Department of Toxicology, School of Public Health, Lanzhou University, Lanzhou, Gansu 730000, China.
Department of Gastroenterology, Third People's Hospital of Gansu Province, Lanzhou, Gansu 730000, China.
Chin Med J (Engl). 2019 Sep 20;132(18):2213-2222. doi: 10.1097/CM9.0000000000000427.
MicroRNAs (miRNAs) have played important roles in the regulation of gene expression in many cancers, but their roles in esophageal squamous cell carcinoma (ESCC) are still unclear. The aim of this study was to determine the potential ESCC-specific key miRNAs from a large sample dataset in The Cancer Genome Atlas (TCGA).
Integrative bioinformatics analysis was used to identify key ESCC-specific miRNAs related to the ESCC patients' tumor histological grade and lymphatic metastasis from TCGA. Next, these key miRNA potential gene regulatory functions and relationships with ESCC patients' clinical characteristics and overall survival were analyzed. Finally, three key miRNAs were selected randomly and quantificational real-time polymerase chain reaction (qRT-PCR) was used to validate in 51 newly diagnosed ESCC patients' tissues samples (collected from Nov. 2017 to Feb. 2019, in Wuwei, China) whether the bioinformatics analyses results were reliable and valid. Two-tailed Student's t test, Pearson Chi-squared test and Kaplan-Meier survival analysis were used in this study.
Thirty-five ESCC-specific miRNAs from TCGA database were investigated (fold-change > 2.0, P < 0.05), and 28 participated in the miRNAs-mRNAs co-expression network construction, while 17 were related with ESCC patients' tumor histological grade, TNM stage, and lymphatic metastasis (P < 0.05). Meanwhile, six miRNAs (including miR-200b-3p, miR-31-5p, miR-15b-5p, miR-141-3p, miR-135b-5p, and miR-195-5p) were correlated with overall survival of ESCC patients (log-rank, P < 0.05). MiR-135b-5p, miR-15b-5p, and miR-195-5p were selected for verification of the expression levels in 51 ESCC patients' tissue samples by using qRT-PCR. We found that the fold-changes between qRT-PCR and TCGA were completely consistent. The results also suggested that miR-135b-5p, miR-15b-5p, and miR-195-5p were significantly correlated with tumor differentiation degrees (P < 0.05), miR-195-5p was significantly correlated with tumor TNM stage (P < 0.05), and miR-135b-5p was significantly correlated with lymph-node metastasis (P < 0.05). MiR-135b-5p, miR-15b-5p, and miR-195-5p expression levels, ESCC patient clinical features association analysis results and the aforementioned TCGA bioinformatics analyses were similar.
This study identified key ESCC-related miRNAs. The key miRNAs are worthy of further investigation as potential novel biomarkers for diagnosis, classification, and prognosis of ESCC.
微小 RNA（miRNA）在许多癌症的基因表达调控中发挥着重要作用，但它们在食管鳞状细胞癌（ESCC）中的作用尚不清楚。本研究的目的是从癌症基因组图谱（TCGA）中的大型样本数据集中确定 ESCC 特异性关键 miRNA。
采用整合生物信息学分析方法，从 TCGA 中鉴定出与 ESCC 患者肿瘤组织学分级和淋巴转移相关的 ESCC 特异性关键 miRNA。接下来，分析这些关键 miRNA 潜在的基因调控功能及其与 ESCC 患者临床特征和总生存期的关系。最后，随机选择 3 个关键 miRNA，并采用定量实时聚合酶链反应（qRT-PCR）在 51 例新诊断的 ESCC 患者组织样本（2017 年 11 月至 2019 年 2 月，中国武威）中验证生物信息学分析结果的可靠性和有效性。本研究采用了双尾学生 t 检验、Pearson 卡方检验和 Kaplan-Meier 生存分析。
从 TCGA 数据库中研究了 35 个 ESCC 特异性 miRNA（fold-change>2.0，P<0.05），其中 28 个参与了 miRNA-mRNAs 共表达网络的构建，而 17 个与 ESCC 患者的肿瘤组织学分级、TNM 分期和淋巴转移有关（P<0.05）。同时，有 6 个 miRNA（包括 miR-200b-3p、miR-31-5p、miR-15b-5p、miR-141-3p、miR-135b-5p 和 miR-195-5p）与 ESCC 患者的总生存期相关（log-rank，P<0.05）。miR-135b-5p、miR-15b-5p 和 miR-195-5p 被选择用于通过 qRT-PCR 验证 51 例 ESCC 患者组织样本中的表达水平。我们发现 qRT-PCR 与 TCGA 的倍数变化完全一致。结果还表明，miR-135b-5p、miR-15b-5p 和 miR-195-5p 与肿瘤分化程度显著相关（P<0.05），miR-195-5p 与肿瘤 TNM 分期显著相关（P<0.05），miR-135b-5p 与淋巴结转移显著相关（P<0.05）。miR-135b-5p、miR-15b-5p 和 miR-195-5p 的表达水平、ESCC 患者临床特征与上述 TCGA 生物信息学分析结果相似。
本研究鉴定了关键的 ESCC 相关 miRNA。这些关键 miRNA作为 ESCC 诊断、分类和预后的潜在新型生物标志物值得进一步研究。
