Dysregulated circRNAs Research Articles

Circular RNA hsa_circ_0044234 as distinct molecular signature of triple negative breast cancer: a potential regulator of GATA3

BackgroundCircular RNAs (circRNAs) have been implicated in the initiation and development of breast cancer as functional non-coding RNAs (ncRNA). The roles of circRNAs as the competing endogenous RNAs (ceRNAs) to sponge microRNAs (miRNAs) have also been indicated. However, the functions of circRNAs in breast cancer have not been totally elucidated. This study aimed to explore the clinical implications and possible roles of circ_0044234 in carcinogenesis of the most problematic BC subtype, triple negative breast cancer (TNBC), which are in desperate need of biomarkers and targeted therapies.MethodsThe importance of circ_0044234 as one of the most dysregulated circRNAs in TNBC was discovered through microarray expression profile analysis. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to confirm the downregulation of circ_0044234 in triple negative tumors and cell lines versus non-triple negative ones. The bioinformatics prediction revealed that circ_0044234 could act as an upstream sponge in the miR-135b/GATA3 axis, two of the most dysregulated transcripts in TNBC.ResultsOur experimental investigation of circ_0044234 expressions in various BC subtypes as well as cell lines reveals that TNBC expresses circ_0044234 at a substantially lower level than non-TNBC. The ROC curve analysis indicates that it could be applied as a discriminative biomarker to identify TNBC from other BC subtypes. Moreover, circ_0044234 expression could be an independent prognostic biomarker in BC. Interestingly, a substantial inverse expression correlation was detected between circ_0044234 and miR-135b-5p as well as between miR-135b-5p and GATA3 in breast tumors.ConclusionsThe possible clinical usefulness of circ_0044234 as a promising distinct biomarker and upcoming therapeutic target for TNBC have been indicated in this research. Our comprehensive approach revealed the potential circ_0044234/miR135b-5p/GATA3 ceRNA axis in TNBC.

CircRNA-ceRNA Network Revealing the Potential Regulatory Roles of CircRNA in Alzheimer's Disease Involved the cGMP-PKG Signal Pathway.

Background: Alzheimer’s disease (AD) is a chronic progressive neurodegenerative disease. The characteristic pathologies include extracellular senile plaques formed by β-amyloid protein deposition, neurofibrillary tangles formed by hyperphosphorylation of tau protein, and neuronal loss with glial cell hyperplasia. Circular RNAs (circRNAs) are rich in miRNA-binding sites (miRNA response elements, MREs), which serve as miRNA sponges or competitive endogenous RNAs (ceRNAs). Although several research groups have identified dysregulated circRNAs in the cerebral cortex of SAMP8 mice or APP/PS1 mice using deep RNA-seq analysis, we need to further explore circRNA expression patterns, targets, functions and the signaling pathways involved in the pathogenesis of AD and in particular the hippocampal circRNA expression profiles in AD.Methods: We used deep RNA sequencing to investigate circRNA-ceRNA network patterns in the hippocampus of APP/PS1 mice.Results: In our study, 70 dysregulated circRNAs, 39 dysregulated miRNAs and 121 dysregulated mRNAs were identified between the APP/PS1 group and the wild-type group at 8 months in the hippocampus of the mice. Through correlation analysis, we identified 11 dysregulated circRNAs, 7 dysregulated miRNAs and 8 dysregulated mRNAs forming 16 relationships in the circRNA-miRNA-mRNA regulatory network. Gene ontology (GO) analysis indicated that the dysregulated circRNAs were most enriched in biological metabolic processes. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the dysregulation of circRNAs was enriched in the cGMP-PKG signaling pathway, cAMP signaling pathway, Hippo signaling pathway, platelet activation, long-term potentiation and axon guidance. In addition, our findings preliminarily verified that the novel_circ_0003012/mmu-miR-298-3p/Smoc2 signaling axis may regulate the pathophysiology of AD by affecting the cGMP-PKG signaling pathway.Conclusions: These newly identified circRNAs in networks and signaling pathways reveal potential diagnostic or therapeutic targets for AD.

Involvement of circRNAs in Proinflammatory Cytokines-Mediated β-Cell Dysfunction.

Aim During the initial stage of type 1 diabetes, prolonged exposure of pancreatic β-cell to proinflammatory cytokines such as IL-1β, TNF-α, and IFN-γ results in a decreased capacity to produce and release insulin, as well as cell loss by apoptosis. Circular RNAs (circRNAs) are a new class of endogenous noncoding RNAs (ncRNAs) with closed loop with no free ends. circRNAs have been reported to be participated in the development of many diseases. As little is known about their role in insulin-secreting cells, this study is aimed at evaluating their contribution in the process of inflammation-induced β-cell damage. Methods circRNA expression profile of MIN6 cells stimulated with a mix of cytokines, including IL-1β, IFN-γ, and TNF-α, was detected by circRNA microarrays. Four dysregulated circRNAs were validated by qRT-PCR. The involvement of the selected circRNAs in β-cell dysfunction was tested after their inhibition in MIN6 cells. MicroRNA target prediction software and multiple bioinformatic approaches were used to predict the targeting genes of circRNAs and analyze possible functions of the circRNAs. Results 1020 upregulated and 902 downregulated circRNAs were identified in cytokines-treated β-cells. Inhibition of circRNAs 000286 and 017277 in β-cells could promote β-cell apoptosis and affect insulin biosynthesis and secretion. GO analysis enriched terms such as regulation of transcription and regulation of gene expression and KEGG analysis enriched top pathways included TGF-β and MAPK signaling pathways. Conclusions The data shows that circRNAs may be involved in proinflammatory cytokines-mediated β-cell dysfunction and suggests the involvement of circRNAs in the development of type 1 diabetes.

Identification of a Potentially Functional circRNA-miRNA-mRNA Regulatory Network in Melanocytes for Investigating Pathogenesis of Vitiligo.

CircRNAs have been reported to play essential roles in regulating immunity and inflammation, which may be an important regulatory factor in the development of vitiligo. However, the expression profile of circRNAs and their potential biological functions in vitiligo have not been reported so far. In our study we found there are 64 dysregulated circRNAs and 14 dysregulated miRNAs in the patients with vitiligo. Through the correlation analysis, we obtained 12 dysregulated circRNAs and 5 dysregulated miRNAs, forming 48 relationships in the circRNA-miRNA-mRNA regulatory network. Gene Ontology analysis indicated dysregulated circRNAs in vitiligo is closely related to the disorder of the metabolic pathway. The KEGG pathway of dysregulation of circRNAs mainly enriched in the biological processes such as ubiquitin mediated proteolysis, endocytosis and RNA degradation, and in Jak-STAT signaling pathway. Therefore, we found the circRNA-miRNA-mRNA regulatory network are involved in the regulation of numerous melanocyte functions, and these dysregulated circRNAs may closely related to the melanocyte metabolism. Our study provides a theoretical basis for studying the vitiligo pathogenesis from the perspective of circRNA-miRNA-mRNA network.

Circ-LAMP1 contributes to the growth and metastasis of cholangiocarcinoma via miR-556-5p and miR-567 mediated YY1 activation.

Dysregulation of circular RNAs (circRNAs) executes important regulatory roles in carcinogenesis. Nonetheless, few studies focused on the mechanisms of circRNAs in cholangiocarcinoma (CCA). qRT‐PCR was applied to verify the dysregulated circRNAs in CCA. Fisher's exact test, Kaplan‐Meier analysis and Cox regression model were utilized to investigate the clinical implications of circ‐LAMP1 in the patients with CCA. The viability, apoptosis, migration and invasion of CCA cells were detected after silencing/overexpression of circ‐LAMP1. Xenograft and lung metastasis assays were performed to verify the in vitro results. The regulatory networks of circ‐LAMP1 were unveiled by bioinformatic analysis, RNA immunoprecipitation (RIP), RNA pulldown and luciferase reporter assays. Up‐regulation of circ‐LAMP1 was found in CCA tissue samples and cell lines. Enhanced level of circ‐LAMP1 was linked to clinical severity, high post‐operative recurrence and poor prognosis for the patients with CCA. Gain/loss‐of‐function assays confirmed the oncogenic role of circ‐LAMP1 in mediating cell growth, apoptosis, migration and invasion. Nevertheless, the level of circ‐LAMP1 had no effect on normal biliary epithelium proliferation and apoptosis. Animal study further verified the in vitro data. Mechanistically, circ‐LAMP1 directly sponged miR‐556‐5p and miR‐567, thereby releasing their suppression on YY1 at post‐transcriptional level. Rescue assay indicated that the oncogenic role of circ‐LAMP1 is partially dependent on its modulation of YY1 in CCA. In summary, this study suggested that circ‐LAMP1 might be used as a promising biomarker/therapeutic target for CCA.

The identification of circular RNAs from peripheral blood mononuclear cells in systemic lupus erythematosus

BackgroundThe diagnosis of systemic lupus erythematosus (SLE) is complicated. This study explores the expression of circular RNAs (circRNAs), which are closed non-coding RNAs in which the 5′ and 3′ ends are covalently linked and which work by sponging microRNAs. CircRNAs were extracted from peripheral blood mononuclear cells (PBMCs) of SLE patients to identify novel circRNA species that might be used for SLE diagnosis.MethodsMicroarray was applied to screening circRNAs changes in PBMCs obtained from SLE patients (n = 10) and healthy participants (n = 10), paired for age and sex. We then verified the selected circRNAs in PBMCs using quantitative reverse transcription-polymerase chain reaction amplification (qRT-PCR) in another cohort, including ten paired SLE patients and healthy participants. The correlation between the differential circRNAs and clinical pathology of SLE were analyzed.Results182 up-regulated and 563 significantly down-regulated circRNAs in PBMCs of patients with SLE were identified. Besides, the qRT-PCR results were consistent with the microarray results. The correlation analysis revealed that has_circRNA_100236, has_circRNA_102489, and has_circRNA_101413 were correlated with positive anti-dsDNA, thrombocytopenia, and positive IgG, respectively. Lastly, their miRNAs targets and the binding sites were predicted.ConclusionWe identified some dysregulated circRNAs in PBMCs from SLE patients, and these circRNAs may be developed as the novel biomarkers for the diagnosis of SLE.

Circular RNA expression profiles and features in NAFLD mice: a study using RNA-seq data

BackgroundNonalcoholic fatty liver disease (NAFLD) is primarily characterized by the hepatic cholesterol accumulation. Circular RNA (circRNA), one of noncoding RNA, involves in many liver diseases progression. However, no recent studies on circRNA expression profiles in NAFLD have been reported previously.MethodsA NAFLD mouse model was constructed by providing high-fat diet (HFD) for 32 weeks. The circRNAs expression profile in normal mice and NAFLD mice were determined using high-output RNA sequencing method and bioinformatics methods, while the differentially expressed circRNAs were confirmed using Sanger sequencing and qRT-PCR. The circRNA-miRNA network was also predicted. The biological functions of circRNAs were annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG).ResultsThe results demonstrated the successful construction of NAFLD mice model by immunohistology and serology assay. In total, 93 dysregulated circRNAs were observed, including 57 upregulated circRNAs and 36 downregulated circRNAs, in the NAFLD group. The circRNA-miRNA network revealed the complex interaction between circRNAs and its potential miRNA targets in NAFLD. The characteristic of tissue-specific expression in circRNA was demonstrated. The differentially expressed circRNAs with important biological function were also annotated using GO and KEGG. Both DDAH1 and VAV3 genes were found to be associated with the NAFLD development.ConclusionsTaken together, this study demonstrated the circRNAs expression profile and features in NAFLD, which may provide potential biological markers for the pathogenesis of NAFLD.

Circ0120816 acts as an oncogene of esophageal squamous cell carcinoma by inhibiting miR-1305 and releasing TXNRD1

BackgroundCircular RNAs (circRNAs) have been discovered to participate in the carcinogenesis of multiple cancers. However, the role of circRNAs in esophageal squamous cell carcinoma (ESCC) progression is yet to be properly understood. This research aimed to investigate and understand the mechanism used by circRNAs to regulate ESCC progression.MethodsBioinformatics analysis was first performed to screen dysregulated circRNAs and differentially expressed genes in ESCC. The ESCC tissue samples and adjacent normal tissue samples utilized in this study were obtained from 36 ESCC patients. All the samples were subjected to qRT-PCR analysis to identify the expression of TXNRD1, circRNAs, and miR-1305. Luciferase reporter assay, RNA immunoprecipitation assay and RNA pull-down assay were later conducted to verify the existing relationship among circ0120816, miR-1305 and TXNRD1. CCK-8, BrdU, cell adhesion, cell cycle, western blot and caspase 3 activity assays were also employed to evaluate the regulation of these three biological molecules in ESCC carcinogenesis. To evaluate the effect of circ0120816 on ESCC tumor growth and metastasis, the xenograft mice model was constructed.ResultsExperimental investigations revealed that circ0120816 was the highest upregulated circRNA in ESCC tissues and that this non-coding RNA acted as a miR-1305 sponge in enhancing cell viability, cell proliferation, and cell adhesion as well as repressing cell apoptosis in ESCC cell lines. Moreover, miR-1305 was observed to exert a tumor-suppressive effect in ESCC cells by directly targeting and repressing TXNRD1. It was also noticed that TXNRD1 could regulate cyclin, cell adhesion molecule, and apoptosis-related proteins. Furthermore, silencing circ0120816 was found to repress ESCC tumor growth and metastasis in vivo.ConclusionsThis research confirmed that circ0120816 played an active role in promoting ESCC development by targeting miR-1305 and upregulating oncogene TXNRD1.

Protective effects of P2X7R antagonist in sepsis-induced acute lung injury in mice via regulation of circ_0001679 and circ_0001212 and downstream Pln, Cdh2, and Nprl3 expression.

Sepsis induces pulmonary P2X7 receptor (P2X7 R) expression and P2X7 R-knockout reduced lung inflammation in mice. The present study investigated the expression of circular RNA (circRNA) and mRNA in sepsis-induced acute lung injury (ALI) treated with a P2X7 R antagonist. Sepsis was induced by tracheal administration of lipopolysaccharide (LPS), and the mice were then divided into two groups: without [sepsis+dimethyl sulfoxide (DMSO)] or with P2X7 R antagonist treatment (sepsis+P2X7 A). Sham mice were administrated sterile normal saline. Serum levels of interleukin (IL)-1β and tumor necrosis factor (TNF)-α, pathological changes, cell apoptosis and P2X7 R expression in lung were assessed, followed by RNA sequencing (RNA-seq) and bioinformatics analyses. A quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) was used to validate circRNAs and mRNAs. Compared to the sham group, LPS-induced sepsis produced obvious pathological changes in lung tissue, as well as increased apoptotic lung cells, serum TNF-α and IL-1β levels, and P2X7 R expression; P2X7 R antagonism significantly ameliorated these changes. RNA-seq identified many dysregulated circRNAs and mRNAs during sepsis, whereas this changed with P2X7 R antagonism. RT-qPCR confirmed that Mus musculus (mmu)_circ_0001679, mmu_circ_0001212, phospholamban (Pln), cadherin-2 (Cdh2) and nitrogen permease regulator 3-like (Nprl3) expression were significantly increased in the sepsis+DMSO group compared to that in the sham group but were decreased in the sepsis+P2X7 A group compared to that in the sepsis+DMSO group. The circRNA-microRNA-mRNA coexpression network indicated that mmu_circ_0001679 may regulate Nprl3 and that mmu_circ_0001212 may similarly regulate Pln, Cdh2 and Nprl3 as a competing endogenous RNA. P2X7 R antagonism attenuates sepsis-induced ALI by inhibiting dysregulated expression of circRNA (circ_0001679, circ_0001212) and mRNA (Pln, Cdh2 and Nprl3).

Hsa_circNFXL1_009 modulates apoptosis, proliferation, migration, and potassium channel activation in pulmonary hypertension

In this study, we explored the circular RNA (circRNA) profile in pulmonary arterial hypertension (PAH) patients caused by chronic obstructive pulmonary disease (COPD) and the effects of hsa_circNFXL1_009 on abnormal proliferation, apoptosis, and migration of human pulmonary arterial smooth muscle cells (hPASMCs) driven by hypoxia. Using microarrays, we screened the circRNA profile in whole-blood samples from three pairs of subjects and found 158 dysregulated circRNAs in patients with PAH-COPD. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) analysis further validated that hsa_circNFXL1_009 was dramatically downregulated with the highest area under a receiver operating characteristic curve (ROC) in 21 pairs of subjects. Consistently, exposure to hypoxia markedly reduced the hsa_circNFXL1_009 level in cultured hPASMCs. Delivery of exogenous hsa_circNFXL1_009 attenuated hypoxia-induced proliferation, apoptotic resistance, and migration of hPASMCs, as evidenced by immunocytochemistry, 5-ethynyl-2′-deoxyuridine incorporation, wound healing, and a TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling) assay. A luciferase assay showed that hsa_circNFXL1_009 directly sponged hsa-miR-29b-2-5p (miR-29b) and positively regulated the expression of voltage-gated potassium (K+) channel subfamily B member 1 (KCNB1) at the mRNA level. Using patch-clamp electrophysiology, we proved that overexpression of hsa_circNFXL1_009 promoted a whole-cell K+ current in hPASMCs. Taken together, these studies identify hsa_circNFXL1_009 as a key regulator of PAH, and it may be used as a potential therapeutic target for the treatment of PAH.

Identification of differentially expressed circular RNAs in human nasopharyngeal carcinoma.

Circular RNAs (circRNAs) are endogenous RNAs that have a covalent closed cycle configuration. circRNAs have been found to be differentially expressed in many human cancers. Therefore, circRNAs may be ideal biomarkers for the diagnosis and treatment of cancer. However, we know very little about the function of circRNAs in nasopharyngeal carcinoma (NPC). The purpose of this study was to evaluate the circRNA expression profiles in NPC. We utilized high-throughput RNA sequencing (RNA-Seq) to evaluate the circRNA expression profile in NPC A total of 13,561 unique circRNA candidates were detected. Selection of aberrantly expressed circRNAs was carried out using a q-value of < 0.001 with a fold change of > 2.0 or < 0.5. We carried out Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses to identify the biological functions of differentially expressed circRNAs. Moreover, bioinformatics analyses were implemented to predict the effects between circRNAs and cancer-related microRNAs (miRNAs), and we used Cytoscape to build a cancer-related circRNA-miRNA target gene map. Finally, to verify dysregulated circRNAs, quantitative real-time PCR was utilized. In NPC tissues, we found that 73 circRNAs were downregulated and 59 were upregulated. The top 12 candidate circRNAs were selected from several vital NPC pathways such as the human papillomavirus and Epstein-Barr virus infection signaling pathways (hsa05165 and hsa05169, respectively), Hepatitis B (hsa05161), and the Ras signaling pathway (hsa04014). A network map of circRNA-miRNA interactions of 12 differentially expressed circRNAs was built. Hsa_circ_0007637 expression distinguished NPC tissues from paired healthy tissues and NPC cell lines (HNE1 6-10B, 5-8F, CNE-2, and so on) from a normal epithelial (NP460) cell line. In this study, we investigated the profiles of differentially expressed circRNAs in NPC, and our results show that hsa_circ_0007637 may be a biomarker for NPC and play a role in its development. This observation-based research identified dysregulated circRNAs in NPC, which may assist in the development of biomarkers for this disease. Further studies on the mechanisms and functions of these circRNAs may promote our understanding of NPC tumorigenesis.

CircRNAome Profiling in Oral Carcinoma Unveils a Novel circFLNB that Mediates Tumour Growth-Regulating Transcriptional Response

Deep sequencing technologies have revealed the once uncharted non-coding transcriptome of circular RNAs (circRNAs). Despite the lack of protein-coding potential, these unorthodox yet highly stable RNA species are known to act as critical gene regulatory hubs, particularly in malignancies. However, their mechanistic implications in tumor outcome and translational potential have not been fully resolved. Using RNA-seq data, we profiled the circRNAomes of tumor specimens derived from oral squamous cell carcinoma (OSCC), which is a prevalently diagnosed cancer with a persistently low survival rate. We further catalogued dysregulated circRNAs in connection with tumorigenic progression. Using comprehensive bioinformatics analyses focused on co-expression maps and miRNA-interaction networks, we delineated the regulatory networks that are centered on circRNAs. Interestingly, we identified a tumor-associated, pro-tumorigenic circRNA, named circFLNB, that was implicated in maintaining several tumor-associated phenotypes in vitro and in vivo. Correspondingly, transcriptome profiling of circFLNB-knockdown cells showed alterations in tumor-related genes. Integrated in silico analyses further deciphered the circFLNB-targeted gene network. Together, our current study demarcates the OSCC-associated circRNAome, and unveils a novel circRNA circuit with functional implication in OSCC progression. These systems-based findings broaden mechanistic understanding of oral malignancies and raise new prospects for translational medicine.

Comprehensive analysis of circular RNA expression profiles in cisplatin-resistant non-small cell lung cancer cell lines.

Platinum-based drugs such as cisplatin are widely used in combination chemotherapy for non-small cell lung cancer (NSCLC) owing to their high clinical response rate; however, acquired resistance to cisplatin is eventually inevitable. Circular RNAs (circRNAs) are involved in the development of diverse types of cancers, but their connection to cisplatin-resistance in NSCLC has not been studied. In the present study, two cisplatin-resistant NSCLC cell lines (A549/DDP and PC9/DDP) were established by gradually increasing concentrations of cisplatin in the media. The resulting cell lines possessed high resistance to cisplatin and strong proliferation, migration, and colony formation abilities compared to the parental cells. Microarray analysis identified 19,161 circRNAs that were dysregulated in cisplatin-resistant cell lines (fold change abs>2), including 11,915 up-regulated and 7246 down-regulated circRNAs. The expression of the top five up-regulated and down-regulated circRNAs was validated using real-time quantitative polymerase chain reaction. A circRNA-micro RNA (miRNA) network of the top 20 dysregulated circRNAs and their predicted miRNAs was constructed using Cytoscape. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that the host genes of the identified circRNAs were involved in the regulation of MAP kinase kinase kinase kinase activity, 6-phosphofructo-2-kinase activity, focal adhesion, ErbB signaling, and ECM-receptor interactions, which may contribute to cisplatin resistance in NSCLC. In summary, this is the first report on circRNA profiling in cisplatin-resistant NSCLC cells and it provides new potential targets for the reversal of cisplatin resistance in NSCLC.

RNA-Seq Profiling of Serum Exosomal Circular RNAs Reveals Circ-PNN as a Potential Biomarker for Human Colorectal Cancer.

Circular RNAs (circRNAs) are emerging as cardinal areas of focus in the non-coding RNA field. Growing evidences have revealed exosomal circRNAs as potential biomarkers for detection of various cancers. However, the clinical importance of most serum exosomal circRNAs in colorectal cancer (CRC) have rarely been investigated. In this study, we examined the possible clinical application of serum exosomal circRNAs in the diagnosis of CRC. Firstly, we conducted RNA sequencing (RNA-seq) analysis using fifty CRC and fifty healthy control serum samples to identify CRC-related circRNAs. The sequencing data showed 122 differentially expressed circRNAs including 100 up-regulated and 22 down-regulated circRNA transcripts in CRC patients. Then, eight most dysregulated circRNAs were selected for validation by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assay. Validation analysis revealed that the serum exosomal circ-PNN (hsa_circ_0101802) levels were significantly up-regulated in CRC cases compared with those in the healthy control groups. Receiver operating characteristic curve (ROC) analysis suggested that circ-PNN had significant value in CRC diagnosis with areas under the ROC curve (AUC) of 0.855 and 0.826 in the training and validation sets, respectively. We also found that the AUC of serum exosomal circ-PNN for early-stage CRC was 0.854. Finally, a network map based on circ-PNN was constructed to determine its potential miRNA-mRNAs binding. We also demonstrated that the expression of hsa-miR-6833-3P, hsa-let-7i-3p and hsa-miR-1301-3P were negatively correlated with circ-PNN in CRC patients. Collectively, our findings indicated that serum exosomal circ-PNN might be a potential non-invasive biomarker for the detection of CRC and may play a crucial role in the pathogenesis of CRC.

Circular RNA expression and association with the clinicopathological characteristics in papillary thyroid carcinoma.

Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer. Circular RNAs (circRNAs) are a novel class of RNAs, with higher stability and tissue specificity, which may be of value as novel clinical markers. High-throughput RNA sequencing was used to profile the expression of circRNAs in 5 pairs of cancer and normal tissues, and reverse transcription-quantitative PCR (RT-qPCR) analysis was employed to verify the results of the RNA sequencing in 45 cases of PTC. The dysregulated circRNA expression and clinicopathological characteristics were assessed and the potential roles of circRNAs in the cellular miRNA and mRNA network were predicted using bioinformatics analysis. The results demonstrated that, compared with normal tissues, a total of 53 circRNAs were dysregulated in tumour tissues, and 8 circRNAs were validated at the mRNA level (P<0.001 and P<0.01). Among those, the expression of chr5:161330882-161336769- (P=0.015), chr9:22046750-22097364+ (P=0.041) and chr8:18765448-18804898- (P=0.036) were obviously associated with the BRAFV600E mutation, chr12:129699809-129700698- was associated with capsular invasion (P=0.025) and chr5:38523418-38530666- was associated with pT stage (P=0.037) and lymph node metastasis (P=0.002). Therefore, some dysregulated circRNAs were found to be associated with BRAFV600E mutation, capsular invasion, advanced pT stage and lymph node metastasis of PTC, indicating that circRNAs may be involved in tumourigenesis and cancer progression, and they may be putative biomarkers for the diagnosis and evaluation of progression of PTC.

Identification of downregulated circRNAs from tissue and plasma of patients with gastric cancer and construction of a circRNA-miRNA-mRNA network.

The connection between circular RNAs (circRNAs) and gastric cancer has been reported widely in recent years. However, previous studies have focused mainly on circRNAs from gastric cancer tissue. The objectives of the present study were to detect dysregulated circRNAs from both tissue and plasma of patients with gastric cancer and to explore their potential roles in the pathogenesis of gastric cancer. Expression profiles of circRNAs were obtained from the Gene Expression Omnibus (GEO) and analyzed using the GEO2R tool to identify differential expressed circRNAs. The significance threshold was set as |log2 (fold change)| > 2 and adjusted P < .05. The microRNA (miRNA) binding sites of the differentially expressed circRNAs were predicted using the Circular RNA Interactome web tool. TargetScan and the miRNet database were used to predict the miRNA target genes. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed using Database for Annotation Visualization and Integrated Discovery. Hub genes were identified and a network was constructed with Cytoscape. The overall survival rates for the selected miRNAs and messenger RNAs were evaluated by Kaplan-Meier Plotter. A total of three downregulated circRNAs (hsa_circ_0001190, hsa_circ_0036287, and hsa_circ_0048607) were identified in this study. Six miRNAs and eight hub genes met the significance criteria and were selected for further analysis. A circRNA-miRNA-hub gene network was constructed based on three circRNAs, six miRNAs, and eight hub genes. Evaluation of overall survival rates for the hub genes showed that low expression levels of GADD45A, PPP1CB, PJA2, and KLF2 were associated with poor overall survival. This study identified potential novel plasma circRNA biomarkers and provides insights into the underlying mechanisms of gastric cancer pathogenesis.

Comprehensive circular RNA profiling reveals the regulatory role of circ_100242/miR-145 pathway in bladder cancer.

Bladder cancer is a common genitourinary carcinoma with high morbidity and mortality rates. Circular RNAs (circRNAs), which are a type of single-stranded RNAs, have been characterized as stable, conserved and tissue-specific molecules in mammalian cells. The present study explored the circRNA expression profile in four bladder cancer tissues and matched normal samples by using microarray analysis. Furthermore, bioinformatics analyses were performed to investigate the potential function of dysregulated circRNAs in bladder cancer. The results demonstrated that 89 circRNAs were downregulated and 210 circRNAs were upregulated in bladder cancer tissues. The results from RT-qPCR demonstrated that hsa_circ_100241, hsa_circ_100242 and hsa_circ_101303 were markedly upregulated whereas hsa_circ_104510 was significantly downregulated in bladder cancer tissues compared with normal tissue. Following circRNA/microRNA (miRNA) interaction network generation via Cytoscape, it was demonstrated that hsa_circ_100242 contained a miRNA response element for miR-145-5p, which is a tumor suppressor in bladder carcinoma. In addition, results from the Kyoto Encyclopedia of Genes and Genomes analysis showed that the MAPK signaling pathway was the most significant pathway of the differentially expressed circRNAs in bladder cancer tissues. In conclusion, the present study demonstrated that circRNAs were dysregulated in bladder cancer tissues compared with matched normal samples. Pathway analysis was also performed to predict the binding of miRNAs to the dysregulated circRNAs. The results revealed that hsa_circ_100242 may be involved in bladder cancer initiation and progression by sponging miR-145. These findings may provide further insights into the functional and therapeutic roles of circRNAs in bladder cancer.

Circular RNA expression profile and its potential regulative role in human abdominal aortic aneurysm

BackgroundThis study aimed to identify the differentially expressed circular RNAs (circRNAs) between human abdominal aortic aneurysm (AAA) and the control group.MethodsHigh-throughput sequencing was applied to determine the circRNA expression profiles of 4 paired aortic samples. Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was carried out to testify 6 randomly selected dysregulated circRNAs. Kyoto Encyclopedia of Genes and Genomes and Gene ontology (GO) analysis were conducted for functional annotation of the parental genes. Additionally, interaction networks between circRNA and 5 putative microRNA (miRNA) partners were constructed.ResultsFinally, 411 differentially expressed circRNAs were discovered, including 266 downregulated and 145 upregulated circRNAs. Compared with the control group, the expression level of hsa (Homo sapiens) _circ_0005360 (LDLR) and hsa_circ_0002168 (TMEM189) were proved significantly lower in the AAA group by qRT-PCR. Regarding upregulated circRNAs, the most enriched GO molecular function, biological process and cellular component terms were poly(A) RNA binding, negative regulation of transcription from RNA polymerase II promoter and nucleoplasm, respectively. Moreover, circRNA/miRNA interaction networks showed that hsa_circ_0005360/miR-181b and hsa_circ_0002168/miR-15a axis might have a regulative role in human AAA.ConclusionsThis study revealed new circRNAs potentially related to the pathogenesis of AAA. Further experimental studies are warranted to clarify the potential molecular mechanisms.

Circular RNAs as biomarkers in liquid biopsy in colorectal cancer.

230 Background: Circular RNAs (circRNAs) are emerging as essential regulators of cancer- related biological hallmarks, as cell proliferation, apoptosis, differentiation, immune regulation and angiogenesis. CircRNAs are abundant, conserved, and have a tissue‐specific expression pattern. These characteristics make them candidate to serve as biomarkers in liquid biopsy (LB) in cancer. The aim of this study is to analyse differential expression of circRNAs in the colorectal cancer (CRC) scenario. Methods: To comprehensively understand the expression patterns of circRNAs we characterized 13,617 circRNAs using a microarray [Arraystar v2 (8x15K)] in 10 human samples, five CRC cell lines, one colorectal human tumour, one normal colon healthy control, vs. Peripheral Human Blood Leukocytes (2 pools) and Human Bone Marrow. Differentially expressed circRNAs were identified using fold change (FC) cut-off or through Volcano Plot filtering respectively. CircRNAs having FC ▪2 and P-values ▪ 0.05 were selected. CircRNA/microRNA interaction was predicted with target prediction software. Results: Hierarchical clustering showed distinguishable circRNA expression profiling among 10 samples. These data indicated that circRNAs have a different expression pattern in colorectal tissues compared with that in blood and bone marrow tissues. The microarray data showed 2329 circRNAs differentially expressed (FC &gt; 2.0, P &lt; 0.05). Among them, 964 circRNAs were upregulated and 1365 were downregulated in colon tissues compared with blood and bone marrow. Using a stringent criterion (FC &gt; 10, P≤ 0.01 and false discovery rate [FDR] &lt; 0.05) we have identified 30 circRNA upregulated in colorectal cancer versus non tumour samples. CircRNA/microRNA interaction prediction analysis showed that most upregulated circRNAs contain miRNA Binding Sites (MREs) for cancer-related miRNA, including among others, miR-17, miR-103, miR-let-7g. Conclusions: Microarray analysis was used to comprehensively identify dysregulated circRNAs in CRC. We identify novel circRNAs could be valuable as blood-based CRC biomarkers.

Circular RNA hsa_circ_001895 serves as a sponge of microRNA-296-5p to promote clear cell renalcell carcinoma progression by regulating SOX12.

There is an urgent need to find novel potential therapeutic targets for the diagnosis and treatment of clear cell renal cell carcinoma (ccRCC) due to its highly invasive ability as a common urological malignant tumor. Circular RNAs (circRNAs) have been indicated as potentially critical mediators in various types of tumor progression. We first used qRT‐PCR analysis to find dysregulated circRNAs in ccRCC. A novel circRNA, hsa_circ_001895, was upregulated in ccRCC specimens and associated with metastatic properties of ccRCC. However, the tumorigenic mechanism of hsa_circ_001895 on ccRCC is yet to be found. We first indicated that hsa_circ_001895 predicted a poor prognosis in ccRCC patients. Additionally, overexpression of hsa_circ_001895 not only promoted cell proliferation, invasion and migration of ccRCC, but also inhibited cell apoptosis, whereas hsa_circ_001895 knockdown reversed the effect on ccRCC progression. In vivo s.c. xenotransplanted tumor model also showed that silencing hsa_circ_001895 could suppress in vivo ccRCC growth. Mechanistically, hsa_circ_001895 directly binds with microRNA (miR)‐296‐5p and inhibits its expression. Moreover, sex determining region Y (SRY)‐box 12 (SOX12) was identified as a target of miR‐296‐5p, the expression of which was suppressed by miR‐296‐5p. Notably, the inhibitory effect of hsa_circ_001895 on ccRCC progression was reversed by miR‐296‐5p inhibitor. In general, our findings indicated that hsa_circ_001895 may sponge miR‐296‐5p and promote SOX12 expression, which is the underlying mechanism of hsa_circ_001895‐induced ccRCC progression.