miR-18b靶向CDKN2B促进大肠癌细胞HCT116增殖和迁移研究
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摘要
目的研究miR-18b靶向CDKN2B对大肠癌细胞HCT116增殖和迁移的作用及miR-18b与CDKN2B相关性。方法大肠癌细胞经过细胞培养、转染分为大肠癌细胞组、miR-18b沉默组、miR-18b过表达组。检测大肠癌细胞增殖、细胞周期、细胞迁移,荧光定量RT-PCR检测miR-18b、CDKN2B mRNA表达,Western Blot检测CDKN2B表达。结果 miR-18b过表达组在24 h、48 h和72 h时的大肠癌细胞增殖显著较高,48 h时大肠癌细胞增殖效果最明显。miR-18b过表达组G0/G1期、S期细胞百分比较高且细胞迁移数量较高。miR-18b过表达组miR-18b表达水平高于大肠癌细胞组、miR-18b沉默组,CDKN2B mRNA低于大肠癌细胞组、miR-18b沉默组(P <0. 05)。miR-18b与CDKN2B mRNA呈负相关(r=-0. 708,P=0. 001)。结论 miR-18b过表达能增强CDKN2B表达水平,miR-18b表达上调能够促进大肠癌细胞HCT116增殖和迁移,miR-18b沉默则相反。
Objective To study the effects of microRNA-18 b targeting CDKN2 B on the proliferation and migration of colorectal cancer cell line HCT116 and the correlation between microRNA-18 b and CDKN2 B. Methods Colorectal cancer cells were cultured and transfected into colorectal cancer cell group, miR-18 b silencing group and miR-18 b overexpression group. The proliferation, cell cycle and cell migration of colorectal cancer cells were detected. The expression of microRNA-18 b and CDKN2 B was detected by fluorescence quantitative RT-OCR, and the expression of CDKN2 B was detected by Western Blot. Results The proliferation of colorectal cancer cells in the miR-18 b overexpression group was significantly higher at 24 h, 48 h and 72 h, and the proliferation effect was the most obvious at 48 h. In the miR-18 b overexpression group, the percentage of cells in G0/G1 phase and S phase was higher and the number of cell migration was higher. The expression level of miR-18 b in the overexpression group was higher than that in the colorectal cancer cell group and the miR-18 b silencing group, and the mRNA level of CDKN2 B was lower than that in the colorectal cancer cell group and the miR-18 b silencing group( P < 0. 05). The expression of microRNA-18 b was negatively correlated with CDKN2 B( r =-0. 708, P = 0. 001). Conclusion Overexpression of microRNA-18 b can enhance the expression level of CDKN2 B.Upregulation of microRNA-18 b can promote the proliferation and migration of colorectal cancer cell line HCT116, while silencing of microRNA-18 b is the opposite.
Objective To study the effects of microRNA-18 b targeting CDKN2 B on the proliferation and migration of colorectal cancer cell line HCT116 and the correlation between microRNA-18 b and CDKN2 B. Methods Colorectal cancer cells were cultured and transfected into colorectal cancer cell group, miR-18 b silencing group and miR-18 b overexpression group. The proliferation, cell cycle and cell migration of colorectal cancer cells were detected. The expression of microRNA-18 b and CDKN2 B was detected by fluorescence quantitative RT-OCR, and the expression of CDKN2 B was detected by Western Blot. Results The proliferation of colorectal cancer cells in the miR-18 b overexpression group was significantly higher at 24 h, 48 h and 72 h, and the proliferation effect was the most obvious at 48 h. In the miR-18 b overexpression group, the percentage of cells in G0/G1 phase and S phase was higher and the number of cell migration was higher. The expression level of miR-18 b in the overexpression group was higher than that in the colorectal cancer cell group and the miR-18 b silencing group, and the mRNA level of CDKN2 B was lower than that in the colorectal cancer cell group and the miR-18 b silencing group( P < 0. 05). The expression of microRNA-18 b was negatively correlated with CDKN2 B( r =-0. 708, P = 0. 001). Conclusion Overexpression of microRNA-18 b can enhance the expression level of CDKN2 B.Upregulation of microRNA-18 b can promote the proliferation and migration of colorectal cancer cell line HCT116, while silencing of microRNA-18 b is the opposite.
引文
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[18] Yu JH,Zhu BM,Wickre M,et al. The transcription factors signal transducer and activator of transcription 5A(STAT5A)and STAT5B negatively regulate cell proliferation through the activation of cyclin-dependent kinase inhibitor 2b(Cdkn2b)and Cdkn1a expression[J]. Hepatology,2010,52(5):1808-18.
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[20] Jang W,Park J,Kwon A,et al. CDKN2B downregulation and other genetic characteristics in T-acute lymphoblastic leukemia[J]. Exp Mol Med,2019,51(1):4.
[2]郭凤军,邹颖刚,李丹,等.微小RNA-92与恶性肿瘤关系的研究进展[J].中国实验诊断学,2018,22(7):1288-1290.
[3]潘成,屈潇,严雪冰,等.大肠癌相关癌前病变患者血浆微小RNA的表达特征[J].中华实验外科杂志,2018,35(12):2342.
[4]陈竟男,王杨,金东辉,等. miR-18b增强乳腺癌MDA-MB-231细胞对顺铂化疗敏感性的作用及其机制[J].吉林大学学报(医学版),2015,41(2):316-320.
[5]贺蓉,邹德学,余振东,等.鼻咽癌组织中细胞周期相关基因CDKN2B/p15的表达及临床意义[J].中国实验诊断学,2009,13(5):618-622.
[6]齐晓艳.长链非编码RNA在大肠癌中的研究进展[J].肿瘤研究与临床,2018,30(9):638-641.
[7]郁雨婷,赵根明,郁建国,等.上海市松江区结直肠癌发病危险因素研究[J].中国全科医学,2018,21(30):3709-3713.
[8]孙颖昊,吴东.微小RNA与大肠癌[J].中华内科杂志,2014,53(6):493-495.
[9]李一鸣,赵娇,刘娟,等. MiR-18b-5p对大肠癌PTEN/PI3K/Akt2信号通路的调控[J].现代肿瘤医学,2015,23(15):2092-2096.
[10] Li Y,Chen M,Liu J,et al. Upregulation of MicroRNA 18b Contributes to the Development of Colorectal Cancer by Inhibiting CDKN2B[J]. Mol Cell Biol,2017,37(22):e00391-17.
[11]周杰,郑远明,沈宗坤,等. MiR-181a和miR-181b靶向调控FUT1对结直肠癌进展的影响[J].中华检验医学杂志,2018,41(11):841-846.
[12] Azizian A,Kramer F,Jo P,et al. Preoperative Prediction of Lymph Node Status by Circulating Mir-18b and Mir-20a During Chemoradiotherapy in Patients with Rectal Cancer[J]. World J Surg,2015,39(9):2329-35.
[13]王高峰,李月灵,彭红,等.长链非编码CDKN2B调控miR-19对慢性髓细胞白血病的影响[J].中国免疫学杂志,2017,33(9):1375-1380.
[14]乔琳,文星艳,窦克非,等. CDKN2B-AS1基因多态性与女性早发冠心病的相关性研究[J].中国循环杂志,2017,32(12):1154-1157.
[15] Du Y,Hao X,Liu X. Low expression of long noncoding RNA CDKN2B-AS1 in patients with idiopathic pulmonary fibrosis predicts lung cancer by regulating the p53-signaling pathway[J]. Oncol Lett,2018,15(4):4912-4918.
[16]马天有,归巧娣,施瑞洁,等. RNA干扰抑制HPV16E6基因的表达对HPV16阳性宫颈癌细胞中基因表达谱的影响[J].现代肿瘤医学,2017,25(19):3058-3063.
[17]刘杰,王淑贞,王巧莲,等.妊娠糖尿病患者血糖、血脂水平与CDKN2A/2B基因多态性的相关性[J].标记免疫分析与临床,2018,25(9):1360-1363.
[18] Yu JH,Zhu BM,Wickre M,et al. The transcription factors signal transducer and activator of transcription 5A(STAT5A)and STAT5B negatively regulate cell proliferation through the activation of cyclin-dependent kinase inhibitor 2b(Cdkn2b)and Cdkn1a expression[J]. Hepatology,2010,52(5):1808-18.
[19] Hu Q,Chen X,Liu S,et al. Methylation of CDKN2B CpG islands is associated with upregulated telomerase activity in children with acute lymphoblastic leukemia[J]. Oncol Lett,2017,13(4):2115-2120.
[20] Jang W,Park J,Kwon A,et al. CDKN2B downregulation and other genetic characteristics in T-acute lymphoblastic leukemia[J]. Exp Mol Med,2019,51(1):4.