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Concomitant Up-Regulation of Hsa- Mir-374 and Down-Regulation of Its Targets, GSK-3β and APC, in Tissue Samples of Colorectal Cancer

机译:伴随HSA-miR-374的上调和下调其靶标,GSK-3β和APC,在结肠直肠癌的组织样本中

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Background:The WNT-pathway is involved in several cancers, including colorectal cancer (CRC). Many cell signaling components and pathways are controlled by microRNAs. The main purpose of the present study was to investigate the expression of hsa-miR-374, and its two target genes of the Wnt-pathway in CRC clinical samples. Methods: In this study, we predicted the miRNAs targeting key genes of WNT-pathway using bioinformatics algorithms. The expression levels of hsa-miR-374, APC and GSK-3β on 48 pairs of Formalin-Fixed ParaffinEmbedded (FFPE) CRC tumors and marginal-tumors were evaluated using real time-PCR. Additionally, the hsa-miR-374a-5p precursor sequence was amplified by whole-blood DNA as a template. This amplicon was cloned into pEGFP-c1 expression vector and transfected into SW742 cells. Aside from this, MTT assay was performed to evaluate the effect of miR-374 on cell viability. Results: The bioinformatics analysis indicated that hsa-miR-374 binds to the regulatory region the key components of WNT-pathway, including APC and GSK-3β considering the recognition elements and mirSVR scores. Our results revealed significant down-regulation of GSK-3β (0.94 times, p= 0.0098) and APC (0.96 times, p= 0.03) and up-regulation of miR-374 (1.22 times, p= 0.0071) on tumor samples compared with their normal pairs. Meanwhile, the results of the over-expression of miR-374 showed down-regulation of APC and GSK-3β. MTT-assay also indicated that the miR-374 increased cell survival. Conclusions:The results of our study indicated a concomitant change in the expression of miR-374 and its two related target genes, in clinical samples of CRC. Hsa-miR-374 might be as a helpful biomarker or therapeutic target in CRC.
机译:背景:WNT途径参与了几种癌症,包括结肠直肠癌(CRC)。许多细胞信号传导组分和途径由MicroRNA控制。本研究的主要目的是研究HSA-MIR-374的表达及其在CRC临床样品中WNT通路的两种靶基因。方法:在本研究中,我们预测使用生物信息学算法预测靶向Wnt-途径的关键基因。使用实时PCR评估HSA-miR-374,APC和GSK-3β的表达水平,在48对福尔马林固定的石蜡床上(FFPE)CRC肿瘤和边缘肿瘤。另外,将HSA-miR-374A-5P前体序列通过全血DNA作为模板扩增。将该扩增子克隆到PEGFP-C1表达载体中并转染到SW742细胞中。除此之外,进行MTT测定以评估miR-374对细胞活力的影响。结果:生物信息学分析表明,HSA-MIR-374与监管区结合,考虑到识别元件和MIRSVR分数,包括APC和GSK-3β的关键组分。我们的结果表明,GSK-3β(0.94倍,P = 0.0098)和APC(0.96倍,P = 0.03)和MIR-374(1.22倍,P = 0.0071)的显着下调(0.96倍,P = 0.0071)与肿瘤样品相比他们的正常对。同时,MIR-374的过表达结果显示了APC和GSK-3β的下调。 MTT测定还表明miR-374增加了细胞存活率。结论:我们研究的结果表明MIR-374及其两种相关靶基因表达的伴随变化,CRC的临床样本。 HSA-MIR-374可能是CRC中有用的生物标志物或治疗靶标。

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