miR-509-3p在肝癌组织中的表达及对肝癌细胞迁移侵袭的影响

背景与目的：肝细胞癌（hepatocellular carcinoma，HCC）是世界范围内致死率第三高的恶性肿瘤，microRNA（miRNA）被认为在HCC的发病机制中起重要作用。本研究旨在探讨在原发性肝癌中miR-509-3p的表达水平、细胞功能及调控的相关基因。方法：通过实时荧光定量聚合酶链反应（real-time fluorescent quantitative polymerase chain reaction，RTFQ-PCR）技术检测miR-509-3p在46例肝癌患者组织样本、人肝癌细胞系及永生化人正常肝细胞系中的表达水平，采用transwell实验检测miR-509-3p对肝癌细胞转移的影响。采用蛋白质印迹法（Western blot）检测miR-509-3p与上皮-间质转化（epithelial-mesenchymal transition，EMT）相关基质金属蛋白酶（matrix metalloproteinase，MMP）之间的关系。结果：RTFQ-PCR技术检测结果显示，miR-509-3p在46例肝癌患者的肝癌组织样本以及MHCC97H、HCCLM3和SMMC-7721肝癌细胞系中异常高表达。体外功能实验证实抑制miR-509-3p表达能降低MHCC97H和HCCLM3肝癌细胞的迁移能力。Western blot检测结果显示，在MHCC97H和HCCLM3肝癌细胞中抑制miR-509-3p的表达可以降低MMP-9和MMP-2蛋白的表达。结论：本研究表明miR-509-3p通过正向调控MMP-9和MMP-2蛋白的表达而促进肝癌细胞的迁移和侵袭。     

微小RN A表达比在骨肉瘤中的预测作用

目的：探讨微小RNA（miRNA）表达比在骨肉瘤中的生物标志物作用。方法选取经病理证实为骨肉瘤患者的血样22例,年龄相近的30例健康者血样作为对照组。运用高通量芯片的方法对差异表达的miRNA进行筛选,对分析结果进行qRT-PCR验证;用受试者工作特征曲线分析miRNA表达比,找出最具预测潜能的组合型标志物。结果与正常血样相比,基因芯片检测出258个差异表达的miRNA（F＝5．564,P＜0．05）,选取其中4个miRNA在22例骨肉瘤患者血样和正常对照组血样中进行验证。其中miR-181b、miR-199b-5p、miR-451在骨肉瘤血样中的表达较正常血样明显增高（F＝6．283, P＜0．05）,而miR-124在骨肉瘤中则是低表达（F＝7．201,P＜0．05）;miR-199b-5p/miR-124的表达比表现出很高的敏感性（96％）和特异性（97％）。结论 miRNA在骨肉瘤中的异常表达使其具备一定的预测作用,而miR-199b-5p/miR-124表达比或许可以成为预测骨肉瘤的生物标志物。     

血清微小RNA(miR-129-3p、miR-767-3p和miR-877)在结直肠癌诊断中的价值

目的:探讨血清微小RNA (microRNA,mi RNA)在结直肠癌诊断中的价值.方法:通过miRNA表达谱芯片检测7例结直肠癌患者血清和10例健康志愿者血清中差异表达的miRNA.应用实时荧光定量PCR法在40例结直肠癌患者血清和18例健康志愿者血清中验证芯片结果,并分析血清特异性miRNA在结直肠癌诊断中的价值.结果:筛选获得10个在结直肠癌中特异性表达的血清miRNA,实时荧光定量PCR验证后获得一组结直肠癌特异性血清miRNA(miR-129-3p、miR-767-3p及miR-877*)生物标志物,这组生物标志物组合检测结直肠癌的灵敏度为77.78％、特异度为100％,并可产生最大受试者工作特征曲线(receiver operator characteristic curve,ROC)的曲线下面积(area under the curve,AUC)为0.914.结论:miR-129-3p、miR-767-3p和miR-877*生物标志物组合有望成为结直肠癌筛查和早期诊断的指标.     

miR-10a-5p基因甲基化对卵巢癌铂类耐药的影响

目的 卵巢癌是死亡率最高的妇科肿瘤,较强的化疗耐药性是其预后差的主要原因之一,为了阐明卵巢癌对铂类药物的耐药机制,本研究探讨miRNA基因的甲基化水平对卵巢癌铂类耐药的影响.方法 将卵巢癌组织分为敏感组和耐药组,每组各3例;采用基因芯片技术,对比分析了两组微RNA(microRNA,miRNA)的表达差异;采用实时荧光定量PCR,分别在6例敏感和3例耐药组织、铂类药物敏感(CoC1)和耐药的卵巢癌细胞系(CoC1/DDP),检测了候选miRNA的表达差异;应用Massarray技术,检测敏感组织(15例)与耐药组织(6例)中miRNA基因启动子的甲基化差异;应用生物信息学分析,鉴定目标miRNA的潜在靶基因.结果 以铂类药物敏感的卵巢癌组织样本为对照,利用基因芯片筛选,鉴定了6条在耐药组织样本中出现表达上调的miRNA(miR-493-3p、miR-10a-5 p、miR-16-2-3p、miR-1248、miR-451a、miR-628-3p)和6条表达下调的miRNA(miR-509-3p、miR-1197、miR-376a-3p、miR-1273a、miR-550a-3p、miR-19b-3p).组织验证发现,miR-509-3p、miR-493-3p、miR-10a-5p、miR-16-2-3p和miR-451a,与芯片结果一致;培养细胞研究发现,4条miRNA的表达调控方式与组织芯片结果一致,miR-10a-5p、miR-16-2-3p、miR-1248和miR-628-3p在耐药细胞系中高表达.进一步研究发现,与敏感肿瘤组织相比,耐药组织中miR-10a-5p基因启动子的甲基化水平出现显著降低,P=0.04.结合生物信息学预测HOXA1和USF2为miR-10a-5p与耐药相关的靶基因.结论 与敏感组相比,耐药组miR-10a-5p基因启动子甲基化水平显著降低,miR-10a-5p表达升高,通过抑制 HOXA1和USF2,抑制细胞凋亡,导致铂类化疗药物耐受.     

miR-124在胃癌中的表达及临床意义

背景与目的：miR-124在多种肿瘤中发挥抑癌基因样功能,如肺癌、前列腺癌、膀胱癌和乳腺癌,但是其在胃癌中的表达及临床意义尚不清楚。该研究旨在研究miR-124在正常胃黏膜上皮细胞与不同胃癌细胞及胃癌组织及正常胃黏膜组织中的表达,分析其表达与胃癌患者性别、年龄、组织学分级、T分期、TNM分期、淋巴结转移及预后之间的相关性。方法：采用实时荧光定量聚合酶链反应(real-time fluorescent quantitative polymerase chain-reaction,RTFQ-PCR)检测miR-124在人胃黏膜上皮细胞及胃癌细胞中的表达,采用原位杂交法检测miR-124在胃癌及癌旁正常组织中的表达。结果：RTFQ-PCR结果显示,miR-124在胃癌MKN-74、MKN-28、MKN-45、MGC-803、SGC-7901及AGS细胞中的表达均低于GES-1细胞;原位杂交实验结果显示,miR-124在正常胃黏膜中呈强阳性表达,在胃腺癌组织中表达下降、局灶阳性或缺失;统计分析显示,miR-124与胃腺癌患者组织学分级、TNM分期及淋巴结转移密切相关,与患者的年龄、性别及肿瘤大小无关;Kaplan-Meier生存曲线统计分析显示,miR-124低表达患者的总生存时间和无病生存时间显明低于miR-124高表达患者;多因素分析结果提示,miR-124表达下调是影响患者生存的独立预后因素。结论：miR-124在胃癌细胞及组织中表达下调,并且与患者的组织学分级、TNM分期、淋巴结转移及预后密切相关。     

血清miR-196a-5p、miR-105-5p在良恶性肺结节鉴别诊断中的价值

目的:探讨miR-196a-5p、miR-105-5p在良、恶性肺结节患者血清中的表达水平差异及其在恶性肺结节中的诊断价值。方法:分析癌症基因组图谱(TCGA)数据库中肺腺癌、肺鳞状细胞癌癌组织以及癌旁组织miRNA表达水平,筛选癌组织与癌旁组织相比表达水平差异明显的miRNA作为目的miRNA。选取2019年6月至2...     

miR-296在卵巢癌中的表达及其靶基因的预测研究

目的 观察miR-296在卵巢癌中的表达及其靶基因的预测.方法 收集卵巢癌患者组织样本22例和妇科活检为正常的样本22例,在提取RNA后进行RNA纯度检测,进而检测miR-296在卵巢癌中的表达;在卵巢癌细胞中稳定转入高表达miR-296的质粒,miRNA芯片进行靶基因检测.结果 卵巢癌组织中miR-296高表达,与正常对照组相比,差异有统计学意义(P＜0.001).在卵巢癌细胞系中,当高表达miR-296时,microRNA143和microRNA215在转染组中高表达,microRNA96、microRNA551b、microRNA502-3p低表达.结论 miR-296在卵巢癌中高表达,且其潜在的靶基因有可能作为临床卵巢癌诊断的标志.     

骨肉瘤血清miR-337-3p、miR-484、miR-582表达及临床意义

目的 检测骨肉瘤血清miR-337-3p、miR-484、miR-582表达及其临床意义。方法 收集我院2016年1月至2022年1月期间收治的67例骨肉瘤患者的临床病历资料，另纳入同期60例健康志愿者作为对照组。采用DEGseq方法对血清样本进行全基因组miRNA分析，采用实时荧光定量PCR(qPCR)分析血清和组织样本中miRNA水平。受试者工作特征(ROC)曲线分析miRNA诊断骨肉瘤的效能。Logistic回归模型分析影响骨肉瘤新辅助化疗疗效的因素。Cox风险比例回归模型分析影响骨肉瘤预后的因素。结果 经全基因组miRNA分析，骨肉瘤患者血清中miR-337-3p、miR-484、miR-582、miR-1912、miR-4535的表达显著下调。qPCR检测结果显示，骨肉瘤血清miR-337-3p(0.519±0.249 vs. 1.015±0.420)、miR-484(0.669±0.254 vs. 1.029±0.515)、miR-582(0.516±0.258 vs. 0.996±0.381)均低于健康对照组(P<0.001),同时骨肉瘤组织中miR-337-3p、...     

血清miRNA作为结直肠癌患者诊断标志物的价值研究

[目的]探讨特征性血液miRNA作为结直肠癌诊断标志物的可行性.[方法]用miRNA的定量PCR芯片,从结直肠癌患者和健康对照者血清中筛选出候选的特征性miR-NA.再以线虫cel-39作为外参,采用相对定量法分析36例结直肠癌患者及25名健康对照者血清中候选miRNA表达情况.[结果]9条miRNA呈特征性表达,其中5条miRNA明显上调,4条明显下调;9条特征性表达的miRNA均得以验证.将9条特征性血清miRNA进行聚类分析,可见其能够清晰地将结直肠癌患者与健康对照者分为两类.采用ROC曲线分析发现被检测结直肠癌样本的曲线下面积(AUC)达0.934 (95％CI:0.877～0.992)(P＜0.001).[结论]miR-21等9条miRNA组合检测可作为结直肠癌诊断的生物标志物.     

循环miR-29a和miR-150与非小细胞肺癌胸部放射治疗剂量的相关性

目的:探讨非小细胞肺癌（non-small cell lung cancer,NSCLC）胸部放射治疗剂量与循环血miR-29a和miR-150的相关性。方法:收集56例2014年1月至2015年12月在我院接受放射治疗的诊断为NSCLC的患者,其中5例NSCLC患者在0、20、40、60 Gy辐照后用miRNA芯片检测循环血miRNA表达差异;51例NSCLC患者在0、20、40 Gy辐照后用实时定量PCR验证循环血中候选miRNA表达;医用直线加速器（2 Gy/天,连续处理3天）辐照A549和MRC5细胞,实时定量PCR检测细胞内和细胞上清外泌体miR-29a和miR-150的表达。结果:miRNA芯片筛选出随着患者放疗剂量的增加而差异表达的10个miRNA（miR-29a、miR-150、miR-142、miR-342、miR-125b、miR-101、miR-425、miR-338、miR-126、miR-15b）。验证发现验证组患者0、20、40 Gy辐照后循环血miR-29a和miR-150表达具有统计学差异（P<0.05）。验证组循环miR-29a和miR-150分别与V5、V20、MLD、Mean Eso呈负相关。A549及MRC5细胞辐照3天后细胞内miR-29a和miR-150表达显著增加（P<0.05）,细胞上清外泌体中表达显著下降（P<0.05）。结论:循环血miR-29a和miR-150与NSCLC胸部放射治疗剂量相关。     

Plasma Circulating Mirnas Profiling for Identification of Potential Breast Cancer Early Detection Biomarkers

Objective: This study aimed to characterize the miRNA expression profiles from plasma samples of our local breast cancer patients in comparison to healthy control by using miRNA PCR Array. Methods: In this study, plasma miRNA profiles from eight early-stage breast cancer patients and nine age-matched (± 2 years) healthy controls were characterized by miRNA array-based approach, followed by differential gene expression analysis, Independent T-test and construction of Receiver Operating Characteristic (ROC) curve to determine the capability of the assays to discriminate between breast cancer and the healthy control. Results: Based on the 372-miRNAs microarray profiling, a set of 40 differential miRNAs was extracted regarding to the fold change value at 2 and above. We further sub grouped 40 miRNAs of breast cancer patients that were significantly expressed at 2-fold change and higher. In this set, we discovered that 24 miRNAs were significantly upregulated and 16 miRNAs were significantly downregulated in breast cancer patients, as compared to the miRNA expression of healthy subjects. ROC curve analysis revealed that seven miRNAs (miR-125b-5p, miR-142-3p, miR-145-5p, miR-193a-5p, miR-27b-3p, miR-22-5p and miR-423-5p) had area under curve (AUC) value > 0.7 (AUC p-value < 0.05). Overlapping findings from differential gene expression analysis, ROC analysis, and Independent T-Test resulted in three miRNAs (miR-27b-3p, miR-22-5p, miR-145-5p). Cohen’s effect size for these three miRNAs was large with d value are more than 0.95. Conclusion: miR-27b-3p, miR-22-5p, miR-145-5p could be potential biomarkers to distinguish breast cancer patients from healthy controls. A validation study for these three miRNAs in an external set of samples is ongoing.     

miR-124, miR-137 and miR-340 regulate colorectal cancer growth via inhibition of the Warburg effect

Colorectal cancer represents one of the most challenging diseases. Increasing evidence indicates that aberrant expression of microRNAs (miRNAs) is related to pathogenesis of colorectal cancer. Cancer cells reprogram metabolic pathways to sustain higher proliferation rates. Whether mechanisms underlying the role of miRNA in colorectal cancer are involved in metabolic reprogramming and the mechanisms through which miRNAs alter cancer metabolism are as yet unknown. Herein, we show that miR-124, miR-137 and miR-340 are associated with poor prognosis of colorectal cancer. Expression of these miRNAs inhibits the growth of colorectal cancer cells. PKM (pyruvate kinase isozyme) alternative splicing proteins (PTB1/hnRNAPA1/hnRNAPA2), which control the inclusion of exon 9 (PKM1) or exon 10 (PKM2), are targeted by miR-124, miR-137 and miR-340. Consequently, miR-124, miR-137 and miR-340 switch PKM gene expression from PKM2 to PKM1. High ratios of PKM1/PKM2 inhibit the glycolysis rate, but elevate the glucose flux into oxidative phosphorylation. These results demonstrate that miRNAs (miR-124, miR-137 and miR-340) impair colorectal cancer growth by counteracting the Warburg effect due to regulating alternative splicing of the PKM gene.     

Circulating exosomal miR-125a-5p as a novel biomarker for cervical cancer

Exosomal microRNAs (miRs/miRNAs) have been reported to be associated with cervical cancer. The aim of the present study was to investigate circulating exosomal miRNA as a biomarker for cervical cancer diagnosis. In the present study, samples from 6 patients with cervical cancer and 6 healthy control subjects were retrieved for exosomal RNA-sequencing. The results revealed that a total of 39 miRNAs were differentially expressed between patients with cervical cancer and healthy controls (P<0.001; fold-change >2.0). Exosomal miR-125a-5p was further quantified in plasma from 60 subjects, which included 22 healthy individuals and 38 patients with cervical cancer. miR-16a-5p served as the reference miRNA for quantitative PCR analysis of exosomal miR-125a-5p in patients with cervical cancer and healthy individuals. The results revealed that exosomal miR-125a-5p expression levels in the patients with cervical cancer were significantly lower than those in the healthy controls (P<0.001). Receiver operating characteristic (ROC) curve analyses were performed and the results revealed that the level of plasma exosomal miR-125a-5p was a potential marker for differentiating between non-cervical cancer and cervical cancer, with an ROC area under the curve of 0.7129. At the cut-off value of 2.537 for miR-125a-5p, cervical cancer diagnostic sensitivities and specificities were 59.1 and 84.2%, respectively. The present study provides confirmation that exosomal miR-125a-5p could potentially serve as a biomarker for cervical cancer diagnosis. The present study involved only a small number of clinical samples; more samples are required to support the conclusions of the present study.     

Expression of microRNAs in the urine of patients with bladder cancer

BackgroundMicroRNAs (miRNA) have been implicated to play an important role in the pathogenesis of a variety of cancers. We studied the levels of miRNAs related to epithelial-mesenchymal transition (EMT) in the urine of patients with bladder cancer.MethodThe expression of the miR-200 family, miR-205, miR-192, miR-155, and miR-146a in the urine sediment and supernatant of 51 patients with bladder cancer and in 24 controls was determined by real-time quantitative polymerase chain reaction.ResultsCompared with controls, the patients with bladder cancer had a lower expression of the miR-200 family, miR-192, and miR-155 in the urinary sediment; lower expression of miR-192; and higher expression of miR-155 in the urinary supernatant. The expression of the miR-200 family, miR-205, and miR-192 in the urine sediment significantly correlated with urinary expression of EMT markers, including zinc finger E-box-binding homeobox 1, vimentin, transforming growth factor β1, and Ras homolog gene family, member A. Furthermore, the levels of miR-200c and miR-141 in the urine sediment became normalized after surgery.ConclusionWe found that the urinary miR-200 family, miR-155, miR-192, and miR-205 levels are depressed in patients with bladder cancer. The level of these miRNA targets in urine has the potential to be developed as noninvasive markers for bladder cancer.     

Identification of Differentially Expressed Mirna by Next Generation Sequencing in Locally Advanced Breast Cancer Patients of South Indian Origin

Purpose: miRNAs are known to be aberrantly expressed in the serum, tissue, and Peripheral Blood Mononuclear Cells (PBMC) of cancer patients and could serve as potential noninvasive diagnostic markers for breast cancer. The aim of this study was to identify the differentially expressed miRNA using next-generation sequencing (NGS) from the paired PBMC samples from breast cancer patients and age-matched healthy individuals and explore their functional significance. Methods: In this study, PBMCs were employed for the detection of miRNAs by NGS in locally advanced breast cancer (LABC) women of South Indian origin who were divided into three age groups, (a) 40yrs-50yrs (b) 50yrs-60yrs and (c) 60yrs-70yrs, compared with age-matched control groups. Results: Four miRNAs (hsa-miR-192-5p, hsa-miR-24-2-2p, hsa-miR-3609, and hsa-miR-664b-3p) were found to be differentially expressed among LABC patients compared with age matched healthy women of the South Indian population. While miR-24-2-5p, miR3609, and miR-664b-3p were down-regulated, miR-192-5p was up-regulated. Gene Ontology (GO) annotations implicated miRNA with signaling pathways in peripheral nerve synapses, glutamatergic synapse, and cell morphogenesis, all of which play a pivotal role in the manifestation of cancer. Conclusion: Four miRNAs- 3 (While miR-24-2-5p, miR3609, and miR-664b-3p) downregulated and one upregulated (miR-192-5p) were identified as potential biomarkers for patients with locally advanced breast cancer. These markers could be validated in studies with a larger sample size.     

Oncogenic role of miR-17-92 cluster in anaplastic thyroid cancer cells

Micro RNAs (miRNAs) are non-coding small RNAs and constitute a novel class of negative gene regulators that are found in both plants and animals. Several miRNAs play crucial roles in cancer cell growth. To identify miRNAs specifically deregulated in anaplastic thyroid cancer (ATC) cells, we performed a comprehensive analysis of miRNA expressions in ARO cells and primary thyrocytes using miRNA microarrays. MiRNAs in a miR-17-92 cluster were overexpressed in ARO cells. We confirmed the overexpression of those miRNAs by Northern blot analysis in ARO and FRO cells. In 3 of 6 clinical ATC samples, miR-17-3p and miR-17-5p were robustly overexpressed in cancer lesions compared to adjacent normal tissue. To investigate the functional role of these miRNAs in ATC cells, ARO and FRO cells were transfected with miRNA inhibitors, antisense oligonucleotides containing locked nucleic acids. Suppression of miR-17-3p caused complete growth arrest, presumably due to caspase activation resulting in apoptosis. MiR-17-5p or miR-19a inhibitor also induced strong growth reduction, but only miR-17-5p inhibitor led to cellular senescence. On the other hand, miR-18a inhibitor only moderately attenuated the cell growth. Thus, we have clarified functional differences among the members of the cluster in ATC cells. In conclusion, these findings suggest that the miR-17-92 cluster plays an important role in certain types of ATCs and could be a novel target for ATC treatment. ( Cancer Sci 2008; 99: 1147–1154)