RNAi沉默p53基因对HeLa细胞核转录因子3表达及其对细胞增殖和侵袭能力的影响

doi:10.3969/j.issn.1004-616x.2017.01.009

癌变·畸变·突变 ›› 2017, Vol. 29 ›› Issue (1): 46-50.doi: 10.3969/j.issn.1004-616x.2017.01.009

RNAi沉默p53基因对HeLa细胞核转录因子3表达及其对细胞增殖和侵袭能力的影响

李慧英¹, 李洋²

1. 广州市增城区妇幼保健院, 广东广州 511000;
2. 东莞市第五高级中学, 广东东莞 523000

收稿日期:2016-05-11 修回日期:2016-07-21 出版日期:2017-01-31 发布日期:2017-01-31
作者简介:李慧英,E-mail:lihuiying192@163.com

Effect of RNAi silenced p53 gene on expression of nuclear transcription factor 3 in cervical cancer HeLa cells

LI Huiying¹, LI Yang²

1. Maternal and Child Care Service Centre of Guangzhou Zengcheng District, Guangzhou 511000;
2. Dongguan City Fifth Senior Middle School, Dongguan 523000, Guangdong, China

Received:2016-05-11 Revised:2016-07-21 Online:2017-01-31 Published:2017-01-31

摘要/Abstract

摘要：

目的：探讨RNAi沉默p53基因对宫颈癌HeLa细胞核转录因子3（STAT3）的表达，及对HeLa细胞增殖和侵袭能力的影响。方法：构建3种靶向p53基因的shRNA1~3序列，并转染HeLa细胞，采用逆转录PCR和Western blot检测细胞p53 mRNA和蛋白的表达，以筛选有效沉默p53的shRNA序列。将筛选的序列转染HeLa细胞后，采用逆转录PCR和Western blot检测STAT3 mRNA和蛋白的表达；采用四甲基噻唑蓝法（MTT）检测细胞增殖；采用Transwell检测细胞侵袭能力。结果：构建的3种p53-shRNA均可沉默宫颈癌HeLa细胞p53的表达，尤以shRNA3的沉默效果最好，转染shRNA3后，HeLa细胞p53 mRNA和蛋白的表达量分别为0.23±0.05和0.41±0.11，显著低于未转染组、空质粒组、p53-shRNA1组和p53-shRNA2组（P均 < 0.05），故后续试验均以shRNA3转染HeLa细胞。将shRNA3序列转染HeLa细胞沉默p53基因后，p53-shRNA3组STAT3 mRNA和蛋白表达量分别为1.09±0.21和1.46±0.25，显著高于未转染组（0.53 ±0.10和0.74±0.14）及空质粒组（0.55±0.11和0.73±0.15）（P均 < 0.05）。MTT法分析结果显示，在孵育24、36、48和72 h时，p53-shRNA3组HeLa细胞增殖能力均显著大于未转染组和空质粒组（P均 < 0.05）。Transwell法分析结果显示，转染12 h后p53-shRNA3组穿膜细胞数目（59.36±5.33）显著高于未转染组（15.73±1.29）和空质粒组（16.01±1.34）（P均 < 0.05）。结论：RNAi沉默p53基因能够上调宫颈癌HeLa细胞STAT3的表达，并促进细胞的增殖及侵袭能力。

关键词: RNAi沉默, p53, HeLa细胞, 核转录因子3, 增殖, 侵袭

Abstract:

OBJECTIVE: To observe the effect of RNAi silenced p53 gene on the expression of nuclear transcription factor STAT3 in cervical cancer HeLa cells,and on their proliferation and invasion. METHODS: 3 kinds of short hairpin RNA (shRNA) which target the p53 gene were constructed and were used for liposome-transfection into HeLa cells. Then,expression of p53 were detected by PCR and Western blot. After silencing the p53 gene,expression of STAT3 was detected by PCR and Western blot. Proliferation and invasion of HeLa cells were detected by the MTT and Transwell assays. RESULTS: The transfection efficiency of p53-shRNA3 group was 85.7%. After the transfection,the expression of p53 mRNA and protein were 0.23±0.05 and 0.41±0.11,respectively,significantly lower than the non-transfection,empty plasmid,p53-shRNA1 and p53-shRNA2 groups (P < 0.05). In addition,expression of STAT3 mRNA and protein of the p53-shRNA3 group were 1.09±0.21 and 1.46±0.25,respectively,significantly higher than non-transfection group and empty plasmid group (P < 0.05). MTT results showed that at 24,36,48 h and 72 h,cell proliferation ability of the p53-shRNA3 group was significantly higher than the non-transfected group and empty plasmid group (P < 0.05). Transwell results showed that the number of cells in the p53-shRNA3 group (0.05±59.36) was significantly higher than the non-transfection group (15.73±1.29) and the empty vector group (16.01±1.34) (P < 0.05). CONCLUSION: Silencing the p53 gene can significantly increase the expression of STAT3,and promote the proliferation and invasive ability of HeLa cells.

Key words: RNAi silencing, p53, HeLa cell, signal transducer and activator of transcription 3, proliferation, invasion

中图分类号:

R730.231

李慧英, 李洋. RNAi沉默p53基因对HeLa细胞核转录因子3表达及其对细胞增殖和侵袭能力的影响[J]. 癌变·畸变·突变, 2017, 29(1): 46-50.

LI Huiying, LI Yang. Effect of RNAi silenced p53 gene on expression of nuclear transcription factor 3 in cervical cancer HeLa cells[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2017, 29(1): 46-50.

参考文献

[1] WEN Y,FENG D,WU H,et al.Defective initiation of liver regeneration in osteopontin-deficient mice after partial hepatec-tomy due to insufficient activation of IL-6/Stat3 pathway[J].Int J Biol Sci,2015,11(10):1236-1247.

[2] JOHNSTON P A,SEN M,HUA Y,et al.HCS campaign to identify selective inhibitors of IL-6-induced STAT3 pathway activation in head and neck cancer cell lines[J].Assay Drug Dev Technol,2015,13(7):356-376.

[3] JUNG I H,CHOI J H,CHUNG Y Y,et al.Predominant activation of JAK/STAT3 pathway by interleukin-6 is implicated in hepatocarcinogenesis[J].Neoplasia,2015,17(7):586-597.

[4] RODRIGUEZ-BARRUECO R,YU J,SAUCEDO-CUEVAS L P,et al. Inhibition of the autocrine IL-6-JAK2-STAT3-calprotectin axis as targeted therapy for HR-/HER2⁺ breast cancers[J].Genes Dev,2015,29(15):1631-1648.

[5] DARVIN P,BAEG S J,JOUNG Y H,et al.Tannic acid inhibits the JAK2/STAT3 pathway and induces G1/S arrest and mitochondrial apoptosis in YD-38 gingival cancer cells[J].Int J Oncol, 2015,47(3):1111-1120.

[6] YU C,HUO X,AGOSTON A T,et al.Mitochondrial STAT3 contributes to transformation of Barrett's epithelial cells that express oncogenic Ras in a p53-independent fashion[J].Am J Physiol Gastrointest Liver Physiol,2015,309(3):G146-161.

[7] GUILIAN N,WRIGHT K L,YIHONG M,et al.Role of Stat3 in regulating p53 expression and function[J].Mol Cell Biol, 2005,25(17):7432-7440.

[8] DAVID D,RAJAPPAN L M,BALACHANDRAN K,et al. Prognostic significance of STAT3 and phosphorylated STAT3 in human soft tissue tumors-a clinicopathological analysis[J].J Exp Clin Cancer Res,2011,30:56.

[9] CHEN J,LAN T,ZHANG W,et al. Feed-forward reciprocal activation of PAFR and STAT3 regulates epithelial-mesenchymal transition in non-small cell lung cancer[J]. Cancer Res, 2015,75(19):4198-4210.

[10] WU K,WANG B,CHEN Y,et al. DAB2IP regulates the chemoresistance to pirarubicin and tumor recurrence of non-muscle invasive bladder cancer through STAT3/Twist1/P-glycoprotein signaling[J].Cell Signal,2015,27(12):2515-2523.

[11] MATTEO-SANTONI M D,MASSARI F,DEL-RE M,et al. Investigational therapies targeting signal transducer and activator of transcription 3 for the treatment of cancer[J].Expert Opin Invest Drugs,2015,24(6):809-824.

[12] KIM S H,AHN K S,JEONG S J,et al.Janus activated kinase 2/signal transducer and activator of transcription 3 pathway mediates icariside Ⅱ-induced apoptosis in U266 multiple myeloma cells[J].Eur J Pharmacol,2011,654(1):10-16.

[1]	赵岩, 孙震晓. 二甲双胍抑制人MDA-MB-231细胞增殖及迁移的体外实验研究[J]. 癌变·畸变·突变, 2022, 34(1): 35-39.
[2]	王晓舟, 李洪石, 于溪, 魏宁, 王梓瑛, 赫丽杰. 青藤碱对HPV阳性宫颈癌SiHa细胞增殖和侵袭的影响[J]. 癌变·畸变·突变, 2021, 33(6): 442-445.
[3]	杨兴肖, 张雪原, 邹乃祎, 单保恩, 马鸣, 祝淑钗. 干扰HMGB1基因表达对食管鳞癌细胞侵袭迁移及放射敏感性的影响[J]. 癌变·畸变·突变, 2021, 33(5): 327-333.
[4]	王琛瑶, 梁百慧, 吴锦源, 唐梓涵, 许小文, 尹梓健, 丁雅琼, 方雄钊, 李栢芝, 文姝佚, 程转, 詹涵, 张粤烽, 王泫哲, 何仕龙, 张可洵, 刘雅骐, 周飞, 熊兴东, 白春英, 许丽艳, 李恩民. RAC1对恶性肿瘤的调控作用及其分子机制研究进展[J]. 癌变·畸变·突变, 2021, 33(5): 401-404.
[5]	黄晓华, 牛永东, 石刚刚. 核受体FXR激活对宫颈癌CaSki细胞增殖的抑制效应及其作用机制[J]. 癌变·畸变·突变, 2021, 33(4): 302-306.
[6]	郑珺文, 常晨, 郝佳洁, 蔡岩, 王明荣, 张钰. NEDD4在食管鳞癌中的表达及作用研究[J]. 癌变·畸变·突变, 2021, 33(3): 203-207,224.
[7]	张娜, 范志露, 杨荔艳, 常晨, 蔡岩, 郝佳洁, 王明荣. 食管鳞癌中SERPINB5表达降低的作用机制及其临床意义[J]. 癌变·畸变·突变, 2021, 33(2): 101-109.
[8]	方磊, 杨涛, 盛磊, 木塔力甫·艾买提, 齐鑫鑫, 艾尼娃尔·艾克木, 伊力亚尔·尼加提. 黑种草子总皂苷对人宫颈癌SiHa细胞生物学行为的影响[J]. 癌变·畸变·突变, 2021, 33(2): 143-148,152.
[9]	刘春燕, 张爱华. p53的翻译后修饰及其与砷中毒致人体多器官损伤关系的研究进展[J]. 癌变·畸变·突变, 2021, 33(2): 157-160.
[10]	王耀坤, 陈珏晓, 张明远, 邵长利, 杨玉, 商宇. 基于数据库的未分化甲状腺癌基因组变异分析及靶向药物筛选[J]. 癌变·畸变·突变, 2021, 33(1): 53-57.
[11]	杜璐, 姜晓燕, 丁库克. 稳定敲降NF-YB基因HeLa细胞系的构建[J]. 癌变·畸变·突变, 2020, 32(6): 409-413.
[12]	兰莉, 陈海丽, 徐蕾. RBMS3-AS3对宫颈癌细胞增殖、凋亡和侵袭的影响及作用机制[J]. 癌变·畸变·突变, 2020, 32(6): 438-443.
[13]	林凯煌, 蔡高阳, 刘新城. mTOR抑制剂AZD2014对肝癌细胞的增殖抑制作用及其机制研究[J]. 癌变·畸变·突变, 2020, 32(6): 448-451.
[14]	刘鹏, 白意晓. miR-29a对人胃癌细胞增殖、迁移的影响及其机制研究[J]. 癌变·畸变·突变, 2020, 32(6): 452-456,463.
[15]	许迎剑, 张航, 张建清. 细胞核受体RXRα在2,2',4,4'-四溴二苯醚神经毒性中的作用及其机制[J]. 癌变·畸变·突变, 2020, 32(5): 336-343,349.

RNAi沉默p53基因对HeLa细胞核转录因子3表达及其对细胞增殖和侵袭能力的影响

Effect of RNAi silenced p53 gene on expression of nuclear transcription factor 3 in cervical cancer HeLa cells

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价