miR-203抑制食管鳞癌细胞迁移、侵袭及其分子机制探讨

目的:探究miR-203对食管鳞癌细胞(TR146、EC109)迁移、侵袭能力的影响及其分子机制。方法:检测miR-203在食管鳞癌细胞系中的表达水平,并通过转染miR-203激动剂agomir使TR146、EC109细胞稳定高表达miR-203,miR-203 agomir阴性对照组(NC)和无处理组(Blank)作为对照。通过划痕实验、Transwell实验检测miR-203对TR146、EC109细胞迁移、侵袭能力的影响。利用生物信息学分析miR-203潜在的靶基因,并通过双荧光素酶报告基因实验、实时定量PCR（qPCR）实验、Western blot实验验证miR-203靶基因。通过拯救实验探究miR-203是否通过抑制靶基因发挥作用。结果:与正常食管上皮细胞相比,miR-203在食管鳞癌细胞系中表达下调。在TR146、EC109细胞内将miR-203表达水平上调数倍,划痕实验证实miR-203能够抑制TR146、EC109细胞迁移能力,Transwell实验证实miR-203能够抑制TR146、EC109细胞侵袭能力。生物信息学、qPCR实验和Western blot实验表明LASP1(LIM and SH3 domain protein 1)是miR-203潜在的靶基因。拯救实验表明miR-203通过靶向抑制LASP1发挥抑制食管鳞癌细胞迁移、侵袭的作用。结论:miR-203能够抑制食管鳞癌细胞迁移、侵袭,并且该抑制作用可能通过miR-203靶向抑制LASP1介导,为食管鳞癌临床诊断和靶向治疗提供了理论依据。     

miR-203a-3p通过靶向PRMT5抑制胃癌细胞增殖

目的:探讨miR-203a-3p在胃癌中的表达及其对胃癌细胞增殖的影响.方法:收集胃癌组织标本44例,采用Real-time PCR方法检测胃癌组织标本中miR-203a-3p的表达,并分析其表达水平与临床病理参数的关系;生物信息学预测miR-203a-3p的靶基因,并采用荧光素酶报告基因实验进行验证;免疫组化检测胃癌组织标本中PRMT5的表达情况,分析其表达水平与miR-203a-3p表达的相关性;利用脂质体介导的瞬时转染方法过表达miR-203a-3p或同时过表达miR-203a-3p和PRMT5,并通过CCK-8实验检测胃癌细胞的增殖情况.结果:胃癌组织中miR-203a-3p的表达水平与正常组织对照相比显著降低(P<0.01),并且miR-203a-3p的表达水平与肿瘤细胞的分化程度显著相关;荧光素酶报告基因实验证实miR-203a-3p可以直接结合在PRMT5 3'-UTR上,即PRMT5是miR-203a-3p的直接靶基因;在胃癌组织标本中,PRMT5表达水平显著高于正常组织对照(P<0.01),并且其表达水平与miR-203a-3p表达呈显著负相关(r=-0.4124,P<0.01);过表达miR-203a-3p后,胃癌BGC823细胞的增殖能力显著低于miR-NC对照组(P<0.01),并且,"挽救"实验表明在过表达miR-203a-3p的细胞中同时过表达PRMT5后会部分恢复miR-203a-3p对细胞增殖的抑制(P<0.01).结论:miR-203a-3p可通过下调靶基因PRMT5的表达,进而抑制胃癌细胞增殖.因此,miR-203a-3p可作为胃癌疾病临床治疗的潜在靶点.     

MicroRNA 203对胃癌中EIF5A2表达的影响

目的:研究microRNA 203(miR-203)对胃癌EIF5A2表达的影响,为进一步阐明miR-203与胃癌的关系提供理论基础.方法:分别应用免疫荧光化学和免疫组织化学检测胃癌细胞和组织中EIF5A2的表达;脂质体miR-203转染干预胃癌细胞后,Real time PCR和Western blot检测细胞中EIF5A2 mRNA和蛋白的表达情况.结果:胃癌细胞株SGC7901胞浆中大量表达EIF5A2蛋白,而细胞核中表达较少;胃癌组织中EIF5A2的表达量明显升高;EIF5A2的表达在脂质体miR-203转染干预胃癌细胞后明显受到抑制.结论:胃癌细胞和组织中存在EIF5A2的表达,且miR-203显著抑制了EIF5A2的表达.     

MicroRNA-7与肺癌关系的研究进展

MicroRNA-7(miR-7)首先在果蝇(Drosophila melanogaster)体内被发现,参与果蝇翼、卵子等形成. 在人类体内miR-7不但参与细胞增殖、分化等,还参与肿瘤的发生发展,尤其在肺癌中起重要作用. 多数研究报道肺癌组织和肺癌细胞株中miR-7呈低表达,低表达组患者预后差,提高miR-7的表达能抑制肿瘤生长. miR-7作为肿瘤抑制因子,负性调控BCL-2、EGFR等的表达,调节肿瘤细胞对放化疗和靶向治疗的敏感性,有望成为肺癌治疗的新靶点.     

miR-203抑制bcl-w表达促进膀胱癌细胞凋亡

摘 要：［目的］ 研究miR-203在膀胱癌组织中的表达,探讨其表达对膀胱癌细胞T24凋亡的影响。［方法］ 实时荧光定量PCR(qRT-PCR)检测23例膀胱癌组织和癌旁正常组织中miR-203表达情况,免疫印迹法检测bcl-w蛋白的表达情况,Pearson法分析miR-203表达与bcl-w蛋白表达相关性;转入miR-203-mimic建立高表达miR-203的T24细胞,流式细胞仪检测miR-203表达对T24细胞凋亡的影响。［结果］ miR-203在膀胱癌组织中的相对表达量为1.40±0.47,显著低于其在癌旁正常组织中的表达量（2.99±0.59）（P=0.0001）;miR-203在膀胱癌组织中的表达与bcl-w蛋白表达呈负相关（r=-0.952,P<0.001）;miR-203靶向抑制T24细胞中bcl-w蛋白的表达,转入miR-203-mimic 24h和72h后,T24细胞凋亡率分别为8.33%±0.58%和17.07%±0.76%,均显著高于对照组T24细胞的3.63%±0.32%和4.16%±0.28%（P=0.012,P<0.001）。［结论］ miR-203在膀胱癌组织中低表达,其可以通过靶向抑制bcl-w的表达促进膀胱癌细胞T24的凋亡。     

miR-203-3p在肝癌细胞中的表达及其对细胞增殖、迁移和侵袭的影响

目的:探讨微小RNA-203-3p（miR-203-3p）在肝细胞癌细胞中的表达,并进一步分析其对肝细胞癌进展的影响。方法:通过qRT-PCR检测miR-203-3p在不同肝细胞癌细胞系中的表达,并利用敲低和过表达的方法分析miR-203-3p表达水平的变化对肝癌细胞生长、运动等的影响,并初步探索其机制。结果:肝细胞癌细胞中的miR-203-3p表达水平低于正常肝细胞。功能上,miR-203-3p抑制肝细胞癌细胞增殖、迁移、侵袭,加速细胞凋亡;机制上,miR-203-3p可与FGF2 mRNA碱基配对,吸附FGF2 mRNA导致FGF2蛋白生成减少。结论:miR-203-3p通过直接靶向作用,抑制FGF2表达,从而抑制肝细胞癌细胞的增殖、迁移和侵袭。     

miR-203在结直肠癌中的表达及其功能分析

目的 探讨microRNA-203（miR 203）在结直肠癌中的表达情况及其临床意义,并在结直肠癌细胞中验证其功能。方法 采用实时荧光定量PCR（qRT-PCR）检测52例结直肠癌及对应癌旁组织中miR-203的水平,分析miR-203表达与临床病理参数及术前癌胚抗原的关系;检测miR 203在3种结直肠癌细胞株SW480、Lovo和HT 29及人正常结肠黏膜上皮细胞NCM460中的表达情况,选取miR-203水平最低的细胞分别转染miR-203模拟物（mimics）和miR-203 control,采用MTT法、流式细胞仪PI／Annexin V双染法和Transwell法检测转染miR 203对细胞增殖、凋亡和侵袭能力的影响。结果 结直肠癌组织中miR-203的相对表达量为0.372±0.019,低于癌旁组织的miR-203水平,且其表达与TNM分期、淋巴结转移、肿瘤大小及术前癌胚抗原水平均有关（P＜0.05）;与NCM460细胞相比（其miR-203表达水平设为1.00）,结直肠癌细胞SW480、Lovo和HT-29的miR 203相对表达量依次为0.26±0.07、0.44±0.05和0.32±0.04,选取SW480细胞进行后续实验。MTT检测显示,转染miR-203 mimics 48、72和96 h的SW480细胞吸光值均低于转染miR-203 control组,差异均有统计学意义（P＜0.05）;且转染miR-203 mimics 48和96 h的SW480细胞凋亡率高于转染miR-203 control组,但穿膜细胞数低于转染miR-203 control组（P＜0.05）。结论 miR-203在结直肠癌组织和细胞中低表达,上调miR 203水平可抑制结直肠癌细胞的增殖、侵袭并诱导凋亡。     

miR-203下调Bmi-1基因对肺腺癌细胞侵袭转移的影响

目的 探讨miR-203在肺腺癌中的表达,并分析其与肺腺癌细胞侵袭转移的关系及其分子机制。方法 实时定量PCR检测40例肺腺癌患者肿瘤组织中miR-203的相对表达水平及其与临床病理特征之间的关系;实时定量PCR检测H1650、A549、H1975、SPC-A-1肺腺癌细胞株中miR-203表达水平;生物信息学软件预测miR-203潜在的靶基因;脂质体2000介导miR-203模拟物、Bmi-1基因或Bmi-1 siRNA转染H1975细胞株;Western blotting检测Bmi-1蛋白水平;双荧光素酶报告基因验证miR-203是否作用于Bmi-1 mRNA的3’UTR 区预测靶位;Transwell小室侵袭实验检测H1975细胞株的侵袭转移能力。结果 40例肺腺癌组织中miR-203的相对表达量为0.065±0.013;肺腺癌miR-203表达与淋巴结转移有关,而与其他临床病理参数均无关;miR-203在肺腺癌细胞株H1650、A549、H1975、SPC-A-1中的相对表达量分别为0.280±0.102、0.308±0.168、0.167±0.073和0.287±0.096。生物信息学软件预测Bmi-1是miR-203的潜在靶基因;过表达miR-203可明显降低Bmi-1蛋白表达水平;双荧光素酶报告基因检测证明miR-203可作用于Bmi-1基因mRNA的 3’UTR区预测靶位。过表达miR-203+Bmi-1 siRNA可显著抑制肺腺癌细胞株H1975的侵袭迁移能力;在miR-203过表达的H1975细胞株中同时过表达Bmi-1可恢复其侵袭能力。结论 miR-203可通过下调Bmi-1基因表达抑制H1975肺腺癌细胞株的侵袭转移,是一种潜在抑制转移的miRNA分子。     

miR-338通过靶向GPX4抑制非小细胞肺癌细胞增殖

目的:探讨miRNA-338（miR-338）通过靶向调控谷胱甘肽过氧化物酶4（GPX4）表达在非小细胞肺癌(non-small cell lung cancer,NSCLC)增殖中的作用及机制研究。方法:通过实时定量聚合酶链式反应（qRT-PCR）检测非小细胞肺癌组织、癌旁、细胞系及对照细胞系中miR-338及GPX4 mRNA的表达水平;通过Western Blot检测非小细胞肺癌组织、癌旁、细胞系及对照细胞系中GPX4蛋白水平;通过荧光素酶报告基因时间验证miR-338直接靶向调节GPX4的表达;通过CCK-8探索miR-338是否通过调节铁死亡影响肿瘤细胞增殖;通过试剂盒检测细胞脂质氧化和活性氧水平。结果:NSCLC组织和细胞系中miR-338表达水平低于癌旁组织和对照细胞系;NSCLC组织和细胞系中GPX4 mRNA及蛋白表达水平高于癌旁组织和对照细胞系;Starbase软件分析发现GPX4 mRNA序列中含有miR-338特异作用位点,荧光素酶报告基因实验结果证实miR-338直接靶向调节GPX4表达;过表达miR-338提高肿瘤细胞中脂质氧化及活性氧水平,并抑制肿瘤细胞增殖;而铁死亡抑制剂预处理可以逆转miR-338的抑癌作用。结论:miR-338通过负向调控GPX4表达进而促进肿瘤细胞铁死亡,最终抑制NSCLC细胞增殖。     

微小RNA-494在肿瘤发生发展中的作用

微小RNA-494 (miR-494)参与正常细胞的细胞周期调控、分化和凋亡等过程.近年来研究表明,miR-494的异常表达与肿瘤的发生密切相关,其参与肿瘤细胞的侵袭和转移等过程.miR-494是一种肿瘤抑制基因,也可作为癌基因,其通过多种靶基因、信号通路对肿瘤发生发展进行调控.     

Curcumin modulates microRNA-203-mediated regulation of the Src-Akt axis in bladder cancer

Bladder cancer is often associated with recurrence and progression to invasive metastatic disease that have palliative therapeutic options. The use of traditional chemotherapeutic agents for bladder cancer management often suffers from toxicity and resistance concerns. This emphasizes the need for development of safer, natural, nontoxic compounds as chemotherapeutic/chemopreventive agents. Curcumin (diferuloylmethane) is a natural compound that has been known to possess anticancer properties in various cancers, including bladder cancer. However, the biological targets of curcumin are not well defined. Recently, it has been proposed that curcumin may mediate epigenetic modulation of expression of microRNAs (miRNA). In this article, we define for the first time, that curcumin directly induces a tumor-suppressive miRNA, miR-203, in bladder cancer. miR-203 is frequently downregulated in bladder cancer due to DNA hypermethylation of its promoter. We studied the functional significance of miR-203 in bladder cancer cell lines and found that miR-203 has tumor suppressive properties. Also, we define Akt2 and Src as novel miR-203 targets in bladder cancer. Curcumin induces hypomethylation of the miR-203 promoter and subsequent upregulation of miR-203 expression. This leads to downregulation of miR-203 target genes Akt2 and Src that culminates in decreased proliferation and increased apoptosis of bladder cancer cells. This is the first report that shows a direct effect of curcumin on inducing epigenetic changes at a miRNA promoter with direct biological consequences. Our study suggests that curcumin may offer a therapeutic advantage in the clinical management of refractory bladder cancer over other standard treatment modalities.     

ING1b-inducible microRNA203 inhibits cell proliferation

Background:

The ING family of type II tumour suppressors serve as both epigenetic ‘readers'' and target histone acetyl transferase (HAT) and histone deacetylase (HDAC) ‘writers'' of the epigenetic histone code. The ING1 protein has also been implicated in regulating microRNA (miRNA) levels. In this study, we identify a link between ING1b and the miRNA epigenetic network.

Methods:

Primary fibroblasts infected with adenoviruses expressing GFP control or GFP plus ING1b were examined for alterations in miRNA profiles using a miRNA PCR array. Additional experiments confirmed specificity and consequences of altered miRNA expression.

Results:

MicroRNAs miR-203, miR-375, miR-449b and miR-200c were increased by ING1b overexpression. Ectopic expression of miR-203 inhibited U2OS and MDA-MB-231 cancer cell growth, and induced G1 cell cycle arrest in U2OS cells as estimated by flow cytometry. Transfection with miR-203 inhibitor reversed the proliferation inhibition induced by ING1b in U2OS cells. CHIP assays showed that ING1b bound to the promoter of miR-203. Western blot analyses showed that CDK6, c-Abl and Src were downregulated by the transfection of miR-203.

Conclusion:

These results indicate that ING1b epigenetically regulates several miRNAs including miR-203. The several-fold increase in miR-203 by ING1b might inhibit cancer cell proliferation through coordinate downregulation of CDK6, c-Abl and Src.     

Down-regulation of miR-203 induced by Helicobacter pylori infection promotes the proliferation and invasion of gastric cancer by targeting CASK

Several microRNAs (miRNA) have been implicated in H. pylori related gastric cancer (GC). However, the molecular mechanism of miRNAs in GC has not been fully understood. In this study, we reported that miR-203 is significantly down-regulated in H. pylori positive tissues and cells and in tumor tissues with important functional consequences. Ectopic expression of miR-203 dramatically suppressed cell proliferation and invasion. We found that miR-203 strongly reduced the expression of CASK oncogene in GC cells. Similar to the restoring miR-203 expression, CASK down-regulation inhibited cell growth and invasion, whereas CASK over-expression rescued the suppressive effect of miR-203. These results can also be found in nude mice. In clinical specimens, CASK was over-expressed in tumors and H. pylori positive tissues and its mRNA levels were inversely correlated with miR-203 expression. Taken together, our results indicated that miR-203 functions as a growth-suppressive miRNA in H. pylori related GC, and that its suppressive effects are mediated mainly by repressing CASK expression.     

microRNA在胃癌中的生物学作用

microRNA（miRNA）是一类含量丰富且高度保守的非编码小RNA分子,调控细胞分裂、分化、调亡等重要的生物学过程,并且在很多疾病中表达失调。miRNA在多种肿瘤的发生、发展中发挥重要作用。目前,对miRNA的研究正逐步从肿瘤分子研究走向临床应用领域,作为生物标志物用于癌症的诊断以及作为靶分子用于治疗已逐渐得到确认。本文对miRNA在胃癌中作为一个潜在的生物标志物用于诊断、预后、药物反应以及易感性的预测等方面的潜能进行综述。      

miR-203a-3p在胰腺癌组织与细胞中的表达及其对BxPC-3细胞增殖、迁移和侵袭的影响

目的:探讨miR-203a-3p对胰腺癌BxPC-3细胞增殖、迁移和侵袭能力的影响.方法:运用癌症基因组图谱(TCGA)数据库筛选胰腺癌组织和癌旁组织中差异表达的miRNA,分析miRNA高表达与低表达时胰腺癌患者的生存率和临床分期;利用TarBase数据库分析miRNA与癌症相关的GO功能与KEGG通路,利用DIAN...     

miRNA-203表达水平对结直肠癌患者预后的临床价值

目的:探讨miRNA-203表达水平对结直肠癌患者预后的预测价值。方法:回顾性选择2010年1月至2013年12月来我院接受手术切除治疗的结直肠癌患者120例,所有患者均经病理确认。根据结直肠癌组织中miRNA-203表达水平将患者分为miRNA-203低表达组（n=42）和miRNA-203高表达组（n=78）。分析结直肠癌组织中miRNA-203表达水平与临床病理特征的关系,应用单因素、多因素非条件Cox回归分析影响结直肠癌患者预后的危险因素,并采用Kaplan-Meier法绘制累积生存曲线。结果:结直肠癌患者组织中miRNA-203表达量与年龄、性别、吸烟史、嗜酒史、组织类型、肿瘤位置、肿瘤直径无关（P>0.05）,与分化程度、TNM分期、淋巴结转移、脉管浸润有关（P<0.05）。单因素、多因素Cox回归分析结果显示,分化程度为高分化、TNM分期为IV期、组织中miRNA-203低表达是影响结直肠癌患者无进展生存率和总生存率的危险性因素（P<0.05）。Kaplan-Meier法生存曲线结果显示,miRNA-203高表达组结直肠癌患者无进展生存率和总生存率显著高于低表达组（P<0.05）。结论:miRNA-203表达水平可作为预测结直肠癌患者预后的指标。     

肿瘤干细胞与microRNA的研究进展

肿瘤干细胞是肿瘤的起源性细胞, 具有高度的致瘤性和耐药性。微小RNA(microRNA或miRNA)是由21~25个核苷酸组成的内源性非编码单链RNA, 是基因表达调控因子, 参与许多生物功能的调节。最近的研究发现, microRNA参与肿瘤干细胞的分化、自我更新等生物学特性的调控。肿瘤干细胞和microRNAs可以作为肿瘤研究的一个新的切入点。本文就近年来的研究进展做简要综述。      

The screening of a microRNA expression during development of human macrophages and mouse dendritic cells

There is increasing evidence showing specific roles of microRNA in cell differentiation and cancer progression. Here we examine miRNA profiles during maturation of monocytes and bone marrow-derived dendritic cells (BMDCs) in human and mouse, respectively. We have identified significant changes of various miRNA expression during monocyte and BMDC monocyte development via miRNA microarrays, confirmed by quantitative PCR. Increases in miR155 expression positively correlated with increasing maturity of monocyte and BMDC in both mouse and human microarrays, indicating its importance in development. We describe a requirement of miR155 for MHCII expression during GM-CSF-induced development and LPS-induced maturation of DCs, suggesting reduced immune function of DC when miR155 is absent. Our study suggests that miRNAs might have an important role in differentiation of myeloid cell such as dendritic cells and macrophages.