TargetScan Database Research Articles

Aloin Induces Gastric Cancer Cell Apoptosis via the miR-5683/HMGB1 Signal Axis

Background: Recent studies conducted by us indicate that aloin (ALO) could promote gastric cancer (GC) cell apoptosis, but the underlying mechanism remains unclear. The high mobility group box 1 (HMGB1) has been reported to regulate the apoptosis, proliferation, and migration of several cancer types. Bioinformatics analysis suggests a possible targeted regulatory relationship between miR5683 and HMGB1. Purpose: Herein, we aimed to investigate the possible role of the miR-5683/HMGB1 axis in ALO-induced GC cell apoptosis. Methods: The expression levels of miR-5683 and HMGB1 in GC tissue and adjacent normal tissue were investigated using bioinformatics analysis, and their expressions at the cellular levels were determined using reverse transcription quantitative real-time polymerase chain reaction and western blotting (WB) assays. The interaction between miR-5683 and HMGB1 was predicted in the TargetScan database and verified by a dual-luciferase assay. Cell viability and apoptosis were assessed by the cell counting kit 8 (CCK-8) assay, 4′,6-diamidino-2-phenylindole (DAPI) staining, and flow cytometry. WB was used to examine the expression levels of apoptosis-related proteins and HMGB1. Results: Our data showed that ALO promoted GC cell apoptosis, downregulated HMGB1, upregulated miR-5683 expression, and dual-luciferase assay confirmed the interaction between HMGB1 and miR-5683. Moreover, miR-5683 mimics enhanced ALO-induced GC cell apoptosis and the inhibitory effects of ALO on the levels of HMGB1. However, miR-5683 inhibitors showed the opposite effects. Conclusion: Our data suggested that ALO promotes GC cell apoptosis through the miR-5683/HMGB1 axis.

Identification of circRNA-miRNA-mRNA network in luminal breast cancers by integrated analysis of microarray datasets.

Introduction: Circular RNAs (circRNAs) regulatory network is important in human cancer. We, therefore, mapped the regulatory networks driven by circRNA in luminal-subtype breast cancer. Methods: Breast cancer-related microarray datasets from GEO database were analyzed for the differentially expressed circRNAs, miRNAs, and mRNAs. The potential downstream RNAs were collected using Circular RNA Interactome or Targetscan database. Protein-protein interaction (PPI) analysis was performed for the filtered genes to identify hub genes. The functions were annotated by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. CircRNA-miRNA-mRNA networks were mapped using Cytoscape software. Hsa_circ_0086735-miR-1296-5p-STAT1 axis was used for verification. The expression levels of hsa_circ_0086735, miR-1296-5p, and STAT1 mRNA were confirmed by qRT-PCR in luminal-subtype tissues and cell lines. The interactions among them were verified by Luciferase reporter assay and RNA pull-down assay. Cell proliferation and apoptosis were assayed. Overall and distant metastasis-free survival was analyzed. Results: A total of 70 genes were finally targeted and enriched in multi-process and multi-pathway. Networks containing 96 circRNA-miRNA-mRNA axes were constructed. Hsa_circ_0086735 and STAT1 mRNA was upregulated in luminal breast cancer, while miR-1296-5p was downregulated. Hsa_circ_0086735-miR-1296-5p-STAT1 axis promotes breast cancer progression and contributes to tamoxifen resistance. High hsa_circ_0086735 was associated with poor overall and distant metastasis-free survival. Discussion: This study identified the hsa_circ_0086735-miR-1296-5p-STAT1 as an important regulatory axis in luminal-subtype breast cancer, aiding to determine potential therapeutic targets.

Genes whose expressions in the primary lung squamous cell carcinoma are able to accurately predict the progression of metastasis through lymphatic system, inferred from a bioinformatics analyses

Lymph node metastasis is the most important prognostic factor in patients with lung squamous cell carcinoma. The current findings show that lymph node metastatic tumor cells can arise by programming metastasis in primary tumor cells. Thereby, the genetic alterations responsible for the metastasis could be detected in the primary tumors. This bioinformatic study aimed to determine novel potential prognostic biomarkers shared between primary lung squamous cell tumors (without lymph node metastasis) and lymphatic metastasis, using the Cancer Genome Atlas database. Differentially expressed genes were screened by limma statistical package in R environment. Gene ontology and biological pathways analyses were performed using Enrichr for up-regulated and down-regulated genes. Also, we selected lymph node metastasis related genes among DEGs using correlation analysis between DEGs and suitable references genes for metastasis. Receiver operating characteristic curves was applied using pROC and R package ggplot2 to evaluate diagnostic value of differentially expressed genes. In addition, survival and drug resistance analyses were performed for differentially expressed genes. The miRNA-mRNA interaction networks were predicted by miRwalk and TargetScan databases and expression levels analysis of the miRNAs which were mainly targeting mRNAs was performed using UALCAN database. Protein–protein interaction network analysis and hub genes identification were performed using FunRich and Cytoscape plugin cytoHubba. In this study, a total of 397 genes were differentially expressed not only with a significant difference between N + vs. normal and N0 vs. normal but also with significant difference between N + vs. N0. Identified GO terms and biological pathways were consistent with DEGs role in the lung squamous cell carcinoma and lymph node metastasis. A significant correlation between 56 genes out of 397 differentially expressed genes with reference genes prompted them being considered for identifying lymph node metastasis of lung squamous cell carcinoma. In addition, SLC46A2, ZNF367, AC107214.1 and NCBP1 genes were identified as survival-related genes of patients with lung squamous cell carcinoma. Moreover, NEDD9, MRPL21, SNRPF, and SCLT1 genes were identified to be involved in lung squamous cell carcinoma drug sensitivity/resistance. We have identified several numbers of miRNAs and their related target genes which could emerge as potential diagnostic biomarkers. Finally, CDK1, PLK1, PCNA, ZWINT and NDC80 identified as hub genes for underlying molecular mechanisms of lung squamous cell carcinoma and lymphatic metastasis. Our study highlights new target genes according to their relation to lymph node metastasis, whose expressions in the primary lung squamous cell carcinoma are able to accurately assess the presence of lymphatic metastasis.

A functional polymorphism at the miR-25-3p binding site in the 3′-untranslated region of the S1PR1 gene decreases the risk of osteoporosis in Chinese postmenopausal women

The low bone mineral density due to abnormally activated osteoclasts can induce bone disorders such as osteopenia and osteoporosis. MiR-25-3p modulates sphingosine-1-phosphate receptor 1 (S1PR1) expression to enhance osteoclast motivation. The association between miR-25 rs41274221 polymorphism and osteoporosis susceptibility is unknown. Herein, 300 patients with osteoporosis and 320 healthy controls were genotyped using polymerase chain reaction and Sanger sequencing to explore the role of miR-25 rs41274221 polymorphism in osteoporosis. The odds ratios (ORs) and 95% confidence intervals (CIs) were determined using logistic regression analysis. Additionally, this study also investigated the effect of miR-25 rs41274221 polymorphism on the expression of miR-25 and its target gene S1PR1 through luciferase reporter gene, qRT-PCR, and Western blot. The rs41274221 in miR-25 was found to be positively associated with osteoporosis and can be viewed as a protective factor. Furthermore, miR-25 rs41274221 polymorphism decreased the risk of osteoporosis among smokers. The TargetScan database predictions and dual-luciferase activity demonstrated that S1PR1 may be the target gene of miR-25. MiR-25 downregulated the S1PR1 mRNA and protein expressions. Patients with the AA genotype of rs41274221 polymorphism showed higher S1PR1 expression compared with the GG genotype. In conclusion, miR-25 rs41274221 polymorphism decreases the risk of osteoporosis through modifying the binding with S1PR1 and may serve as a novel biomarker for early diagnosis of osteoporosis.

Comprehensive analysis of exosomal circRNA-miRNA-mRNA network in breast cancer.

Exosomal circRNAs played critical roles in tumor development and progression and might be novel biomarkers in the diagnosis and treatment of various cancers. However, the biological functions and clinical implications of exosomal circRNAs in breast cancer are unclear. Expression profiles of exosomal circRNAs in exoRBase 2.0 database were used to identify differentially expressed exosomal circRNAs in breast cancer. The LASSO and SVM-RFE algorithm followed by multivariate logistic regression analysis were performed to construct the diagnostic model. The target genes of circRNAs were selected by combing differential expression analysis and CSCD, TargetScan and ENCORI database. Univariate and multivariate survival analysis were conducted to construct a survival-associated exosomal circRNA-miRNA-mRNA network. GSVA and CIBERSORT algorithm were used to evaluate the cancer hallmarks and immune cells in breast cancer and spearman correlation analysis was used to investigate their correlations with the circRNA-miRNA-mRNA network. A total of 347 upregulated and 3 downregulated exosomal circRNAs were identified in breast cancer patients. The diagnostic model basing on 14 exosomal circRNAs showed a high area under the curve (AUC) value in both the training (AUC = 0.98) and validation dataset (AUC = 0.94). A total of 70 miRNAs and 1147 mRNAs were selected as the downstream targets of circRNAs and were revealed to participate into tumor-associated pathways including PI3K-AKT, MAPK, RAS and RAP1 pathways, calcium signaling pathways and transcriptional misregulations. The constructed survival-associated exosomal circRNA-miRNA-mRNA network contained 9 exosomal circRNAs, 12 miRNAs and 10 mRNAs and showed complicated correlations and interactions within networks. Cancer hallmark pathways including TGF-β, KRAS and MYC signaling pathways, tumor angiogenesis, epithelial-mesenchymal transition, DNA repair and G2M checkpoint and immune cells including CD4+ and CD8+ T cells, dendritic cells, mast cells, macrophage cells, memory B cells and natural killer cells were closely correlated with the circRNA-miRNA-mRNA network. Our study was the first to systematically analyze the exosomal circRNAs in breast cancer. We established an exosomal circRNA diagnostic model and constructed a survival-associated exosomal circRNA-miRNA-mRNA network. Our results revealed the complicated functions and potential mechanisms of exosomal circRNA-miRNA-mRNA network in breast cancer which need to be further validated in the future.

Analysis and Validation of the Prognosis ability of the M7GRelated miRNAs in Lung Adenocarcinoma.

N7-methylguanosine (m7G) modification plays a crucial role in the development and progression of lung cancers. MicroRNAs (miRNAs) are closely involved in programmed cell death and the mechanism of tumor growth. The m7G-associated miRNAs genes in lung adenocarcinoma (LUAD), and their prognosis prediction ability of LUAD, however, had not been investigated. The RNA transcriptomes, clinical indices, and immune scores of LUAD patients were searched and downloaded from The Cancer Genome Atlas (TCGA) and the ESTIMATE database. The miRNAs targeting METTL1 and WDR4 were extracted from the TargetScan database. Differentially expressed m7G-related miRNAs were identified and their prediction power of LUAD prognosis was systematically investigated. Among 40 the differentially expressed m7G-related miRNAs in LUAD, five (hsa-miR-31-5p, hsa-miR-5571-3p, hsa-miR-4697-3p, hsa-miR-6858-5p, and hsa-miR-873-3p) demonstrate significant predictive value for prognosis. The risk score constructed by these five miRNAs was an independent prognostic factor (univariate Cox regression results: hazard ratio (HR) = 1.6619, 95% confidential interval (CI) = 1.2103-2.2819, p = 0.0017; multivariate Cox regression results: HR = 1.6004, 95% CI = 1.1633-2.2017, p = 0.0039). The survival curves showed that patients with high-risk scores had a poor prognosis. Calibration curves indicated good predictability in a nomogram constructed combining the miRNA and the clinical indices of age, sex, chemotherapy, radiotherapy, stage, and risk score. GO and KEGG analysis of the overlapping genes showed that the prognostic miRNAs were closely associated with the neuropeptide signaling pathway. Besides, the immune infiltration analysis showed that the expression of the AMPD1 gene was strongly associated with immune cells and immunology functions in LUAD. This study identified DE m7G-related miRNAs and demonstrated their prediction ability in the prognosis of LUAD patients. The risk signature based on these miRNAs demonstrates high accuracy in predicting the prognosis of LUAD patients.

Identification of epidermal growth factor receptor as an immune-related biomarker in epilepsy using multi-transcriptome data.

Epilepsy is a chronic disease that is characterized by transient brain dysfunction caused by an abrupt abnormal neuronal discharge. Recent studies have indicated that the pathways related to inflammation and innate immunity play significant roles in the pathogenesis of epilepsy, suggesting an interrelationship between immunity and inflammatory processes and epilepsy. However, the immune-related mechanisms are still not precisely understood; therefore, this study aimed to explore the immune-related mechanisms in epilepsy disorders, highlight the role of immune cells at the molecular level in epilepsy, and provide therapeutic targets for patients with epilepsy. Brain tissue samples from healthy and epileptic individuals were collected for transcriptome sequencing to identify differentially expressed genes (DEGs) and differentially expressed (DE)-long coding RNAs (lncRNAs). Based on interactions from the miRcode, starBase2.0, miRDB, miRTarBase, TargetScan, and ENCORI databases, a lncRNA-associated competitive endogenous RNA (ceRNA) network was created. Gene ontology and the Kyoto encyclopedia of genes analyses established that the genes in the ceRNA network were mainly enriched in immune-related pathways. Immune cell infiltration, screening, and protein-protein interaction analyses of the immune-related ceRNAs, and correlation analysis between immune-related core messenger RNA (mRNA) and immune cells were also performed. Nine hub genes (EGFR, GRB2, KRAS, FOS, ESR1, MAPK1, MAPK14, MAPK8, and PPARG) were obtained. Also, 38 lncRNAs, one miRNA (hsa-miR-27a-3p), and one mRNA (EGFR) comprised the final core ceRNA network. Mast cells, plasmacytoid dendritic cells, and immature dendritic cells all showed positive correlations with EGFR, while Cluster of differentiation 56 dim natural killer cells (CD56dim natural killer cells) showed negative correlations. Finally, we employed an epilepsy mouse model to validate EGFR, which is consistent with disease progression. In conclusion, the pathophysiology of epilepsy was correlated with EGFR. Thus, EGFR could be a novel biomarker of juvenile focal epilepsies, and our findings provide promising therapeutic targets for epilepsy.

CircRNA_17725 Promotes Macrophage Polarization towards M2 by Targeting FAM46C to Alleviate Arthritis.

Accumulating studies have implicated that circular RNAs (circRNAs) play vital roles in the pathogenesis of rheumatoid arthritis (RA). Dysregulation of macrophage polarization leads to immune homeostatic imbalance in RA. However, the altering effects and mechanisms of circRNAs on macrophages polarization and immune homeostatic balance remain largely unclear. We aimed to investigate the potential role of circRNA_17725 in RA. The high-throughput sequence was performed to identify the dysregulated circRNAs in RA. We confirmed the data by CCK-8, EdU, and Annexin V/PI staining to elucidate the proliferation and apoptosis. The expressions of M1/M2-associated markers were confirmed using real-time PCR and flow cytometry analysis. Luciferase reporter assay and RNA Binding Protein Immunoprecipitation (RIP) were used to demonstrate the underlying mechanism of circRNA_17725. The altering effect of circRNA_17725 on macrophages in vivo was evaluated using collagen-induced arthritis (CIA) mouse model. circRNA_17725 was demonstrated to be downregulated in peripheral blood mononuclear cells and CD14+ monocytes from RA cases in contrast to healthy controls. The negative association between circRNA_17725 and the disease activity indexes (CRP, ESR, and DAS28) was observed, suggesting a vital role of circRNA_17725 in RA disease activity. Besides, after a coexpression analysis based on high-input sequencing and the bioinformatics analysis in MiRanda and TargetScan databases, a circRNA_17725-miR-4668-5p-FAM46C competing endogenous RNA (ceRNA) network was hypothesized. A series of cytology experiments in vitro have implicated that circRNA_17725 could inhibit the proliferation but enhance the apoptosis of macrophages. Decreased expression of TNF-α, IL-1β, and MMP-9 were observed in the supernatant of circRNA_17725-overexpressed Raw264.7 macrophages, implicating the inhibitory effect of circRNA_17725 on macrophage inflammatory mediators. Furthermore, circRNA_17725 could promote macrophage polarization towards M2 by targeting miR-4668-5p/FAM46C as a miRNA sponge. Additionally, circRNA_17725-overexpressed macrophages alleviated arthritis and protected against joint injuries and bone destruction by inducing macrophage polarization towards M2 in collagen-induced arthritis (CIA) mice. This study has suggested that circRNA_17725 regulated macrophage proliferation, apoptosis, inflammation, and polarization by sponging miR-4668-5p and upregulating FAM46C in RA.

MiR-223-3p increases resistance of colorectal cancer cells to 5-fluorouracil via targeting SORBS1.

5-Fluorouracil (5-FU) is the first-line drug for treating colorectal cancer (CRC), and the resistance of tumor cells to 5-FU is the main cause of chemotherapeutic failure. However, the resistant mechanism is still unclear. This study aims to explore the tumor suppressor genes involved in 5-FU resistance in CRC, and to find the microRNA (miRNA) that regulates these genes. CRC data sets GSE28702 and GSE69657 were downloaded from Gene Expression Omnibus (GEO) database, and gene expression profiles of patients in the FOLFOX chemotherapeutic response group and the non-response group were analyzed, and differential expression genes were identified between the 2 groups. Target gene was then selected. Online bioinformatics databases TargetScan, miRwalk, and miRDB were used to predict miRNA targeting the interested gene sorbin and SH3 domain containing 1 (SORBS1). siSORBS1, HA-SORBS1, miR-223-3p mimic, anti-miR-223-3p, and their corresponding negative controls (siNC, HA, miR-NC, and anti-miR-NC) were transfected into CRC cell lines of HCT116 and SW620 by transient transfection technique, respectively. Co-transfection was done with miRNA and plasmid (miR-NC+HA, miR-223-3p mimic+HA, or miR-223-3p mimic+HA-SORBS1) or anti-miRNA and siRNA (anti-miR-NC+siNC, anti-miR-223-3p+siNC, or anti-miR-223-3p+siSORBS1) in HCT116 cells. Real-time reverse transcription PCR (real-time RT-PCR) and/or Western blotting were used to detect the expression levels of SORBS1 and miR-223-3p in cells. After transfection, the cells were treated with different concentrations of 5-FU, and the cell viability was detected by methyl thiazolyl tetrazolium (MTT) method. The targeting relationship between miR-223-3p and SORBS1 was comfirmed by dual luciferase reporter gene assay. There were 409 and 528 highly expressed genes in the FOLFOX chemotherapeutic response group of GSE69657 and GSE28702, respectively. There were 22 overlapping genes in the response group, among which exist 3 tumor suppressor genes might be involved in chemosensitivity in CRC, and SORBS1 was selected as the target gene for further study. Three online bioinformatics databases predicted miRNAs targeting SORBS1 and obtained an intersection molecule miR-223-3p. After treatment with 5-FU (25 µmol/L) for 12-36 h, the levels of miR-223-3p in HCT116 and SW620 cells were significantly down-regulated (all P<0.05). After transfection with siSORBS1 or miR-223-3p mimic, the expression levels of SORBS1 in HCT116 and SW620 cells were down-regulated, and the cell viability was increased (all P<0.05). After transfection with HA-SORBS1 or anti-miR-223-3p, the expression levels of SORBS1 in HCT116 and SW620 cells were up-regulated, and the cell viability was decreased (all P<0.05). The result of dual luciferase reporter gene assay showed that the luciferase activity of cells co-transfected with SORBS1 3'-UTR wild plasmid and miR-223-3p mimic was significantly lower than that of the 3'-UTR wild plasmid and miR-NC cells (P<0.05). Compared with co-transfection with miR-223-3p mimic and HA, the cell viability of cells co-transfected with miR-223-3p mimic and HA-SORBS1 was decreased significantly (P<0.01). Compared with the co-transfected anti-miR-223-3p and siNC, the cell viability of the co-transfected anti-miR-223-3p and siSORBS1 was significantly increased (P<0.05). MiR-223-3p increases 5-FU resistance in CRC cells by targeting SORBS1,and miR-223-3p is expected to become a new target for clinical treatment of CRC.

LncRNA ERVH48-1 Contributes to the Drug Resistance of Prostate Cancer and Proliferation through Sponging of miR-4784 to the Activation of the Wnt/β-Catenin Pathway

Long noncoding RNAs (LncRNAs) are very important in the way that docetaxel resistance (DR) happens in prostate cancer (PCa) patients. ImmuneScore and StromalScore were calculated using PCa-related expression data from TCGA and the ESTIMATE algorithm. We finally found the DEGs that were related to the immune system and the stroma of the patients by making profiles of the DEGs in ImmuneScore and StromalScore. The CancerSubtypes algorithm identified prognosis-related PCa subtypes, and the GSVA assessed their pathway activity. A UniCox regression analysis was used to identify a prognosis-related differential gene set. We then used intersection analysis to identify immunological and prognostic (IP)-related genes (IPGs). The coexpression of long noncoding RNAs (lncRNAs) and IPGs was used to identify IP-related lncRNAs (IPLs). Three methods (SVM-RFE, random forest, and LASSO) were used to find genes that overlap in the GEO database. A gene signature was then validated by building an ROC curve. CIBERSORT technology was used to look at the possibility of a link between the gene signature and immune cells. LncRNA-miRNA pairs and miRNA-mRNA pairs from the miRDB and TargetScan databases were used to construct the ERVH48-1-miR-4784-WNT2B ceRNA regulation network. The concentration of docetaxel elevated the expression of ERVH48-1. Overexpression of ERVH48-1 increased PCa-DR cell proliferation, invasion, and migration while inhibiting apoptosis. ERVH48-1 increased the tumorigenicity of PCa-DR cells in nude mice. ERVH48-1, acting as a ceRNA, targeted miR-4784 to increase WNT2B expression. ICG001 therapy increased Wnt/-catenin signaling activity in PCa-DR cells by inhibiting ERVH48-1. Finally, ERVH48-1 increased docetaxel resistance in a WNT2B-dependent manner via the miR-4784/Wnt/-catenin pathway.

Construct ceRNA Network and Risk Model of Breast Cancer Using Machine Learning Methods under the Mechanism of Cuproptosis

Breast cancer (BRCA) has an undesirable prognosis and is the second most common cancer among women after lung cancer. A novel mechanism of programmed cell death called cuproptosis is linked to the development and spread of tumor cells. However, the function of cuproptosis in BRCA remains unknown. To this date, no studies have used machine learning methods to screen for characteristic genes to explore the role of cuproptosis-related genes (CRGs) in breast cancer. Therefore, 14 cuproptosis-related characteristic genes (CRCGs) were discovered by the feature selection of 39 differentially expressed CRGs using the three machine learning methods LASSO, SVM-RFE, and random forest. Through the PPI network and immune infiltration analysis, we found that PRNP was the key CRCG. The miRTarBase, TargetScan, and miRDB databases were then used to identify hsa-miR-192-5p and hsa-miR-215-5p as the upstream miRNA of PRNP, and the upstream lncRNA, CARMN, was identified by the StarBase database. Thus, the mRNA PRNP/miRNA hsa-miR-192-5p and hsa-miR-215-5p/lncRNA CARMN ceRNA network was constructed. This ceRNA network, which has not been studied before, is extremely innovative. Furthermore, four cuproptosis-related lncRNAs (CRLs) were screened in TCGA-BRCA by univariate Cox, LASSO, and multivariate Cox regression analysis. The risk model was constructed by using these four CRLs, and the risk score = C9orf163 * (1.8365) + PHC2-AS1 * (-2.2985) + AC087741.1 * (-0.9504) + AL109824.1 * (0.6016). The ROC curve and C-index demonstrated the superior predictive capacity of the risk model, and the ROC curve demonstrated that the AUC of 1-, 3-, and 5-year OS in all samples was 0.721, 0.695, and 0.633, respectively. Finally, 50 prospective sensitive medicines were screened with the pRRophetic R package, among which 17-AAG may be a therapeutic agent for high-risk patients, while the other 49 medicines may be suitable for the treatment of low-risk patients. In conclusion, our study constructs a new ceRNA network and a novel risk model, which offer a theoretical foundation for the treatment of BRCA and will aid in improving the prognosis of BRCA.

Early diagnosis and prognosis of hepatocellular carcinoma based on a ceRNA array

Abstract Objectives Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-related deaths, due to high morbidity, a low early diagnosis rate, and poor prognosis. It is essential to explore competitive endogenous RNA (ceRNA) arrays for early diagnosis and prognosis of HCC. Methods The original gene expression profiles of differentially expressed lncRNA, miRNA and mRNA in HCC were downloaded from TCGA database. Differentially expressed lncRNA-miRNA and miRNA-mRNA interaction pairs were extracted from miRcode and starBase, a ceRNA network was constructed, and GO annotation and KEGG pathway analyses were performed. Cox regression and Kaplan-Meier survival analysis screening identified core genes in the network associated with HCC survival, centering on miRNA, which were screened using ceRNA arrays. qRT-PCR was used to detect the expression level of key genes in clinical samples. Dual luciferase reporter gene assays were used to verify the target binding relationship among lncRNA, mRNA, and miRNA. ROC curves were used to analyze the diagnostic efficacy of the ceRNA array. Results A total of 8 lncRNAs, 5 miRNAs, and 21 mRNAs were used to construct a ceRNA network. Functional enrichment analysis showed that mRNAs in the ceRNA network were mainly enriched in 14 signaling pathways, especially microRNAs in cancer. Survival analysis showed that lncRNA FOXD2-AS1 and miRNA miR-9-5p were related to the prognosis of HCC, and the targeted binding relationships between mRNAs. STMN1, COL15A1, and CCNE2 and miR-9-5p from the TargetScan, starBase, miRDB, and PicTar databases were reliable. qRT-PCR showed that expression levels of FOXD2-AS1, miR-9-5p, STMN1, COL15A1, and CCNE2 were upregulated in HCC tissues. Dual luciferase reporter assays showed that FOXD2-AS1 and STMN1 had a targeted binding relationship with miR-9-5p, but not with COL15A1 or CCNE2. The area under the curve of the candidate ceRNA array (FOXD2-AS1/miR-9-5p/STMN1/COL15A1/CCNE2) was higher than that of each member and ceRNA combination (FOXD2-AS1/miR-9-5p/STMN1). Conclusions The candidate ceRNA array formed by FOXD2-AS1/miR-9-5p/STMN1/COL15A1/CCNE2 could be a biomarker for early diagnosis and prognosis of HCC.

Analysis of Communal Molecular Mechanism Between Chronic Obstructive Pulmonary Disease and Osteoporosis.

Chronic obstructive pulmonary disease (COPD) patients with osteoporosis (OP) usually experience more frequent exacerbations, worse quality of life, and heavier economic burden, however, few studies have investigated common molecular mechanisms of COPD and OP. To explore the relationship between COPD and OP through bioinformatics analysis. The miRNA microarray data of COPD and OP were retrieved from the Gene Expression Database (GEO), and the differentially expressed microRNAs (DEmiRNAs) were screened and the intersection was obtained. The Targetscan, miRDB, and miRWalk databases were used to predict the target genes of DEmiRNA, and the gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed using the R package clusterProfiler, the STRING database was used to analyze the target protein-protein interaction network (PPI) and screens to determine the core modules and core genes. Two DEmiRNAs (miR-23a-5p, miR-194-3p) have been found in COPD and OP, which have predicted 76 and 114 target genes, respectively. GO functional annotations of miR-23a-5p were significantly enriched in CD40 signaling pathway, ubiquitin-conjugating enzyme activity, etc; KEGG pathways of miR-23a-5p were significantly enriched in ubiquitin-mediated proteolysis, folate biosynthesis, and regulation of actin cytoskeleton. GO function annotations of miR-194-3p were significantly enriched in T cell activation regulation, ubiquitin protein ligase activity, and DNA transcription factor binding; KEGG pathways of miR-194-3p were significantly enriched in cell adhesion molecules, intercellular tight junctions, and lysosomal pathway. PPI analysis found target coding proteins formed complex regulatory networks. Ten core genes (TP53, SRC, PXN, CHD4, SYK, TNRC6B, PML, KAT5, BRD1 and IGF2) were picked out among them, then we used the MCODE plugin found three core subnetworks. Two identical DEmiRNAs (miR-23a-5p, miR-194-3p) exist in the peripheral blood of COPD and OP patients, which are important biomarkers for COPD patients with OP and may represent novel targets for diagnosis and treatment of COPD patients with OP.

Differential expression profile of microRNAs in the lung tissues of coal workers with pneumoconiosis and patients with silicosis.

The purpose of this study was to characterize the microRNA (miRNA) profile of the lung tissues from coal workers' pneumoconiosis (CWP) and silicosis and to analyze the changes in downstream genes, biological processes, and signaling pathways based on the differently expressed miRNAs. Lung tissues from three CWP patients, eight silicosis patients, and four healthy controls were collected and analyzed for their miRNA profiles using Affymetrix® GeneChip® miRNA Arrays. Differentially expressed miRNAs (DEMs) were identified between the different groups. The miRanda and TargetScan databases were used to predict the putative target genes, and volcano and heat maps were drawn. Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses were then performed to screen the DEMs-associated biological process and signaling pathways, respectively. Further identification with a comprehensive literature research involving particle exposure, fibrosis, inflammation and lung cancer were used to further screen DEMs of CWP and silicosis. Microarray data showed that 375 and 88 miRNAs were differentially expressed in CWP and silicosis lung tissues compared with healthy lung tissues, while 34 miRNAs were differentially expressed in CWP compared with silicosis lung tissues. The GO and KEGG pathway analyses showed that, the target genes were mainly enriched in the TGF-β, MAPK, p53 and other signal pathways. These results provided insight into the miRNA-related underlying mechanisms of CWP and silicosis, and they provided new clues for miRNAs as biomarkers for the diagnosis and differential diagnosis of these two diseases.

MiR-4757-3p Inhibited the Migration and Invasion of Lung Cancer Cell via Targeting Wnt Signaling Pathway.

Lung cancer accounts for the vast majority of cancer-related deaths worldwide, and aberrant miRNA expression is commonly observed as the disease progresses. The current study aimed to determine the role of miR-4757-3p in the development of lung cancer. The real-time PCR test was performed to determine the expression of miR-4757-3p in lung cancer cell lines. miR-4757-3p was downregulated in A549 cells. CCK8 and transwell assays demonstrated that overexpression of miR-4757-3p significantly reduced A549 cell invasion and migration. Bioinformatic analysis by the TargetScan database predicted the possible targets of miR-4757-3p. A luciferase activity test was used to determine the direct relationship between miR-4757-3p, Wnt5a, and Wnt8b. The overexpression of miR-4757-3p drastically inhibited the expression of Wnt5a and Wnt8b. Furthermore, we discovered that silencing Wnt5a and Wnt8b significantly lowered β-catenin expression and hampered invasion and migration. Finally, miR-4757-3p inhibited lung cancer cell migration and invasion by inhibiting the activation of the Wnt signaling pathway. Our study provided evidence that miR-4757-3p could be developed as an indicator or an anticancer target in the clinical application.

FZD1/KLF10-hsa-miR-4762-5p/miR-224-3p-circular RNAs axis as prognostic biomarkers and therapeutic targets for glioblastoma: a comprehensive report

BackgroundThe circular RNA (circRNA) plays a vital role in the pathogenesis of tumors as a competitive endogenous RNA (ceRNA). Given the high aggressiveness and fatality rate of glioblastoma (GBM) as well as poor prognosis, it is necessary to construct a circRNA-related ceRNA network for further studies on the mechanism of GBM and identify possible biomarkers as well as therapeutic drugs.MethodsThree datasets from the gene expression omnibus (GEO) database were downloaded to distinguish differential circRNAs, microRNAs, and messenger RNAs respectively in GBM. With the help of GEPIA2, circBank, CSCD, TargetScan, miRDB, and miRTarBase databases, we established a circRNAs-related ceRNA network in GBM. Functional enrichments were employed to profile the most relevant mRNAs to indirectly clarify the mechanisms of the ceRNA network. Based on the expression profile data and survival information of GBM patients from the GEO and the cancer genome atlas (TCGA) databases, we performed survival analysis to select prognostic mRNAs and constructed a novel circRNA-miRNA-mRNA central regulatory subnetwork. The DGIdb database was used to find potential drug–gene interactions.ResultsThe datasets obtained from the GEO and TCGA databases were analyzed, and 504 differentially expressed mRNAs (DEmRNAs), 71 differentially expressed microRNAs (DEmiRNAs), and 270 differentially expressed circRNAs (DEcircRNAs) were screened out. The novel ceRNA regulatory network included 22 circRNAs, 11 miRNAs, and 15 mRNAs. FZD1 and KLF10 were significantly correlated with the overall survival rate of patients with GBM (P < 0.05). The final survival subnetwork contained six circRNAs, two miRNAs, and two mRNAs. Two small-molecule compounds and one antibody could be used as therapeutic drugs for GBM. Interestingly, the Wnt signaling pathway appeared in both KEGG and GO functional terms.ConclusionsResults of this study demonstrate that FZD1 and KLF10 may exert regulatory functions in GBM, and the ceRNA-mediated network could be a therapeutic strategy for GBM.

MiR 26 targeting methyltetrahydrofolate reductase inhibits the proliferation of acute myeloid leukemia cells via JAK/STAT pathway

Purpose: To investigate the effect of microribonucleic acid (miR)-26 targeting methyltetrahydrofolate reductase (MTHFR) on cell proliferation, cycle, and apoptosis in acute myeloid leukemia (AML).Methods: The expressions of miR-26 in three types of human AML cell lines (HL-60, Kasumi-1, and KG-1a) and normal myeloid cell line (HS-5) were determined via quantitative reverse transcriptionpolymerase chain reaction (qRT-PCR), while the effect of the over-expression of miR-26 on the proliferation, cell cycle and apoptosis of AML cells was evaluated using cell counting kit-8 (CCK-8) assay and flow cytometry. The potential target for miR-26 was predicted using public miRNA database TargetScan, and whether miR-26 binds to the predicted target was determined using a dual-luciferase reporter assay. Western blotting was performed to determine the effect of miR-26 on Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway-associated proteins.Results: Expression level of miR-26 was significantly lower in all AML cell lines than in HS-5 cells (p &lt; 0.05). Overexpression of miR-26 inhibited the proliferation of KG-1a cells, reduced the percentage of cells in S phase, increased those in G0/G1 phase, and enhanced apoptosis of KG-1a cells (p &lt; 0.05). After overexpression of miR-26, protein expression levels of phosphorylated (p)-JAK and p-STAT were down-regulated, while those of JAK and STAT did not change significantly.Conclusion: Expression of miR-26 is down-regulated in AML, while MiR-26 targeting of MTHFR induces apoptosis and cycle arrest of AML cells through the JAK/STAT pathway, thus inhibiting AML cell proliferation in vitro.

The lncRNA XIST/miR-29b-3p/COL3A1 axis regulates central carbon metabolism in head and neck squamous cell carcinoma and is associated with poor tumor prognosis.

Recent evidence shows that COL3A1 promotes the progression of many types of cancer. The purpose of our study is to explore the correlation between COL3A1 and the prognosis of patients with head and neck squamous cell carcinoma (HNSCC) and its potential mechanism. We initially screened the differentially expressed gene COL3A1 in The Cancer Genome Atlas (TCGA) database, and the association between the expression level of COL3A1, prognosis, and the clinical parameters of HNSCC patients was verified. A nomogram was constructed according to the multivariate analysis results. Next, a heatmap of COL3A1 co-expressed genes was constructed in TCGA database. The TargetScan database is used to explore the microRNAs (miRNA) related to COL3A1. The starBase database was used to explore and predict the long non-coding RNAs (lncRNAs) that the candidate miRNAs might bind to. Finally, the potential mechanism of action was investigated using Gene Set Enrichment Analysis (GSEA). COL3A1 expression is elevated in HNSCC tumor tissues, and HNSCC patients with high COL3A1 expression have worse prognostic factors. COL3A1 was positively correlated with the central carbon metabolism-related proteins: epidermal growth factor receptor (EGFR), phosphoglycerate mutase 1 (PGAM1), hexokinase 3 (HK3), and phosphofructokinase, platelet (PFKP). The TargetScan database showed that the best candidate miRNA for binding to the three prime untranslated region (3'UTR) end of COL3A1 mRNA was hsa-miR-29b-3p, which was negatively correlated with COL3A1. The starBase database showed that the lncRNA X Inactive Specific Transcript (lncRNA XIST) was the best candidate upstream non-coding RNA for regulating hsa-miR-29b-3p. GSEA showed that COL3A1 may be involved in the poor prognosis of HNSCC by participating in carbon metabolism, glucose metabolism, oxidative stress, and the Wingless-Type MMTV Integration Site Family (Wnt) and vascular endothelial growth factor A-vascular endothelial growth factor receptor 2 (VEGFA-VEGFR2) pathways. Low COL3A1 expression can be employed as a new HNSCC predictive biomarker, and the prognosis of HNSCC patients with lower COL3A1 expression can be greatly improved. At the same time, we found that the lncRNA XIST/miR-29b-3p/COL3A1 axis may regulate the central carbon metabolism of HNSCC and is associated with poor prognosis. These findings point to a potential target for developing HNSCC anticancer therapies.

Molecular modeling of PARP1 inhibition through miRNA gene silencing for the treatment of retinitis pigmentosa.

Retinitis Pigmentosa (RP) is a category of diseases that are characterized by the death of rod photoreceptors and subsequent death of cone receptors. Parthanatos, a form of regulated necrosis, has been reported as being the underlying mechanism of cell death in RP. Parthanatos in RP occurs with the downstream expression of poly-ADP-ribose-polymerase (PARP) and the subsequent nuclear translocation of the mitochondrial apoptosis-inducing factor (AIF), leading to condensing of chromatin and DNA fragmentation. Extracellular vesicles (EVs) are small-scale liposomes that carry miRNAs and the RNA-induced silencing complex (RISC) associated with gene regulation. miRNAs, loaded on the argonaute protein 2 (Ago-2) bind to the mature mRNA and initiate the recruitment of the RISC complex triggering the degradation of the mRNA, dramatically scaling down gene expression. In vitro validation of gene silencing is a time expensive process and computational resources provide faster and cheaper options. Here, we identified the miRNAs target sequences on the mature PARP1 mRNA construct from the TargetScan database and obtained their complementary miRNA sequences from the miRbase. Using the human Ago-2 crystal structure deposited to the PDB database (PDB ID: 4W5Q) as a template, mutational studies were conducted on the protein/mRNA/miRNA complex to reproduce PARP1 mRNA seed pairing to miRNAs. Upon initial geometry optimization, further conformational search studies were done for thorough exploration of the conformational space of the protein/RNA complex by implementing conformational flooding (CF) molecular dynamics (MD) simulations. CF MD introduces a flooding potential to the force field, thus lowering energy barriers and facilitating observations of macroscale conformational changes. Clustering of the CF trajectories reveal the stable conformations of the Ago-2/miRNA/mRNA complexes. Energetically favorable miRNAs discovered in silico will further be validated with in vitro silencing assays.

Common Genetic Factors and Pathways in Alzheimer's Disease and Ischemic Stroke: Evidences from GWAS.

Alzheimer's disease (AD) and ischemic stroke (IS) are common neurological disorders, and the comorbidity of these two brain diseases is often seen. Although AD and IS were regarded as two distinct disease entities, in terms of different etiologies and clinical presentation, recent genome-wide association studies (GWASs) revealed that there were common risk genes between AD and IS, indicating common molecular pathways and their common pathophysiology. In this review, we summarize AD and IS risk single nucleotide polymorphisms (SNPs) and their representative genes from the GWAS Catalog database, and find thirteen common risk genes, but no common risk SNPs. Furthermore, the common molecular pathways associated with these risk gene products are summarized from the GeneCards database and clustered into inflammation and immunity, G protein-coupled receptor, and signal transduction. At least seven of these thirteen genes can be regulated by 23 microRNAs identified from the TargetScan database. Taken together, the imbalance of these molecular pathways may give rise to these two common brain disorders. This review sheds light on the pathogenesis of comorbidity of AD and IS, and provides molecular targets for disease prevention, manipulation, and brain health maintenance.