miRNA-101在甲状腺髓样癌中抑癌机制的研究
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摘要
目的:研究microRNA-101在甲状腺髓样细胞中的抑癌机制。方法:首先用荧光定量PCR技术检测甲状腺髓样癌组织中microRNA-101的表达。接着采用microRNA-101模拟物干预法,并用流式细胞仪技术,Transwell实验等方法,对实验组和对照组细胞的增殖、迁移和凋亡进行比较。此外,还通过Western blot、报告基因检测法、流式细胞仪技术和Transwell实验确定miRNA-101靶基因以及相应的抑癌机制。结果:miRNA-101在甲状腺髓样癌组织中的表达相比癌旁组织下调(P<0.01)。与阴性对照组相比,miRNA-101模拟物转染组的甲状腺髓样癌细胞的增殖和迁移受到显著抑制,而凋亡率显著提高。Western blot和荧光素酶报告实验证实SOX9基因是miRNA-101下游的靶基因并被其下调。SOX9基因敲除可抑制甲状腺髓样癌细胞增殖,迁移和促进癌细胞凋亡。结论:microRNA-101通过靶向下调SOX9基因的表达抑制甲状腺髓样癌细胞的增殖和迁移,加速甲状腺髓样癌细胞的凋亡。
AIM: To explore the tumor suppression mechanism of microRNA-101 in human medullary thyroid carcinoma cells( TT cells). METHODS: The expression of microRNA-101 in tissue samples from patients with medullary thyroid carcinoma was analyzed by quantitative RT-PCR. Then TT cells were divided into two groups and transfected with miRNA-101 mimics/negative control. Flow cytometry and Transwell assay were used to detect TT cell cycle distribution,apoptotic rate as well as cell migration. Moreover,Western blot,flow cytometry,luciferase and Transwell assays were used to predict target gene of miRNA-101 as well as its underlying mechanism. RESULTS: MiRNA-101 was significantly down-regulated in medullary thyroid carcinoma tissues. MiRNA-101 mimics' transfection significantly inhibited TT cell proliferation and migration,induced cell apoptosis comparing with negative control. Western blot and luciferase assay revealed that SOX9 gene expression was down-regulated by miRNA-101. Silencing of SOX9 by siRNA showed similar effects with miRNA-101 over-expression. CONCLUSION: MicroRNA-101 inhibits proliferation,migration and induces apoptosis of human medullary thyroid carcinoma cells by targeting SOX9 gene.
AIM: To explore the tumor suppression mechanism of microRNA-101 in human medullary thyroid carcinoma cells( TT cells). METHODS: The expression of microRNA-101 in tissue samples from patients with medullary thyroid carcinoma was analyzed by quantitative RT-PCR. Then TT cells were divided into two groups and transfected with miRNA-101 mimics/negative control. Flow cytometry and Transwell assay were used to detect TT cell cycle distribution,apoptotic rate as well as cell migration. Moreover,Western blot,flow cytometry,luciferase and Transwell assays were used to predict target gene of miRNA-101 as well as its underlying mechanism. RESULTS: MiRNA-101 was significantly down-regulated in medullary thyroid carcinoma tissues. MiRNA-101 mimics' transfection significantly inhibited TT cell proliferation and migration,induced cell apoptosis comparing with negative control. Western blot and luciferase assay revealed that SOX9 gene expression was down-regulated by miRNA-101. Silencing of SOX9 by siRNA showed similar effects with miRNA-101 over-expression. CONCLUSION: MicroRNA-101 inhibits proliferation,migration and induces apoptosis of human medullary thyroid carcinoma cells by targeting SOX9 gene.
引文
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[17]Chakravarthi BVSK,Goswami MT,Pathi SS,et al.MicroRNA-101 regulated transcriptional modulator SUB1 plays a role in prostate cancer[J].Oncogene,2016,35(49):6330-6340.
[18]Huang F,Lin C,Shi YH,et al.MicroRNA-101 inhibits cell proliferation,invasion,and promotes apoptosis by regulating cyclooxygenase-2 in Hela cervical carcinoma cells[J].Asian Pac J Cancer Prev,2013,14(10):5915-5920.
[19]Long Y,Wu Z,Yang X,et al.MicroRNA-101 inhibits the proliferation and invasion of bladder cancer cells via targeting c-FOS[J].Mol Med Rep,2016,14(3):2651-2656.
[20]Wang C,Lu S,Jiang J,et al.Hsa-microRNA-101suppresses migration and invasion by targeting Rac1 in thyroid cancer cells[J].Oncology Lett,2014,8(4):1815-1821.
[21]陈维,王昌留,王磊.SOX9基因的结构、功能及进化[J].生命的化学,2012,32(4):310-315.
[22]Zhang Y,Guo X,Xiong L,et al.MicroRNA-101suppresses SOX9-dependent tumorigenicity and promotes favorable prognosis of human hepatocellular carcinoma[J].FEBS Lett,2012,586(24):4362-4370.
[23]Liu N,Zhang L,Wang Z,et al.MicroRNA-101 inhibits proliferation,migration and invasion of human glioblastoma by targeting SOX9[J].Oncotarget,2017,8(12):19244-19254.
[2]黄彩平,朱永学,田敖龙.147例甲状腺髓样癌临床分析[J].中华肿瘤杂志,2003,25(5):490-492.
[3]Lee RC,Feinbaum RL,Ambros V,et al.Elegans heterochromic gene Lin-4 encodes small RNAs with antisense complementarity to Lin-14[J].Cell,1993,75(5):843-845.
[4]Kozomara A,Griffiths-Jones S.miRBase:annotating high confidence microRNAs using deep sequencing data[J].Nucleic Acids Res,2014,42:68-73
[5]Murakami Y,Yasuda T,Saigo K,et al.Comprehensive analysis of microRNA expression patterns in hepatocellular carcinoma and non-tumorous tissues[J].Oncogene,2006,25(17):2537-2545.
[6]Lowery A J,Miller N,Devaney A,et al.MicroRNA signatures predict oestrogen receptor,progesterone receptor and HER/neu receptor status in breast cancer[J].Breast Cancer Res,2009,11(3):R27.
[7]Yanaihara N,Caplen N,Bowman E,et al.Unique microRNA molecular profiles in lung cancer diagnosis and prognosis[J].Cancer Cell.2006,9(3):189-198.
[8]Ratert N,Meyer HA,Jung M,et al.miRNA profiling identifies candidate mirnas for bladder cancer diagnosis and clinical outcome[J].J Mol Diagn.2013,15(5):695-705.
[9]Corte H,Manceau G,Blons H,et al.MicroRNA and colorectal cancer[J].Dig Liver Dis.2012,44(3):195-200.
[10]胡明珠,李佩玲.非编码RNA在卵巢癌中的研究进展[J].疑难病杂志,2016,15(11):1197-1200,1205.
[11]Abraham D,Jackson N,Gundara JS,et al.MicroRNA profiling of sporadic and hereditary medullary thyroid cancer identifies predictors of nodal metastasis,prognosis,and potential therapeutic targets[J].Clin Cnacer Res,2011,17(4):4772-4781.
[12]Pennelli G,Galuppini F,Barollo S,et al.The PDCD4/miR-21 pathway in medullary thyroid carcinoma[J].Hum Pathol.2015,46(1):50-57.
[13]Hudson JH,Duncavage E,Tamburrino A,et al.Overexpression of miR-10a and miR-375 and downregulation of YAP1 in medullary thyroid carcinoma[J].Exp Mol Pathol,2013,95(1):62-67.
[14]Duan L,Hao X,Liu Z,et al.MiR-129-5p is downregulated and involved in the growth,apoptosis and migration of medullary thyroid carcinoma cells through targeting RET[J].FEBS Lett,2014,588(9):1644-1651.
[15]Su H,Yang J R,Xu T,et al.MicroRNA-101,downregulated in hepatocellular carcinoma,promotes apoptosis and suppresses tumorigenicity[J].Cancer Res,2009,69(3):1135-1142.
[16]Cho HM,Jeon HS,Lee SY,et al.microRNA-101 inhibits lung cancer invasion through the regulation of enhancer of zeste homolog 2[J].Exp Ther Med,2011,2(5):963-967.
[17]Chakravarthi BVSK,Goswami MT,Pathi SS,et al.MicroRNA-101 regulated transcriptional modulator SUB1 plays a role in prostate cancer[J].Oncogene,2016,35(49):6330-6340.
[18]Huang F,Lin C,Shi YH,et al.MicroRNA-101 inhibits cell proliferation,invasion,and promotes apoptosis by regulating cyclooxygenase-2 in Hela cervical carcinoma cells[J].Asian Pac J Cancer Prev,2013,14(10):5915-5920.
[19]Long Y,Wu Z,Yang X,et al.MicroRNA-101 inhibits the proliferation and invasion of bladder cancer cells via targeting c-FOS[J].Mol Med Rep,2016,14(3):2651-2656.
[20]Wang C,Lu S,Jiang J,et al.Hsa-microRNA-101suppresses migration and invasion by targeting Rac1 in thyroid cancer cells[J].Oncology Lett,2014,8(4):1815-1821.
[21]陈维,王昌留,王磊.SOX9基因的结构、功能及进化[J].生命的化学,2012,32(4):310-315.
[22]Zhang Y,Guo X,Xiong L,et al.MicroRNA-101suppresses SOX9-dependent tumorigenicity and promotes favorable prognosis of human hepatocellular carcinoma[J].FEBS Lett,2012,586(24):4362-4370.
[23]Liu N,Zhang L,Wang Z,et al.MicroRNA-101 inhibits proliferation,migration and invasion of human glioblastoma by targeting SOX9[J].Oncotarget,2017,8(12):19244-19254.