|本期目录/Table of Contents|

[1]陈剑晖,李晓帆,穆细院,等.低氧微环境下siRNA靶向沉默HIF2α基因对前列腺癌PC3细胞增殖和凋亡的影响[J].福建医科大学学报,2017,51(05):275-281.
 CHEN Jianhui,LI Xiaofan,MU Xiyuan,et al.Effects of Silencing HIF2α Gene by siRNA on Human Prostate Cancer PC3 Cell Apoptosis And Proliferation Under Hypoxic Conditions[J].Journal of Fujian Medical University,2017,51(05):275-281.
点击复制

低氧微环境下siRNA靶向沉默HIF2α基因对前列腺癌PC3细胞增殖和凋亡的影响(PDF)
分享到:

《福建医科大学学报》[ISSN:1672-4194/CN:35-1192/R]

卷:
第51卷
期数:
2017年05期
页码:
275-281
栏目:
论著
出版日期:
2017-10-30

文章信息/Info

Title:
Effects of Silencing HIF2α Gene by siRNA on Human Prostate Cancer PC3 Cell Apoptosis And Proliferation Under Hypoxic Conditions
文章编号:
16724194(2017)05027507
作者:
陈剑晖1 李晓帆2 穆细院1 李永生1 蔡伟忠1
福建医科大学 附属协和医院,福州3500011. 泌尿外科;2. 血液科,福建省血液病研究所
Author(s):
CHEN Jianhui1 LI Xiaofan2 MU Xiyuan1 LI Yongsheng1 CAI Weizhong1
1. Department of Urology, Fujian Medical University Union Hospital, Fuzhou 350001, China; 2. Department of Hematology, Fujian Institute of Hematology, Fujian Provincial Key Laboratory on Hematology, Fujian Medical University Union Hospital, Fuzhou 350001, China
关键词:
前列腺肿瘤 RNA干扰 基因 细胞增殖 细胞凋亡
Keywords:
KEY WORDS:prostatic neoplasms RNA interference genes cell proliferation apoptosis
分类号:
R341.31; R737.25; R977.6
DOI:
-
文献标志码:
A
摘要:
目的观察体外低氧条件下缺氧诱导因子2α(HIF2α)在人前列腺癌细胞PC3中的表达情况,并研究siRNA沉默HIF2α基因对前列腺癌PC3细胞生长及凋亡的影响。方法在体外培养的PC3细胞中加入化学诱导剂CoCl2建立低氧模型,采用逆转录PCR(RTPCR)和Western印迹法检测CoCl2诱导HIF2α mRNA和蛋白表达的时效和量效关系。合成HIF2α siRNA片段并转染前列腺癌PC3细胞,采用RTPCR及Western印迹法检测HIF2α siRNA对HIF2α基因表达的影响,采用MTT法检测转染HIF2α siRNA后对PC3细胞生长的抑制情况,采用流式细胞仪检测RNA干扰对PC3细胞凋亡的影响。结果(1)低氧可诱导PC3细胞的HIF2α mRNA表达,与常氧组比较,低氧12,24及48 h组的HIF2α mRNA和蛋白表达量逐渐升高(P<0.05)。100 μmol/L的CoCl2作用48 h可作为最佳前列腺癌细胞PC3低氧模型。(2)低氧+HIF2α siRNA干扰组的HIF2α mRNA和蛋白表达水平明显低于低氧+阴性对照组和低氧组(P<0.05),而低氧+阴性对照组与低氧组的HIF2α mRNA和蛋白表达水平无显著性差异(P>0.05)。低氧+HIF2α siRNA干扰组细胞增殖受抑制、细胞凋亡增加(P<0.05),而常氧组、低氧组、低氧+阴性对照组细胞增殖活力和凋亡差别均无统计学意义(P>0.05)。结论低氧可促进前列腺癌PC3细胞表达HIF2α逐渐上调,siRNA干扰HIF2α可以抑制低氧状态下前列腺癌PC3细胞HIF2α基因的表达,达到抑制细胞增殖、促进细胞凋亡的目的。阻断HIF2α信号通路可作为激素非依赖前列腺癌治疗的新策略。
Abstract:
ABSTRACT:ObjectiveTo observe the expression of hypoxiainducible factor2alpha (HIF2α) in human prostate cancer PC3 cell under hypoxia, and investigate the effects of silencing HIF2α gene by siRNA on PC3 cell apoptosis and proliferation under hypoxia.MethodsTumor hypoxia was induced by CoCl2 chemical hypoxia method.RTPCR and Westernblot was performed to detect the expression of HIF2α mRNA and protein in human prostate cancer PC3 cells, and the relations of the quantityefficiency and the timeefficiency were analyzed.siRNA was chemically synthesized and transfected into PC3 cells.The expressions of HIF2α in hypoxia microenvironment were respectively detected by RTPCR and Westernblot.The effects of apoptosis and proliferation were detected by FCM and MTT assay.Results(1)Hypoxia improved HIF2α expression in PC3 cells in a timedependent manner.Compared with the normal oxygen group, the HIF2α mRNA and protein expression level was significantly higher in 12 h, 24 h, and 48 h group(P<0.05).The best model of hypoxia was 48 h group induced by 100 μmol/L CoCl2.(2)The levels of mRNA and protein of HIF2α were suppressed significantly after being treated with HIF2α siRNA in PC3 cells(P<0.05), but there was no significant difference in cell proliferation and apoptosis between CoCl2 negative control group and CoCl2 group(P>0.05).The proliferation potential and apoptosis in PC3 cells were changed significantly after transfecting HIF2α siRNA(P>0.05), but there was no significant difference among normal oxygen group, CoCl2 groups, and CoCl2 negative control group(P>0.05).ConclusionHIF2α is overexpressed in human prostate cancer cell PC3 after CoCl2 treatment.HIF2α siRNA effectively reduces the HIF2α mRNA expressions in PC3 cells.Tansfecting HIF2α siRNA inhibits PC3 cells proliferation and increases PC3 cells apoptosis.Blocking HIF2α signal pathway can be a potential strategy in the treatment of androgen independent prostate cancer.

参考文献/References:

[1]Poon E, Harris A L, Ashcroft M. Targeting the hypoxiainducicble factor(HIF) pathway in cancer[J]. Expert Rev Mol Med, 2009,11(1):e26.
[2]Gassmann M, Chilov D, Wenger R H. Regulation of the hypoxiainducible factor1 alpha in the nucleus[J]. Adv Exp Med Biol, 2000,475(1):8799.
[3]Amankwah E K, Sellers T A, Park J Y. Gene variants in the angiogenesis pathway and prostate cancer[J]. Carcinogenesis, 2012,33(7):12591269.
[4]GonzalezFlores A, AguilarQuesada R, Siles E, et al. Intercaction between PARP1 and HIF2 alpha in the hypoxie response[J]. Oncogene, 2014,33(7):891898.
[5]Li L, Madu C O, Lu A, et al. HIF1 alpha promotes a hypoxiaindependent cell migration[J]. Open Biol J, 2010,3(1):814.
[6]Com P G, Ricci M S, Scata K A, et al. Mxi1 is induced by hypoxia in a HIF1dependent manner and protects cells from cMycinduced apoptosis[J]. Cancer Biol Ther, 2005,4(11):12851294.
[7]Loboda A, Jozkowicz A, Dulak J. HIF1 and HIF2 transcription factorssimilar but not identical[J]. Mol Cells, 2010,29(5):435442.
[8]Salminen A, Kai K, Kauppinen A. AMPK and HIF signaling pathways regulate both longevity and cancer growth: the good news and the bad news about survival mechanisms[J]. Biogerontology, 2016,17(4):655680.
[9]Semenza G L. Oxyen homeostasis[J].Wiley Interdiscip Rev Syst Biol Med, 2010,22(2):177180.
[10]Chen L, Endler A, Shibasaki F. Hypoxia and angiogenesis:regulation of hypoxiainducible factors via novel binding lactors[J]. Exp Mol Med, 2009,41(12):4249.
[11]Zhdanov A V, Dmitriev R I, Golubeva A V, et al. Chronic hypoxia leads to a glycolytic phenotype and suppressed HIF2 signaling in PC12 cells[J]. Biochim Biophys Acta, 2013,1830(6):35533569.
[12]Uchida T, Rossignol F, Matthay M A, et al. Prolonged hypoxia differentially regulates hypoxiainducible factor (HIF)2α and HIF2α expression in lung epithelial cells[J]. J Biol Chem, 2004,279(15):1487114878.
[13]Ji P, Xuan J W, Onita T, et al. Correlation study showing no concordance between EPAS1/HIF2alpha mRNA and protein expression in transitional cell cancer of the bladder[J]. Urology, 2003,61(4):851857.
[14]Martínezsez O, Gajate B P, Alonsogordoa M A, et al. Targeting HIF2α in clear cell renal cell carcinoma: A promising therapeutic strategy[J]. Crit Rev Oncol Hematol, 2017,111(5):117123.
[15]Foreman K E, Patel D, Davidson V, et al. MP4509 emetine dihydrochloride preferentially inhibits HIF1α and HIF2α expression in bladder cancer cells[J]. Journal of Urology, 2015,193(4):e538e539.
[16]Mei Y K, Lemos R, Liu X, et al. The hypoxiaassociated factor switches cells from HIF1alpha to HIF2alphadependent signaling promoting stem cell characteristics,aggressive tumor growth and invasion[J]. Cancer Res, 2011,71(11):40154027.
[17]Mimeault M, Batra S K. Hypoxiainducing factors as master regulators of stemness properties and altered metabolism of cancerand metastasisinitiating cells[J]. J Cell Mol Med, 2013,17(1):3054.
[18]Pecot C V, Calin G A, Coleman R L, et al. RNA interference in the clinic:challenges and future directions[J]. Nat Rev Cancer, 2011,11(1):5967.
[19]Chitkara D, Singh S, Mittal A. Nanocarrierbased codelivery of small molecules and siRNA/miRNA for treatment of cancer[J]. Therapeutic Delivery, 2016,7(4):245.
[20]Maduri S. Applicability of RNA interference in cancer therapy: Current status.[J]. Indian J Cancer, 2015,52(1):11.
[21]王健. 缺氧诱导因子2α对肝癌细胞生物学行为的影响[D].武汉:华中科技大学,2006.
[22]BenShoshan J, Schwartz S, Luboshits G, et al. Constitutive expression of HIF1α and HIF2α in bone marrow stromal cells differentially promotes their proangiogenic properties[J]. Stem Cells, 2008,26(10):26342643.
[23]Luo J, Lee S O, Cui Y, et al. Infiltrating bone marrow mesenchymal stem cells(BMMSCs) increase prostate cancer cell invasion via altering the CCL5/HIF2α/androgen receptor signals[J]. Oncotarget, 2015,6(29):2755527565.

相似文献/References:

[1]张晓艳,温春燕,吕玉华,等.靶向YWHAE基因shRNA慢病毒表达质粒的构建及功能验证[J].福建医科大学学报,2017,51(03):139.
 ZHANG Xiaoyan,WEN Chunyan,LYU Yuhua,et al.Construction and Function Identification of Short Hairpin RNA Lentiviral Expression Plasmid Targeting YWHAE[J].Journal of Fujian Medical University,2017,51(05):139.

备注/Memo

备注/Memo:
基金项目: 福建省卫生厅青年基金(2010217);福建省青年拔尖创新人才及福建省杰出青年科研人才培育计划(JA14130) 作者单位: 福建医科大学 附属协和医院,福州3500011. 泌尿外科;2. 血液科,福建省血液病研究所 作者简介: 陈剑晖,男,主治医师,医学硕士 通讯作者: 蔡伟忠. Email: 59506952@qq.com
更新日期/Last Update: 2017-10-30