Microarray data analysis on gene and miRNA expression to identify biomarkers in non-small cell lung cancer.
ABSTRACT: BACKGROUND:The aim of this study was to gain further investigation of non-small cell lung cancer (NSCLC) tumorigenesis and identify biomarkers for clinical management of patients through comprehensive bioinformatics analysis. METHODS:miRNA and mRNA microarray datasets were downloaded from GEO (Gene Expression Omnibus) database under the accession number GSE102286 and GSE101929, respectively. Genes and miRNAs with differential expression were identified in NSCLC samples compared with controls, respectively. The interaction between differentially expressed genes (DEGs) and differentially expressed miRNAs (DEmiRs) was predicted, followed by functional enrichment analysis, and construction of miRNA-gene regulatory network, protein-protein interaction (PPI) network, and competing endogenous RNA (ceRNA) network. Through comprehensive bioinformatics analysis, we anticipate to find novel therapeutic targets and biomarkers for NSCLC. RESULTS:A total of 123 DEmiRs (5 up- and 118 down-regulated miRNAs) and 924 DEGs (309 up- and 615 down-regulated genes) were identified. These genes and miRNAs were significantly involved in different pathways including adherens junction, relaxin signaling pathway, and axon guidance. Furthermore, hsa-miR-9-5p, has-miR-196a-5p and hsa-miR-31-5p, as well as hsa-miR-1, hsa-miR-218-5p and hsa-miR-135a-5p were shown to have higher degree in the miRNA-gene regulatory network and ceRNA network, respectively. Furthermore, BIRC5 and FGF2, as well as RTKN2 and SLIT3 were hubs in the PPI network and ceRNA network, respectively. CONCLUSION:Several pathways (adherens junction, relaxin signaling pathway, and axon guidance) miRNAs (hsa-miR-9-5p, has-miR-196a-5p, hsa-miR-31-5p, hsa-miR-1, hsa-miR-218-5p and hsa-miR-135a-5p) and genes (BIRC5, FGF2, RTKN2 and SLIT3) may play important roles in the pathogenesis of NSCLC.
Project description:Background Circular RNA (circRNA) is a noncoding RNA that forms a closed-loop structure, and its abnormal expression may cause disease. We aimed to find potential network for circRNA-related competitive endogenous RNA (ceRNA) in atrial fibrillation (AF). Methods The circRNA, miRNA, and mRNA expression profiles in the heart tissue from AF patients were retrieved from the Gene Expression Omnibus database and analyzed comprehensively. Differentially expressed circRNAs (DEcircRNAs), differentially expressed miRNAs (DEmiRNAs), and differentially expressed mRNAs (DEmRNAs) were identified, followed by the establishment of DEcircRNA-DEmiRNA-DEmRNA regulatory network. Functional annotation analysis of host gene of DEcircRNAs and DEmRNAs in ceRNA regulatory network was performed. In vitro experiment and electronic validation were used to validate the expression of DEcircRNAs, DEmiRNAs, and DEmRNAs. Results A total of 1611 DEcircRNAs, 51 DEmiRNAs, and 1250 DEmRNAs were identified in AF. The DEcircRNA-DEmiRNA-DEmRNA network contained 62 circRNAs, 14 miRNAs, and 728 mRNAs. Among which, two ceRNA regulatory pairs of hsa-circRNA-100053-hsa-miR-455-5p-TRPV1 and hsa-circRNA-005843-hsa-miR-188-5p-SPON1 were identified. In addition, six miRNA-mRNA regulatory pairs including hsa-miR-34c-5p-INMT, hsa-miR-1253-DDIT4L, hsa-miR-508-5p-SMOC2, hsa-miR-943-ACTA1, hsa-miR-338-3p-WIPI1, and hsa-miR-199a-3p-RAP1GAP2 were also obtained. MTOR was a significantly enriched signaling pathway of host gene of DEcircRNAs. In addition, arrhythmogenic right ventricular cardiomyopathy, dilated cardiomyopathy, and hypertrophic cardiomyopathy were remarkably enriched signaling pathways of DEmRNAs in DEcircRNA-DEmiRNA-DEmRNA regulatory network. The expression validation of hsa-circRNA-402565, hsa-miR-34c-5p, hsa-miR-188-5p, SPON1, DDIT4L, SMOC2, and WIPI1 was consistent with the integrated analysis. Conclusion We speculated that hsa-circRNA-100053-hsa-miR-455-5p-TRPV1 and hsa-circRNA-005843-hsa-miR-188-5p-SPON1 interaction pairs may be involved in AF.
Project description:Purpose:This research explores the aberrant expression of the long non-coding RNA (lncRNA), microRNA (miRNA), and messenger RNA (mRNA) in pterygium. A competitive endogenous RNA (ceRNA) network was constructed to elucidate the molecular mechanisms in pterygium. Methods:We obtained the differentially expressed mRNAs based on three datasets (GSE2513, GSE51995, and GSE83627), and summarized the differentially expressed miRNAs (DEmiRs) and differentially expressed lncRNAs (DELs) data by published literature. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, protein-protein interaction (PPI), and gene set enrichment analysis (GSEA) analysis were performed. DEmiRs were verified in GSE21346, and the regulatory network of hub mRNAs, DELs, and DEmiRs were constructed. Results:Overall, 40 upregulated and 40 downregulated differentially expressed genes (DEGs) were obtained. The KEGG enrichment showed the DEGs mainly involved in extracellular matrix (ECM)-receptor interaction, focal adhesion, and PI3K-Akt signaling pathway. The GSEA results showed that cornification, keratinization, and cornified envelope were significantly enriched. The validation outcome confirmed six upregulated DEmiRs (miR-766-3p, miR-184, miR-143-3p, miR-138-5p, miR-518b, and miR-1236-3p) and two downregulated DEmiRs (miR-200b-3p and miR-200a-3p). Then, a ceRNA regulatory network was constructed with 22 upregulated and 15 downregulated DEmiRs, 4 downregulated DELs, and 26 upregulated and 33 downregulated DEGs. The network showed that lncRNA SNHG1/miR-766-3p/FOS and some miRNA-mRNA axes were dysregulated in pterygium. Conclusions:Our study provides a novel perspective on the regulatory mechanism of pterygium, and lncRNA SNHG1/miR-766-3p/FOS may contribute to pterygium development.
Project description:Objective:Lung cancer has high incidence and mortality rate, and non-small cell lung cancer (NSCLC) takes up approximately 85% of lung cancer cases. This study is aimed to reveal miRNAs and genes involved in the mechanisms of NSCLC. Materials and Methods:In this retrospective study, GSE21933 (21 NSCLC samples and 21 normal samples), GSE27262 (25 NSCLC samples and 25 normal samples), GSE43458 (40 NSCLC samples and 30 normal samples) and GSE74706 (18 NSCLC samples and 18 normal samples) were searched from gene expression omnibus (GEO) database. The differentially expressed genes (DEGs) were screened from the four microarray datasets using MetaDE package, and then conducted with functional annotation using DAVID tool. Afterwards, protein-protein interaction (PPI) network and module analyses were carried out using Cytoscape software. Based on miR2Disease and Mirwalk2 databases, microRNAs (miRNAs)-DEG pairs were selected. Finally, Cytoscape software was applied to construct miRNA-DEG regulatory network. Results:Totally, 727 DEGs (382 up-regulated and 345 down-regulated) had the same expression trends in all of the four microarray datasets. In the PPI network, TP53 and FOS could interact with each other and they were among the top 10 nodes. Besides, five network modules were found. After construction of the miRNA-gene network, top 10 miRNAs (such as hsa-miR-16-5p, hsa-let-7b-5p, hsa-miR-15a-5p, hsa-miR-15b-5p, hsa-let-7a-5p and hsa-miR-34a- 5p) and genes (such as HMGA1, BTG2, SOD2 and TP53) were selected. Conclusion:These miRNAs and genes might contribute to the pathogenesis of NSCLC.
Project description:Idiopathic pulmonary fibrosis (IPF) is a fibrotic interstitial lung disease with lesions confined to the lungs. To identify meaningful microRNA (miRNA) and gene modules related to the IPF progression, GSE32537 (RNA-sequencing data) and GSE32538 (miRNA-sequencing data) were downloaded and processed, and then weighted gene co-expression network analysis (WGCNA) was applied to construct gene co-expression networks and miRNA co-expression networks. GSE10667, GSE70866, and GSE27430 were used to make a reasonable validation for the results and evaluate the clinical significance of the genes and the miRNAs. Six hub genes (COL3A1, COL1A2, OGN, COL15A1, ASPN, and MXRA5) and seven hub miRNAs (hsa-let-7b-5p, hsa-miR-26a-5p, hsa-miR-25-3p, hsa-miR-29c-3p, hsa-let-7c-5p, hsa-miR-29b-3p, and hsa-miR-26b-5p) were clarified and validated. Meanwhile, iteration network of hub miRNAs-hub genes was constructed, and the emerging role of the network being involved in non-small cell lung cancer (NSCLC) was also analyzed by several webtools. The expression levels of hub genes were different between normal lung tissues and NSCLC tissues. Six genes (COL3A1, COL1A2, OGN, COL15A1, ASPN, and MXRA5) and three miRNAs (hsa-miR-29c-3p, hsa-let-7c-5p, and hsa-miR-29b-3p) were related to the survival time of lung adenocarcinoma (LUAD). The interaction network of hub miRNAs-hub genes might provide common mechanisms involving in IPF and NSCLC. More importantly, useful clues were provided for clinical treatment of both diseases based on novel molecular advances.
Project description:Increasing evidence has shown competitive endogenous RNAs (ceRNAs) play key roles in numerous cancers. Nevertheless, the ceRNA network that can predict the prognosis of lung adenocarcinoma (LUAD) is still lacking. The aim of the present study was to identify the prognostic value of key ceRNAs in lung tumorigenesis. Differentially expressed (DE) RNAs were identified between LUAD and adjacent normal samples by limma package in R using The Cancer Genome Atlas database (TCGA). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway function enrichment analysis was performed using the clusterProfiler package in R. Subsequently, the LUAD ceRNA network was established in three steps based on ceRNA hypothesis. Hub RNAs were identified using degree analysis methods based on Cytoscape plugin cytoHubba. Multivariate Cox regression analysis was implemented to calculate the risk score using the candidate ceRNAs and overall survival information. The survival differences between the high-risk and low-risk ceRNA groups were determined by the Kaplan-Meier and log-rank test using survival and survminer package in R. A total of 2,989 mRNAs, 185 lncRNAs, and 153 miRNAs were identified. GO and KEGG pathway function enrichment analysis showed that DE mRNAs were mainly associated with "sister chromatid segregation," "regulation of angiogenesis," "cell adhesion molecules (CAMs)," "cell cycle," and "ECM-receptor interaction." LUAD-related ceRNA network was constructed, which comprised of 54 nodes and 78 edges. Top ten hub RNAs (hsa-miR-374a-5p, hsa-miR-374b-5p, hsa-miR-340-5p, hsa-miR-377-3p, hsa-miR-21-5p, hsa-miR-326, SNHG1, RALGPS2, and PITX2) were identified according to their degree. Kaplan-Meier survival analyses demonstrated that hsa-miR-21-5p and RALGPS2 had a significant prognostic value. Finally, we found that a high risk of three novel ceRNA interactions (SNHG1-hsa-miR-21-5p-RALGPS2, SNHG1-hsa-miR-326-RALGPS2, and SNHG1-hsa-miR-377-3p-RALGPS2) was positively associated with worse prognosis. Three novel ceRNAs (SNHG1-hsa-miR-21-5p-RALGPS2, SNHG1-hsa-miR-326-RALGPS2, and SNHG1-hsa-miR-377-3p-RALGPS2) might be potential biomarkers for the prognosis and treatment of LUAD.
Project description:Long non?coding RNAs (lncRNAs) have been implicated in the development and progression of cancer. However, the mechanisms of lncRNAs in hepatitis B virus (HBV) infection?induced hepatocellular carcinoma (HCC) remain unclear. The study aimed to reveal the roles of lncRNAs for HBV?HCC based on the hypothesis of competing endogenous RNA (ceRNA). The lncRNA (GSE27462), miRNA (GSE76903) and mRNA (GSE121248) expression profiles were collected from the Gene Expression Omnibus database. Differentially expressed lncRNAs (DELs), genes (DEGs) and miRNAs (DEMs) were identified using the LIMMA or EdgeR package, respectively. The ceRNA network was constructed based on interaction pairs between miRNAs and mRNAs/lncRNAs. The functions of DEGs in the ceRNA network were predicted using the DAVID database, which was overlapped with the known HCC pathways of Comparative Toxicogenomics Database (CTD) to construct the HCC?related ceRNA network. The prognosis values [overall survival, (OS); recurrence?free survival (RFS)] of genes were validated using the Cancer Genome Atlas (TCGA) data with Cox regression analysis. The present study screened 38 DELs, 127 DEMs and 721 DEGs. A ceRNA network was constructed among 17 DELs, 12 DEMs and 173 DEGs, including the FAM138B?hsa?miR?30c?CCNE2/RRM2 and SSTR5?AS1?hsa?miR?15b?5p?CA2 ceRNA axes. Function enrichment analysis revealed the genes in the ceRNA network that participated in the p53 signaling pathway [cyclin E2 (CCNE2), ribonucleotide reductase M2 subunit (RRM2)] and nitrogen metabolism [carbonic anhydrase 2 (CA2)], which were also included in the pathways of the CTD. Univariate Cox regression analysis revealed that six RNAs (2 DELs: FAM138B, SSTR5?AS1; 2 DEMs: hsa?miR?149, hsa?miR?7; 2 DEGs: CCNE2, RRM2) were significantly associated with OS; while seven RNAs (1 DEL: LINC00284; 3 DEMs: hsa?miR?7, hsa?miR?15b, hsa?miR?30c?2; and 3 DEGs: RRM2, CCNE2, CA2) were significantly associated with RFS. In conclusion, FAM138B?hsa?miR?30c?CCNE2/RRM2 and the SSTR5?AS1?hsa?miR?15b?5p?CA2 ceRNA axes may be important mechanisms for HBV?related HCC.
Project description:microRNAs (miRNAs) have been implicated in the control of many biological processes and their deregulation has been associated with many cancers. In recent years, the cancer stem cell (CSC) concept has been applied to many cancers including pediatric. We hypothesized that a common signature of deregulated miRNAs in the CSCs fraction may explain the disrupted signaling pathways in CSCs.Using a high throughput qPCR approach we identified 26 CSC associated differentially expressed miRNAs (DEmiRs). Using BCmicrO algorithm 865 potential CSC associated DEmiR targets were obtained. These potential targets were subjected to KEGG, Biocarta and Gene Ontology pathway and biological processes analysis. Four annotated pathways were enriched: cell cycle, cell proliferation, p53 and TGF-beta/BMP. Knocking down hsa-miR-21-5p, hsa-miR-181c-5p and hsa-miR-135b-5p using antisense oligonucleotides and small interfering RNA in cell lines led to the depletion of the CSC fraction and impairment of sphere formation (CSC surrogate assays).Our findings indicated that CSC associated DEmiRs and the putative pathways they regulate may have potential therapeutic applications in pediatric cancers.
Project description:Growing evidence has illustrated critical roles of competing endogenous RNA (ceRNA) regulatory network in human cancers including hepatocellular carcinoma. In this study, we aimed to find promising diagnostic and prognostic biomarkers for patients with hepatocellular carcinoma. Three novel unfavorable prognosis-associated genes (CELSR3, GPSM2, and CHEK1) was first identified. We also demonstrated that these genes were significantly upregulated in hepatocellular carcinoma cell lines and tissues. Next, 154 potential miRNAs of CELSR3, GPSM2, and CHEK1 were predicted. CHEK1-hsa-mir-195-5p/hsa-mir-497-5p and GPSM2-hsa-mir-122-5p axes were defined as two key pathways in carcinogenesis of hepatocellular carcinoma by combination of in silico analysis and experimental validation. Subsequently, lncRNAs binding to hsa-mir-195-5p, hsa-mir-497-5p, and hsa-mir-122-5p were predicted via starBase and miRNet databases. After performing expression analysis and survival analysis for these predicted lncRNAs, we showed that nine lncRNAs (SNHG1, SNHG12, LINC00511, HCG18, FGD5-AS1, CERS6-AS1, NUTM2A-AS1, SNHG16, and ASB16-AS1) were markedly increased in hepatocellular carcinoma and their upregulation indicated poor prognosis. Moreover, a similar mRNA-miRNA-lncRNA analysis for six "known" genes (CLEC3B, DNASE1L3, PTTG1, KIF2C, XPO5, and UBE2S) was performed. Subsequently, a comprehensive mRNA-miRNA-lncRNA triple ceRNA network linked to prognosis of patients with hepatocellular carcinoma was established. Moreover, all RNAs in this network exhibited significantly diagnostic values for patients with hepatocellular carcinoma. In summary, the current study constructed a mRNA-miRNA-lncRNA ceRNA network associated with diagnosis and prognosis of hepatocellular carcinoma.
Project description:Accumulating evidence has indicated that noncoding RNAs are involved in intervertebral disc degeneration (IDD); however, the competing endogenous RNA (ceRNA)?mediated regulatory mechanisms in IDD remain rarely reported. The present study aimed to comprehensively investigate the alterations in expression levels of circular RNA (circRNA), long noncoding RNA (lncRNA), microRNA (miRNA/miR) and mRNA in the nucleus pulposus (NP) of patients with IDD. In addition, crucial lncRNA/circRNA?miRNA?mRNA ceRNA interaction axes were screened using the GSE67567 microarray dataset obtained from the Gene Expression Omnibus database. After data preprocessing, differentially expressed circRNAs (DECs), lncRNAs (DELs), miRNAs (DEMs) or genes (DEGs) between IDD and normal controls were identified using the Linear Models for Microarray data method. A protein?protein interaction (PPI) network was constructed for DEGs based on protein databases, followed by module analysis. The ceRNA network was constructed based on the interaction between miRNAs and mRNAs, and lncRNAs/circRNAs and miRNAs. The underlying functions of mRNAs were predicted using the Database for Annotation, Visualization and Integrated Discovery database. The present study identified 636 DECs, 115 DELs, 84 DEMs and 1,040 DEGs between patients with IDD and control individuals. PPI network analysis demonstrated that Fos proto?oncogene, AP?1 transcription factor subunit (FOS), mitogen?activated protein kinase 1 (MAPK1), hypoxia inducible factor 1 subunit ? (HIF1A) and transforming growth factor ?1 (TGFB1) were hub genes and enriched in modules. Metastasis?associated lung adenocarcinoma transcript 1 (MALAT1)/hsa_circRNA_102348?hsa??miR?185?5p?TGFB1/FOS, MALAT1?hsa?miR?155?5p?HIF1A, hsa_circRNA_102399?hsa?miR?302a?3p?HIF1A, MALAT1?hsa??miR?519d?3p?MAPK1 and hsa_circRNA_100086?hsa?miR?509?3p?MAPK1 ceRNA axes were obtained by constructing the ceRNA networks. In conclusion, these identified ceRNA interaction axes may be crucial targets for the treatment of IDD.
Project description:The purpose of this work was to extract key players such as mRNAs and long non-coding RNA (lncRNAs) in the etiopathogenesis of osteosarcoma (OS). The sequencing analyses (mRNAs and lncRNAs) of OS were conducted followed by differentially expressed mRNAs and lncRNAs (DEmRNAs and DElncRNAs) identification between U-2OS cells with has-miR-590-5p overexpression and negative control cells. Following this, the co-expression and functional enrichment analyses of DEmRNAs and DElncRNAs were carried out. Also, the miRNAs-DElncRNAs-DEmRNAs regulatory network was constructed with DElncRNAs-miRNAs and DElncRNAs-DEmRNAs pairs after the target gene analysis of miRNA. In addition, the ceRNA-has-miR-590-5p was further extracted based on the has-miR-590-5p-DElncRNAs and DElncRNAs-DEmRNAs interactions. Finally, the results of the bioinformatics analysis was verified by reverse-transcription polymerase chain reaction (RT-PCR). Totally, 980 DEmRNAs (539 up-regulated DEmRNAs and 441 down-regulated DEmRNAs) and 682 DElncRNAs (352 up-regulated DElncRNAs and 330 down-regulated DElncRNAs) were extracted between cells with hsa-miR-590-5p overexpression and normal cells. The functional analyses suggested that up-regulated genes were significantly enriched in several GO terms such as signal transduction and cytokine-cytokine receptor interaction pathway while down-regulated genes (SCUBE3, HIST1H4E and EDIL3) were associated with calcium ion binding, cell surface function and nucleosome assembly. Additionally, the miRNAs-DEmRNAs-DEmRNAs network represented 220 pairs among 41 miRNAs, 38 DElncRNAs and 61 DEmRNAs. Furthermore, the ceRNA-hsa-miR-590-5p network consisted of 70 interaction pairs including hsa-miR-590-5p-SCUBE3-CTB-113D17.1, hsa-miR-590-5p-EDIL3-CTB-113D17.1 and hsa-miR-590-5p-HIST1H4E-CTB-113D17.1) among hsa-miR-590-5p, 30 DEmRNAs and 4 down-regulated DElncRNAs. Meanwhile, the RT-PCR results incidated that compared with the blank (KB) and negative control (NC) group, the mRNA expression of SCUBE3, HIST1H4E, and EDIL3 were significantly descreased in mimics group (P value <0.05). The lncRNA CTB-113D17.1 might implicate with OS development probably via serving as a hsa-miR-590-5p sponge to regulate gene targets (SCUBE3, EDIL3 and HIST1H4E), which will facilitate the deep understandings of OS progression.