lincRNA-p21通过STAT3信号抑制结直肠癌HCT116细胞增殖
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摘要
目的:研究基因间区长链非编码RNA-p21(lincRNA-p21)通过STAT3信号通路对结直肠癌HCT116细胞生长抑制的影响。方法:通过细胞转染法构建lincRNA-p21过表达的人结直肠癌细胞株HCT116,转染空载体pcDNA3.1作为阴性对照组。转染后采用RT-qPCR法检测细胞中lincRNA-p21的水平,分别采用MTT法和平板集落形成实验检测细胞的活力和增殖情况,采用Western blot法测定细胞中STAT3和磷酸化STAT3(p-STAT3)的蛋白水平。用STAT3信号通路激活剂SD19处理lincRNA-p21过表达的HCT116细胞,Western blot检测STAT3和p-STAT3的蛋白水平,MTT法检测细胞活力的变化,流式细胞术检测细胞凋亡情况。结果:与control组和pcDNA组相比,pcDNA-lincRNA-p21组细胞中lincRNA-p21的表达明显上调,细胞生长受到抑制,STAT3和p-STAT3的蛋白水平降低(P<0.05)。STAT3激活剂SD19处理过表达lincRNA-p21的HCT116细胞后,与pcDNA-lincRNA-p21组相比,pcDNA-lincRNA-p21+SD19组细胞中STAT3和p-STAT3的蛋白水平升高,细胞活力升高,细胞凋亡率降低(P<0.05)。结论:lincRNA-p21过表达可以抑制结直肠癌HCT116细胞的生长;激活STAT3信号通路可促进HCT116细胞的生长;lincRNA-p21可通过抑制STAT3信号激活而抑制HCT116细胞的增殖。
AIM: To study the effect of long intergenic non-coding RNA-p21(lincRNA-p21) on the growth inhibition of colorectal cancer HCT116 cells via STAT3 signaling pathway. METHODS: The human colorectal cancer cell line HCT116 was used to construct the cells with over-expression of lincRNA-p21 by transfection of pcDNA-lincRNA-p21, and negative control cells were also set up. After transfection, the expression level of lincRNA-p21 was detected by RT-qPCR. The cell viability and proliferation were examined by MTT assay and plate colony formation assay, respectively. The protein levels of STAT3 and phosphorylated STAT3(p-STAT3) were determined by Western blot. After STAT3 signaling pathway activator SD19 was used to treat the colorectal cancer HCT116 cells with over-expression of lincRNA-p21, Western blot was used to detect the protein levels of STAT3 and p-STAT3, MTT assay was used to measure the viability of the cells, and flow cytometry analysis was used to determine the cell apoptosis. RESULTS: Compared with control group and pcDNA group, the expression of lincRNA-p21 in pcDNA-lincRNA-p21 group was significantly up-regulated, the cell proliferation was inhibited, and the protein levels of STAT3 and p-STAT3 were significantly decreased(P<0.05). After treatment with STAT3 activator SD19, the protein levels of STAT3 and p-STAT3 in pcDNA-lincRNA-p21+SD19 group were higher than those in pcDNA-lincRNA-p21 group, the cell viability was increased, and the apoptotic rate was decreased significantly(P<0.05). CONCLUSION: Over-expression of lincRNA-p21 inhibits the growth of colorectal cancer HCT116 cells. STAT3 signaling pathway activator abolishes the growth inhibitory effect of lincRNA-p21 over-expression. lincRNA-p21 inhibits the growth of colorectal cancer cells by inhibiting the activation of STAT3 signaling.
AIM: To study the effect of long intergenic non-coding RNA-p21(lincRNA-p21) on the growth inhibition of colorectal cancer HCT116 cells via STAT3 signaling pathway. METHODS: The human colorectal cancer cell line HCT116 was used to construct the cells with over-expression of lincRNA-p21 by transfection of pcDNA-lincRNA-p21, and negative control cells were also set up. After transfection, the expression level of lincRNA-p21 was detected by RT-qPCR. The cell viability and proliferation were examined by MTT assay and plate colony formation assay, respectively. The protein levels of STAT3 and phosphorylated STAT3(p-STAT3) were determined by Western blot. After STAT3 signaling pathway activator SD19 was used to treat the colorectal cancer HCT116 cells with over-expression of lincRNA-p21, Western blot was used to detect the protein levels of STAT3 and p-STAT3, MTT assay was used to measure the viability of the cells, and flow cytometry analysis was used to determine the cell apoptosis. RESULTS: Compared with control group and pcDNA group, the expression of lincRNA-p21 in pcDNA-lincRNA-p21 group was significantly up-regulated, the cell proliferation was inhibited, and the protein levels of STAT3 and p-STAT3 were significantly decreased(P<0.05). After treatment with STAT3 activator SD19, the protein levels of STAT3 and p-STAT3 in pcDNA-lincRNA-p21+SD19 group were higher than those in pcDNA-lincRNA-p21 group, the cell viability was increased, and the apoptotic rate was decreased significantly(P<0.05). CONCLUSION: Over-expression of lincRNA-p21 inhibits the growth of colorectal cancer HCT116 cells. STAT3 signaling pathway activator abolishes the growth inhibitory effect of lincRNA-p21 over-expression. lincRNA-p21 inhibits the growth of colorectal cancer cells by inhibiting the activation of STAT3 signaling.
引文
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[27] Wang G,Li Z,Zhao Q,et al.LincRNA-p21 enhances the sensitivity of radiotherapy for human colorectal cancer by targeting the Wnt/β-catenin signaling pathway[J].Oncol Rep,2014,31(4):1839-1846.
[2] Rassouli FB,Matin MM,Saeinasab M.Cancer stem cells in human digestive tract malignancies[J].Tumor Biol,2016,37(1):7-21.
[3] Pan J,Xin L,Ma YF,et al.Colonoscopy reduces colo-rectal cancer incidence and mortality in patients with non-malignant findings:a meta-analysis[J].Am J Gastroenterol,2016,111(3):355-365.
[4] Engreitz JM,Haines JE,Perez EM,et al.Local regulation of gene expression by lncRNA promoters,transcription and splicing[J].Nature,2016,539(7629):452-455.
[5] Pradeepa MM,Mckenna F,Taylor GC,et al.Psip1/p52 regulates posterior Hoxa genes through activation of lncRNA Hottip[J].PLoS Genet,2017,13(4):e1006677.
[6] Nie W,Ge HJ,Yang XQ,et al.LncRNA-UCA1 exerts oncogenic functions in non-small cell lung cancer by targeting miR-193a-3p[J].Cancer Lett,2016,371(1):99-106.
[7] Xiang JF,Yin QF,Chen T,et al.Human colorectal cancer-specific CCAT1-L lncRNA regulates long-range chromatin interactions at the MYC locus[J].Cell Res,2014,24(5):513-531.
[8] Chen Y,Luo D,Tian W,et al.Demethylation of miR-495 inhibits cell proliferation,migration and promotes apoptosis by targeting STAT-3 in breast cancer[J].Oncol Rep,2017,37(6):3581-3589.
[9] Li X,Wang D,Zhao QC,et al.Resveratrol inhibited non-small cell lung cancer through inhibiting STAT-3 signaling[J].Am J Med Sci,2016,352(5):524-530.
[10] Na S,Huang X,Li J.Upregulation of miR-129-5p affects laryngeal cancer cell proliferation,invasiveness,and migration by affecting STAT3 expression[J].Tumor Biol,2016,37(2):1789-1796.
[11] Banerjee K,Resat H.Constitutive activation of STAT3 in breast cancer cells:a review[J].Int J Cancer,2016,138(11):2570-2578.
[12] Reyes ME,Ye Y,Zhou Y,et al.Predictors of health-related quality of life and association with survival may identify colorectal cancer patients at high risk of poor prognosis[J].Qual Life Res,2017,26(2):319-330.
[13] Bonasio R,Shiekhattar R.Regulation of transcription by long noncoding RNAs[J].Annu Rev Genet,2014,48:433-455.
[14] Qu X,Dang X,Wang W,et al.Long noncoding RNAs and mRNA regulation in peripheral blood mononuclear cells of patients with chronic obstructive pulmonary disease[J].Mediators Inflamm,2018,2018:7501851.
[15] Zhou M,Sun Y,Sun Y,et al.Comprehensive analysis of lncRNA expression profiles reveals a novel lncRNA signature to discriminate nonequivalent outcomes in patients with ovarian cancer[J].Oncotarget,2016,7(22):32433-32448.
[16] Weng M,Di W,Chao Y,et al.Noncoding RNAs in the development,diagnosis,and prognosis of colorectal cancer[J].Transl Res,2017,181:108-120.
[17] Jiang X,Liu W.Long noncoding RNA highly upregulated in liver cancer activates p53-p21 pathway and promotes nasopharyngeal carcinoma cell growth[J].DNA Cell Biol,2017,36(7):596-602.
[18] 潘珠玛,黄文瀚,李达学.lincRNA-p21抑制肾癌细胞增殖作用[J].医学研究杂志,2017,46(3):149-152.
[19] Siveen KS,Sikka S,Surana R,et al.Targeting the STAT3 signaling pathway in cancer:role of synthetic and natural inhibitors[J].Biochim Biophys Acta,2014,1845(2):136-154.
[20] Ye S,Zhang D,Cheng F,et al.Wnt/β-catenin and LIF-Stat3 signaling pathways converge on Sp5 to promote mouse embryonic stem cell self-renewal[J].J Cell Sci,2016,129(2):269-276.
[21] 杨芳,赵秋,王渝,等.姜黄素抑制STAT3信号通路对胰腺癌细胞增殖的影响[J].世界华人消化杂志,2011,19(30):3149-3153.
[22] 朱建宁,吴开杰,陈玉乐,等.2’-羟基黄烷酮通过阻断AKT/STAT3信号通路抑制膀胱癌细胞增殖、侵袭及迁移[J].细胞与分子免疫学杂志,2014,30(3):237-240.
[23] Zhai H,Fesler A,Schee K,et al.Clinical significance of long intergenic noncoding RNA-p21 in colorectal cancer[J].Clin Colorectal Cancer,2013,12(4):261-266.
[24] Liu T,Zhang J,Chai Z,et al.Ginkgo biloba extract EGb 761-induced upregulation of LincRNA-p21 inhibits colo-rectal cancer metastasis by associating with EZH2[J].Oncotarget,2017,8(53):91614-91627.
[25] 沈月明,杨巍.敲低长链基因间非编码RNA-p21对乏氧肿瘤细胞放射敏感性的影响[J].中华放射医学与防护杂志,2016,36(1):19-23.
[26] Yang F,Zhang HF,Mei Y,et al.Reciprocal regulation of HIF-1α and lincRNA-p21 modulates the warburg effect[J].Mol Cell,2014,53(1):88-100.
[27] Wang G,Li Z,Zhao Q,et al.LincRNA-p21 enhances the sensitivity of radiotherapy for human colorectal cancer by targeting the Wnt/β-catenin signaling pathway[J].Oncol Rep,2014,31(4):1839-1846.