沉默FOXQ1基因对甲状腺乳头状癌TPC-1细胞增殖的影响
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摘要
目的 FOXQ1基因在甲状腺乳头状癌(PTC)组织中呈现高表达,有关FOXQ1在PTC的生物学功能尚不清楚。文中旨在探讨FOXQ1-siRAN对PTC的TPC-1细胞增殖的影响及其可能的机制。方法通过lipofectamineTM2000将3条人工合成的FOXQ1-siRNA和NC-siRNA分别转入到PTC的TPC-1细胞,实验分为空白对照组(加入等量的磷酸盐缓冲液PBS)、NC-siRAN组(转染无意义序列siRNA)、FOXQ1-1组(空白培养基+lipofectamineTM2000+FOXQ1-1)、FOXQ1-2组(空白培养基+lipofectamineTM2000+FOXQ1-2)、FOXQ1-3组(空白培养基+lipofectamineTM2000+FOXQ1-3)。利用qRT-PCR和Western blot法分别检测FOXQ1 mRNA和蛋白的表达水平,MTT法观察细胞增殖活性的变化,qRT-PCR和Western blot分别检测细胞中细胞周期蛋白(cyclin D1)和c-Myc的表达水平。结果与空白对照组、NC-siRNA组FOXQ1 mRNA相对表达量比较,FOXQ1-3组明显抑制FOXQ1mRNA的表达(P<0.05)。FOXQ1-1组、FOXQ1-2组、FOXQ-3组的FOXQ1蛋白相对表达量(0.54±0.07、0.40±0.07、0.26±0.06)较NC-siRNA组(0.78±0.08)明显降低,FOXQ1-3组下降最为明显(P<0.05)。MTT结果显示,与空白对照组、NC-siRAN组比较,FOXQ1-3组可明显抑制TPC-1细胞的增殖活性(P<0.05)。FOXQ1-3组c-Myc、cyclin D1蛋白表达量(0.57±0.04、0.51±0.10)较空白对照组(1.05±0.07、0.94±0.12)、NC-siRNA组(0.92±0.06、0.91±0.11)明显降低(P<0.05)。结论沉默FOXQ1基因可以有效地抑制PTC的TPC-1细胞的增殖活性,这一作用可能通过抑制cyclin D1和c-Myc的表达水平来实现的。
Objective Forkhead box Q1( FOXQ1) is highly expressed but its biological role remains unclear in papillary thyroid carcinoma( PTC). This article aims to investigate the effect of FOXQ1-siRAN on the proliferation of TPC-1 cells and its possible mechanism. Methods Synthetic FOXQ1-siRNA and NC-siRNA were transfected into TPC-1 cells by lipofectamineTM2000. The cells were divided into five groups: NC-siRAN,FOXQ1-1,FOXQ1-2,FOXQ1-3 and blank control. After transfection,the expressions of the FOXQ1 mRNA and protein,as well as those of c-Myc and cyclin D1 proteins,were determined by qRT-PCR and Western blot,and the changes in the proliferation of the TPC-1 cells observed by MTT. Results Compared with the blank control and NC-siRAN groups, the FOXQ1-3 group showed a significantly inhibited expression of FOXQ1 mRNA( P < 0. 05). The protein expression of FOXQ1 was markedly decreased in the FOXQ1-1,FOXQ1-2 and FOXQ1-3 groups in comparison with that in the NC-siRAN group( 0.54±0.07,0.40±0.07 and 0.26±0.06 vs 0.78±0.08,P<0.05). The proliferation of the TPC-1 cells was remarkably lower in the FOXQ1-3 than in the blank control and NC-siRNA groups( P<0.05),and so were the relative protein expressions of c-Myc( 0.57± 0.04 vs 1.05 ±0.07 and 0.92±0.06,P < 0. 05) and cyclin D1( 0. 51 ± 0. 10 vs 0. 94 ± 0. 12 and 0. 91 ± 0. 11,P < 0. 05). Conclusion Silencing the FOXQ1 gene can effectively inhibit the proliferation of TPC-1 cells in TPC,probably by suppressing the expressions of c-Myc and cyclin D1.
Objective Forkhead box Q1( FOXQ1) is highly expressed but its biological role remains unclear in papillary thyroid carcinoma( PTC). This article aims to investigate the effect of FOXQ1-siRAN on the proliferation of TPC-1 cells and its possible mechanism. Methods Synthetic FOXQ1-siRNA and NC-siRNA were transfected into TPC-1 cells by lipofectamineTM2000. The cells were divided into five groups: NC-siRAN,FOXQ1-1,FOXQ1-2,FOXQ1-3 and blank control. After transfection,the expressions of the FOXQ1 mRNA and protein,as well as those of c-Myc and cyclin D1 proteins,were determined by qRT-PCR and Western blot,and the changes in the proliferation of the TPC-1 cells observed by MTT. Results Compared with the blank control and NC-siRAN groups, the FOXQ1-3 group showed a significantly inhibited expression of FOXQ1 mRNA( P < 0. 05). The protein expression of FOXQ1 was markedly decreased in the FOXQ1-1,FOXQ1-2 and FOXQ1-3 groups in comparison with that in the NC-siRAN group( 0.54±0.07,0.40±0.07 and 0.26±0.06 vs 0.78±0.08,P<0.05). The proliferation of the TPC-1 cells was remarkably lower in the FOXQ1-3 than in the blank control and NC-siRNA groups( P<0.05),and so were the relative protein expressions of c-Myc( 0.57± 0.04 vs 1.05 ±0.07 and 0.92±0.06,P < 0. 05) and cyclin D1( 0. 51 ± 0. 10 vs 0. 94 ± 0. 12 and 0. 91 ± 0. 11,P < 0. 05). Conclusion Silencing the FOXQ1 gene can effectively inhibit the proliferation of TPC-1 cells in TPC,probably by suppressing the expressions of c-Myc and cyclin D1.
引文
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[15]Song HM,Song JL,Li DF,et al.Inhibition of FOXO1 by small interfering RNA enhances proliferation and inhibits apoptosis of papillary thyroid carcinoma cells via Akt/FOXO1/Bim pathway[J].Onco Targets Ther,2015,8:3565-3573.
[16]Liang S,Mu K,Wang Y,et al.CyclinD1,a prominent prognostic marker for endometrial diseases[J].Diagn Pathol,2013,8:138.
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[18]Larue L,Delmas V.The WNT/Beta-catenin pathway in melanoma[J].Front Biosci,2006,11:733-742.
[19]魏广民,周文波,武伦,等.CyclinD1和Bax在甲状腺微小乳头状癌中的表达及意义[J].重庆医学,2017,46(3):325-328.
[20]Uezono S,Goto T,Terui K,et al.Emergence agitation after sevoflurane versus propofol in pediatric patients[J].Anesth Analg,2000,91(3):563-566.
[21]Díaz VM,de Herreros AG.F-box proteins:Keeping the epithelial-to-mesenchymal transition(EMT)in check[J].Semin Cancer Biol,2016,36:71-79.
[22]Bao B,Azmi AS,Aboukameel A,et al.Pancreatic cancer stemlike cells display aggressive behavior mediated via activation of Fox Q1[J].J Biol Chem,2014,289(21):14520-14533.
[23]李建水,邓大炜,曾丽娟.FOXQ1促进肝癌细胞系SMMC-7721细胞的增殖[J].中国生物化学与分子生物学报,2016,32(4):446-451.
[24]Zhang J,Liu Y,Zhang J,et al.FOXQ1 promotes gastric cancer metastasis through upregulation of Snail[J].Oncol Rep,2016,35(6):3607-3613.
[25]朱双伟,李相述,彭旭东,等.FOXQ1基因介导了Shh诱导的SW480细胞血管生成及增殖[J].中国病理生理杂志,2016,32(3):470-476.
[2]马晔,刘柳,沈美萍.甲状腺乳头状癌中促甲状腺激素受体与钠碘转运体mRNA的表达[J].医学研究生学报,2012,25(7):699-701.
[3]Riesco-Eizaguirre G,Santisteban P.New insights in thyroid follicular cell biology and its impact in thyroid cancer therapy[J].Endocr Relat Cancer,2007,14(4):957-977.
[4]刘泽兵,王丽,叶宣光,等.老年前期及老年期甲状腺乳头状癌C-MET和PP60C-SRC蛋白的表达[J].中国老年学杂志,2011,31(1):9-11.
[5]Kaestner KH,Knochel W,Martinez DE.Unified nomenclature for the winged helix/forkhead transcription factors[J].Genes Dev,2000,14(2):142-146.
[6]Li Y,Zhang Y,Yao Z,et al.Forkhead box Q1:A key player in the pathogenesis of tumors(Review)[J].Int J Oncol,2016,49(1):51-58.
[7]Zhang H,Meng F,Liu G,et al.Forkhead transcription factor foxq1 promotes epithelial-mesenchymal transition and breast cancer metastasis[J].Cancer Res,2011,71(4):1292-1301.
[8]张锦锦,唐慧.叉头框Q1基因调控肿瘤发生和发展的研究进展[J].肿瘤,2017,37(2):177-183.
[9]Katoh M,Katoh M.Human FOX gene family(Review)[J].Int JOncol,2004,25(5):1495-1500.
[10]Hannenhalli S,Kaestner KH.The evolution of Fox genes and their role in development and disease[J].Nat Rev Genet,2009,10(4):233-240.
[11]Myatt SS,Lam EW.The emerging roles of forkhead box(Fox)proteins in cancer[J].Nat Rev Cancer,2007,7(11):847-859.
[12]伍慧慧,沈沁彦,李益波,等.FOXQ1在甲状腺乳头状癌中的表达及意义[J].中国细胞生物学学报,2017,39(4):419-425.
[13]Reynolds A,Leake D,Boese Q,et al.Rational siRNA design for RNA interference[J].Nat Biotechnol,2004,22(3):326-330.
[14]程文,高建平,张征宇.微小RNA与肿瘤相关性研究[J].医学研究生学报,2011,24(2):203-207.
[15]Song HM,Song JL,Li DF,et al.Inhibition of FOXO1 by small interfering RNA enhances proliferation and inhibits apoptosis of papillary thyroid carcinoma cells via Akt/FOXO1/Bim pathway[J].Onco Targets Ther,2015,8:3565-3573.
[16]Liang S,Mu K,Wang Y,et al.CyclinD1,a prominent prognostic marker for endometrial diseases[J].Diagn Pathol,2013,8:138.
[17]Moghaddam SJ,Haghighi EN,Samiee S,et al.Immunohistochemical analysis of p53,cyclinD1,RB1,c-fos and N-ras gene expression in hepatocellular carcinoma in Iran[J].World J Gastroenterol,2007,13(4):588-593.
[18]Larue L,Delmas V.The WNT/Beta-catenin pathway in melanoma[J].Front Biosci,2006,11:733-742.
[19]魏广民,周文波,武伦,等.CyclinD1和Bax在甲状腺微小乳头状癌中的表达及意义[J].重庆医学,2017,46(3):325-328.
[20]Uezono S,Goto T,Terui K,et al.Emergence agitation after sevoflurane versus propofol in pediatric patients[J].Anesth Analg,2000,91(3):563-566.
[21]Díaz VM,de Herreros AG.F-box proteins:Keeping the epithelial-to-mesenchymal transition(EMT)in check[J].Semin Cancer Biol,2016,36:71-79.
[22]Bao B,Azmi AS,Aboukameel A,et al.Pancreatic cancer stemlike cells display aggressive behavior mediated via activation of Fox Q1[J].J Biol Chem,2014,289(21):14520-14533.
[23]李建水,邓大炜,曾丽娟.FOXQ1促进肝癌细胞系SMMC-7721细胞的增殖[J].中国生物化学与分子生物学报,2016,32(4):446-451.
[24]Zhang J,Liu Y,Zhang J,et al.FOXQ1 promotes gastric cancer metastasis through upregulation of Snail[J].Oncol Rep,2016,35(6):3607-3613.
[25]朱双伟,李相述,彭旭东,等.FOXQ1基因介导了Shh诱导的SW480细胞血管生成及增殖[J].中国病理生理杂志,2016,32(3):470-476.
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