ,et al.Altered MicroRNA expression in cervical carcinomas[J].Clin Cancer Res,2008;14(9):2535-42.
21 Pereira PM,Marques JP,Soares AR,et al.MicroRNA expression variability in human cervical tissues[J].PLoS One,2010;5(7):e11780.
22 邹阮敏,胡芝,陈昊,等.miR-199a在宫颈癌及宫颈上皮内瘤样病变中的表达及意义[J].医学研究杂志,2011;40(7):55-9.
2 Brodersen J,Siersma V,Thorsen H.Consequences of screening in cervical cancer:development and dimensionality of a questionnaire[J].BMC Psychol,2018;6(1):39.
3 Kaneko N.Factors associated with cervical cancer screening among young unmarried Japanese women:results from an internet-based survey[J].BMC Womens Health,2018;18(1):132.
4 Jr SW,Bacon MA,Bajaj A,et al.Cervical cancer:a global health crisis[J].Cancer,2017;123(13):2404-12.
5 Brosnan CA,Voinnet O.The long and the short of noncoding RNAs[J].Curr Opin Cell Biol,2009;21(3):416-25.
6 Guttman M,Amit I,Garber M,et al.Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals[J].Nature,2009;458(7235):223-7.
7 沈钲杰,朱银银,沈凡含,等.lncRNA与miRNA调节关系和ceRNA关系在疾病中的研究进展[J].中国细胞生物学学报,2018;40(10):1774-80.
8 涂超峰,武明花,李桂源.LncRNA与miRNA相互调控作用及其与肿瘤的关系研究[J].中国生物化学与分子生物学报,2013;29(11):1029-34.
9 Zhan L,Jun L,Bing W.Long non-coding RNAs in ovarian cancer[J].J Exp Clin Cancer Res,2018;27(1):120.
10 Zheng ZG,Xu H,Suo SS,et al.The essential role of H19 contributing to cisplatin resistance by regulating glutathione metabolism in high-grade serous ovarian cancer[J].Sci Rep,2016;6:26093.
11 尹凌帝,孙倩,孙明,等.人类癌症中长链非编码RNA与miRNA相互作用的研究进展[J].临床肿瘤学杂志,2014;19(7):662-6.
12 Thai P,Statt S,Chen CH,et al.Characterization of a novel long noncoding RNA,SCAL1,induced by cigarette smoke and elevated in lung cancer cell lines[J].Am J Respir Cell Mol Biol,2013;49(2):204-11.
13 Yoon JH,You BH,Park CH,et al.The long noncoding RNA LUCAT1 promotes tumorigenesis by controlling ubiquitination and stability of DNA methyltransferase 1 in esophageal squamous cell carcinoma[J].Cancer Lett,2017;417:47-57.
14 Yu H,Xu Y,Zhang D,et al.Long noncoding RNA LUCAT1 promotes malignancy of ovarian cancer through regulation of miR-612/HOXA13 pathway[J].Biochem Biophys Res Commun,2018;503(3):2095-100.
15 于贵林,林杰,赵国华,等.长链非编码RNA lucat1在胃癌组织中的表达及其临床意义[J].现代肿瘤医学,2019;16(4):607-11.
16 Vishnoi A,Rani S.MiRNA biogenesis and regulation of diseases:an overview[J].Methods Mol Biol,2017;1509(1):1-10.
17 Dong ZQ,Qing ZQ,Lemei D,et al.MiR-199a-5p inhibits the growth and metastasis of colorectal cancer cells by targeting ROCK1[J].Technol Cancer Res Treat,2018;17:1533034618775509.
18 Ma S,Jia W,Ni S.miR-199a-5p inhibits the progression of papillary thyroid carcinoma by targeting SNAI1[J].Biochem Biophys Res Commun,2018;497(1):181-6.
19 Sakurai K,Furukawa C,Haraguchi T,et al.MicroRNAs miR-199a-5p and-3p target the brm subunit of SWI/SNF to generate a double-negative feedback loop in a variety of human cancers[J].Cancer Res,2011;71(5):1680-9.
20 Lee JW,Choi CH,Choi JJ,et al.Altered MicroRNA expression in cervical carcinomas[J].Clin Cancer Res,2008;14(9):2535-42.
21 Pereira PM,Marques JP,Soares AR,et al.MicroRNA expression variability in human cervical tissues[J].PLoS One,2010;5(7):e11780.
22 邹阮敏,胡芝,陈昊,等.miR-199a在宫颈癌及宫颈上皮内瘤样病变中的表达及意义[J].医学研究杂志,2011;40(7):55-9.
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