MiR-150-5p通过靶向调控Rab1A抑制乳腺癌细胞MDA-231的上皮-间质转化

先前的研究表明,miR-150-5p发挥抑癌基因的作用,调控肿瘤细胞的侵袭与转移。然而,关于其在乳腺癌细胞侵袭与转移中的机制尚不明确。本实验旨在研究miR-150-5p负向调控Rab1A在乳腺癌细胞上皮-间质转化（epithelial-mesenchymal transition,EMT）中的作用。双荧光素酶的结果显示,miR-150-5p可负向调控Rab1A。荧光定量PCR (qRT-PCR) 结果显示,miR-150-5p在乳腺癌细胞MCF-7及MDA-MB-231（MDA-231）中的表达水平明显低于正常乳腺上皮细胞MCF-10A; 在MDA-231中过表达miR-150-5p后,qRT-PCR结果显示,Rab1A mRNA的表达水平明显降低。Western印迹结果显示,过表达miR-150-5p后,miR-150-5p组细胞中的Rab1A、波形蛋白(vimentin)及N-钙黏着蛋白(N-cadherin)的表达水平相对于对照组（NC）细胞明显降低,而E-钙黏着蛋白(E-cadherin)的表达水平明显增加。Transwell侵袭和划痕实验显示,与miR-150-5p+Con组细胞相比,miR-150-5p+Rab1A组细胞的侵袭和迁移能力明显增加。qRT-PCR结果显示,miR-150-5p+Rab1A组细胞的Rab1A mRNA表达水平明显增加。Western印迹结果显示,miR-150-5p+Rab1A组细胞中的波形蛋白、N-钙黏着蛋白表达水平明显增加, 而E-钙黏着蛋白表达明显降低,过表达Rab1A后显著逆转了miR-150-5p对EMT的影响。综上所述,miR-150-5p可以通过负向调控Rab1A抑制EMT,进而抑制乳腺癌细胞的侵袭和迁移。     

沉默FOXQ1可逆转SW480细胞的上皮-间质转化

FOXQ1是FOX家族的的重要成员之一,其参与了多种人类肿瘤的上皮间质转化(epithelial- mesenchymal transition,EMT).本研究设计合成了FOXQ1基因的shRNA（short hairpin RNA）,用此转染SW480细胞,通过显微镜观察细胞形态,Transwell小室、细胞黏附试验检测转移能力及黏附能力,以探索FOXQ1与结直肠癌细胞EMT的关系.结果显示,沉默FOXQ1后,SW480细胞顶底极性及细胞间紧密连接增加,侵袭、迁移的细胞数目减少,同种黏附能力增加,异种黏附能力降低.进一步的机制研究表明,沉默FOXQ1基因可以导致SW480细胞的上皮标志因子E-cadherin表达显著增高,而间质细胞标志因子N-cadherin、Vimentin及MMP2表达均降低.以上结果表明,沉默FOXQ1基因可以逆转SW480细胞EMT,其机制可能与E-cadherin的上调和N cadherin、Vimentin、MMP2的下调有关,这为进一步研究FOXQ1在结直肠癌发生发展中的作用提供实验基础.     

Autocrine/Paracrine Human Growth Hormone-stimulated MicroRNA 96-182-183 Cluster Promotes Epithelial-Mesenchymal Transition and Invasion in Breast Cancer

Human growth hormone (hGH) plays critical roles in pubertal mammary gland growth, development, and sexual maturation. Accumulated studies have reported that autocrine/paracrine hGH is an orthotopically expressed oncoprotein that promotes normal mammary epithelial cell oncogenic transformation. Autocrine/paracrine hGH has also been reported to promote mammary epithelial cell epithelial-mesenchymal transition (EMT) and invasion. However, the underlying mechanism remains largely obscure. MicroRNAs (miRNAs) are reported to be involved in regulation of multiple cellular functions of cancer. To determine whether autocrine/paracrine hGH promotes EMT and invasion through modulation of miRNA expression, we performed microarray profiling using MCF-7 cells stably expressing wild type or a translation-deficient hGH gene and identified miR-96-182-183 as an autocrine/paracrine hGH-regulated miRNA cluster. Forced expression of miR-96-182-183 conferred on epithelioid MCF-7 cells a mesenchymal phenotype and promoted invasive behavior in vitro and dissemination in vivo. Moreover, we observed that miR-96-182-183 promoted EMT and invasion by directly and simultaneously suppressing BRMS1L (breast cancer metastasis suppressor 1-like) gene expression. miR-96 and miR-182 also targeted GHR, providing a potential negative feedback loop in the hGH-GHR signaling pathway. We further demonstrated that autocrine/paracrine hGH stimulated miR-96-182-183 expression and facilitated EMT and invasion via STAT3 and STAT5 signaling. Consistent with elevated expression of autocrine/paracrine hGH in metastatic breast cancer tissue, miR-96-182-183 expression was also remarkably enhanced. Hence, we delineate the roles of the miRNA-96-182-183 cluster and elucidate a novel hGH-GHR-STAT3/STAT5-miR-96-182-183-BRMS1L-ZEB1/E47-EMT/invasion axis, which provides further understanding of the mechanism of autocrine/paracrine hGH-stimulated EMT and invasion in breast cancer.     

MMP-3与乳腺癌上皮间质转化的关系及其意义

为了探讨MMP-3在乳腺癌中表达与乳腺癌上皮间质转化的关系及其在乳腺癌浸润转移中的作用.采用免疫组织化学检测正常乳腺组织、乳腺癌癌旁组织、乳腺癌组织中MMP-3、Claudin-1、Vimentin的表达情况.研究发现:1)MMP-3在乳腺癌组织的阳性表达率较癌旁组织及正常乳腺组织高且具有统计学意义(P<0.05).Claudin-1在乳腺癌中的阳性表达率明显低于癌旁组织及正常乳腺组织具有统计学意义(P<0.05).Vimentin在乳腺癌组织阳性表达率高于癌旁组织及正常乳腺组织具有统计学意义(P<0.05);2)随着乳腺癌病理分期的增加MMP-3表达逐渐增加,同时出现Claudin-1蛋白表达降低及Vimentin蛋白表达升高.相关分析表明MMP-3与Claudin-1的表达呈负相关(r=-0.301,P=0.019),与Vimentin的表达呈正相关(r=0.378,P=0.03),Claudin-1与Vimentin的表达呈负相关(r=-0.278,P=0.031).结果提示,MMP-3可能是通过介导乳腺癌细胞发生上皮间质转化来参与乳腺癌的浸润转移.     

MicroRNA对EMT的调节作用及其与肿瘤侵袭的关系

上皮间质转化（epithelial mesenchymal transition,EMT）是指上皮细胞表型由上皮向间质转换的生物学过程,可发生在生理过程中促进发育、组织愈合和修复。近年对肿瘤的研究发现,EMT与肿瘤的发生发展密切相关。肿瘤细胞发生EMT时,伴随着迁移、侵袭能力的增强,进而促进肿瘤的转移。EMT发生的程度以及相关标志分子的检测还可以用于判断肿瘤转移的危险和评估预后。MicroRNA(miRNA)作为非编码小RNA,通过与特定mRNA的3′UTR结合,在蛋白翻译水平抑制基因表达。本文主要综述目前发现的作用于EMT相关转录因子,如ZEB、SNAIL、TWIST的miRNA,以及在各种肿瘤中的表达情况和作用。其中,有些转录因子和miRNA之间,还存在相互抑制的复杂调节网络,因此,了解miRNA在肿瘤中对EMT的作用可能为肿瘤的治疗提供新的方法和策略。     

RSPO2 enhances cell invasion and migration via the WNT/β-catenin pathway in human gastric cancer

R-spondins comprise a group of secreted WNT agonists. R-spondin2 (RSPO2) plays a crucial role in the activation of the WNT/β-catenin pathway and oncogenesis, though its specific role in human gastric cancer (GC) remains unclear. In the current study, RSPO2 expression levels were upregulated in cancer specimens and cell lines (AGS and BGC-823). Inhibition of RSPO2 expression levels had distinct effects on cell invasion, migration, and epithelial-mesenchymal transition (EMT) in AGS and BGC-823 cells in vitro. Furthermore, RSPO2 positively correlated with leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), the receptor of RSPO2. Silencing RSPO2 reduced the expression of LGR5 and WNT/β-catenin effector molecule β-catenin together with downstream targets TCF-4 and Cyclin-D1. These observations demonstrate that upregulation of RSPO2 in GC specimens and cell lines is closely related to tumor invasion and migration and that RSPO2 promotes EMT in gastric cancer cells by activating WNT/β-catenin signaling.