microRNA Target Genes Research Articles

Network Analysis Integrating microRNA Expression Profiling with MRI Biomarkers and Clinical Data for Prostate Cancer Early Detection: A Proof of Concept Study.

The MRI of the prostate is the gold standard for the detection of clinically significant prostate cancer (csPCa). Nonetheless, MRI still misses around 11% of clinically significant disease. The aim was to comprehensively integrate tissue and circulating microRNA profiling, MRI biomarkers and clinical data to implement PCa early detection. In this prospective cohort study, 76 biopsy naïve patients underwent MRI and MRI directed biopsy. A sentinel sample of 15 patients was selected for a pilot molecular analysis. Weighted gene coexpression network analysis was applied to identify the microRNAs drivers of csPCa. MicroRNA–target gene interaction maps were constructed, and enrichment analysis performed. The ANOVA on ranks test and ROC analysis were performed for statistics. Disease status was associated with the underexpression of the miRNA profiled; a correlation was found with ADC (r = −0.51, p = 0.02) and normalized ADC values (r = −0.64, p = 0.002). The overexpression of miRNAs from plasma was associated with csPCa (r = 0.72; p = 0.02), and with PI-RADS assessment score (r = 0.73; p = 0.02); a linear correlation was found with biomarkers of diffusion and perfusion. Among the 800 profiled microRNA, eleven were identified as correlating with PCa, among which miR-548a-3p, miR-138-5p and miR-520d-3p were confirmed using the RT-qPCR approach on an additional cohort of ten subjects. ROC analysis showed an accuracy of >90%. Provided an additional validation set of the identified miRNAs on a larger cohort, we propose a diagnostic paradigm shift that sees molecular data and MRI biomarkers as the prebiopsy triage of patients at risk for PCa. This approach will allow for accurate patient allocation to biopsy, and for stratification into risk group categories, reducing overdiagnosis and overtreatment.

β-arrestin1 Promotes the Concentration of Mitochondrial Reactive Oxygen in T-ALL Cells via MiR-652-5p

To investigate the effect of β-arrestin1 on the concentration of reactive oxygen species (ROS) in the mitochondria of acute T-lymphocytic leukemia (T-ALL) cells and its possible mechanisms. The stable T-ALL cell line with knocked down β-arrestin1 (Jurkat Siβ1) was constructed. Flow cytometry and probe assays were used to detect ROS content in cell and mitochondrial, respectively. The relationship between β-arrestin1 and microRNA was detected, analyzed and Q-PCR confirmed by microRNA microarray. The target genes of microRNA were predicated by miRbase software, identified by Western blot, and validated by Dual luciferase reporter gene. Jurkat Siβ1 stable cell line was successfully constructed and it was found that ROS content was slightly reduced in Jurkat Siβ1 at the whole cell level, and the ROS content was also significantly reduced in mitochondria. MicroRNA microarray analysis revealed that multiple T-ALL related microRNAs showed differentially expressed, in which the expression of miR-652-5p was significantly increased in Jurkat Siβ1 (P<0.05 fold>2.0), and Q-PCR showed that miR-652-5p was nearly 5-fold up-regulated in Jurkat Siβ1. miRbase predicted that the P62 gene was the target gene of miR-652-5p which could regulates mitochondrial function. P62 protein showed highly expressed in stably knocked down miR-652-5p in Jurkat cells. Dual luciferase reporter gene assay confirms that P62 was the target gene of miR-652-5p. β-arrestin1 can decreases the expression of miR-652-5p and deregulates the translational inhibition of P62 mRNA, thus to increase ROS content in mitochondria of T-ALL cells.

Uncovering miRNA-mRNA Regulatory Modules in Developing Xylem of Pinus massoniana via Small RNA and Degradome Sequencing.

Xylem is required for the growth and development of higher plants to provide water and mineral elements. The thickening of the xylem secondary cell wall (SCW) not only improves plant survival, but also provides raw materials for industrial production. Numerous studies have found that transcription factors and non-coding RNAs regulate the process of SCW thickening. Pinus massoniana is an important woody tree species in China and is widely used to produce materials for construction, furniture, and packaging. However, the target genes of microRNAs (miRNAs) in the developing xylem of P. massoniana are not known. In this study, a total of 25 conserved miRNAs and 173 novel miRNAs were identified via small RNA sequencing, and 58 differentially expressed miRNAs were identified between the developing xylem (PM_X) and protoplasts isolated from the developing xylem (PM_XP); 26 of these miRNAs were significantly up-regulated in PM_XP compared with PM_X, and 32 were significantly down-regulated. A total of 153 target genes of 20 conserved miRNAs and 712 target genes of 113 novel miRNAs were verified by degradome sequencing. There may be conserved miRNA-mRNA modules (miRNA-MYB, miRNA-ARF, and miRNA-LAC) involved in softwood and hardwood formation. The results of qRT-PCR-based parallel validation were in relatively high agreement. This study explored the potential regulatory network of miRNAs in the developing xylem of P. massoniana and provides new insights into wood formation in coniferous species.

MicroRNA-206 relieves irradiation-induced neuroinflammation by regulating connexin 43.

Radiation therapy has been widely used for the treatment of various types of cancer; however, it may cause neuroinflammation during the pathological process of the disease. Astrocytes, the most abundant cell type in the central nervous system, have been confirmed to play vital roles in various diseases. Connexin (Cx)43, the main Cx type in astrocytes, which has been identified as a direct target gene of microRNA (miR)-206, was found to be involved in diseases pathologies in regions with astrocytes. The aim of the present study was to investigate the mechanism through which γ-radiation may cause astrocyte neuroinflammation and determine the specific mechanism underlying the effects of miR-206 in irradiation-induced HA-1800 cells. A dual-luciferase reporter system was used to predict and verify the target binding site between Cx43 and miR-206. HA-1800 cell viability and apoptosis were determined using a MTT assay and flow cytometry, respectively. In addition, the HA-1800 cells were induced by γ-radiation, then the protein and mRNA expression levels of Cx43, miR-206 and cleaved-caspase-3 were determined using western blot and reverse transcription-quantitative PCR analyses, respectively. ELISA was also performed to evaluate the concentrations of different inflammatory cytokines (TNF-α, IL-β, IL-6 and IFN-γ). The dual-luciferase reporter system indicated that Cx43 was a direct target of miR-206. miR-206 mimics increased the expression level of miR-206 in the astrocytes. Irradiation suppressed cell proliferation, increased apoptotic cells and enhanced cleaved-caspase-3 expression and inflammatory cytokines secretion in astrocytes. Furthermore, miR-206 was found to be downregulated and its expression was inversely associated with that of Cx43 in γ-radiation-induced astrocytes. Overexpression of miR-206 enhanced miR-206 and suppressed Cx43 expression, while Cx43 was upregulated in HA-1800 cells transfected with miR-206 mimic + Cx43-plasmid. However, the expression level of miR-206 was not significantly different in the Cx43-plasmid transfected group. In addition, it was found that miR-206 mimics relieved irradiation-induced neuroinflammation, which was confirmed by increased cell viability, and reduced cell apoptosis and cleaved caspase-3 protein expression, as well as decreased inflammatory cytokine secretion. Furthermore, all the effects of miR-206 mimics on γ-radiation-induced astrocytes were reversed by Cx43-plasmid. In summary, the results of the present study indicated that miR-206 may relieve irradiation-induced neural damage by regulating Cx43, which may provide a novel research direction and a potential therapeutic target for the clinical treatment of inflammation-associated neuronal injury following irradiation.

Comprehensive computational analysis reveals human respiratory syncytial virus encoded microRNA and host specific target genes associated with antiviral immune responses and protein binding

ObjectiveAbout half-century ago, human respiratory syncytial virus (hRSV) was observed in infants and children under age of five years. As the mechanism of its pathogenesis inside the host is still lacking, in this in-silico study we hypothesized that RSV might create miRNAs, which could target the genes associated with host cellular regulatory pathways, thus provide persistent refuge to virus. MethodologyPre-miRNAs in RSV genome (accession no. NC_001803.1) were extracted through VMir software, and the identification of putative pre-miRNAs and mature miRNAs was accessed using iMiRNA-SSF and FOMmiR tool, respectively. Later, prediction of host specific target gene was accompanied by RNAhybrid tool. Moreover, bioinformatics analysis was done for the validation of target genes using microarray dataset (GSE80179). Lastly, Drug-gene interaction database was used to explore the small drug like candidate against RSV infection. ResultsSearching RSV genome for their pre-miRNAs yielded 15 pre-miRNAs like sequence with length vary for each pre-miRNA sequences. FOMmiR tool revealed a total of 7 mature miRNAs from 6 real pre-miRNA hairpins. Functional enrichment analysis of targeted genes, revealed their involvement in many biological pathways which facilitate their pathogenesis in host. The microarray dataset (GSE80179) was analyzed to validate altered expression level of target genes and found genes linked with pathways such as T-cell activation, apoptosis, NF-kappa B signaling, cell differentiation and autophagy. CYCS, CTLA4, and BTK were chosen as possible targets of 21 drugs. A total of 21 drugs were explored using DGIdb that might have potential to treat RSV patient. ConclusionThis study updates the information and yield a new perspective in context of understanding the pathogenesis of RSV. Our study presents novel miRNAs and their targeted hub genes, which upon experimental validation could facilitate in developing new therapeutics against RSV infection. In vivo and in vitro investigation of miRNAs and pathway interaction is essential to delineate the specific roles of the novel miRNAs, which may help to reveal the mechanisms behind the pathogenesis. Based on the hub genes and miRNAs, experimental models may be designed in terms of the detection of pathogenesis, evaluation of risk, and determining the targeted therapies of RSV infections.

Silver birch pollen-derived microRNAs promote NF-κB-mediated inflammation in human lung cells

The pollen of Betula pendula Roth (silver birch) is considered to be the main cause of allergy-related rhinitis in Europe and its protein-based allergens such as Bet v 1 are well characterized. However, little is known about non-protein components of birch pollen, e.g., small RNAs and their proinflammatory activity. In the present study, next-generation sequencing (NGS) and bioinformatic approaches were used for silver birch pollen (SBP)-derived microRNA profiling and evaluation of microRNA target genes and pathways in human. Human lung cells, namely WI-38 fibroblasts and A549 alveolar epithelial cells were then stimulated with SBP microRNA in vitro and imaging cytometry-based analysis of the levels of proinflammatory cytokines, autophagy parameters and small RNA processing regulators was conducted. Bioinformatic analysis revealed that SBP microRNA may interfere with autophagy, inflammation and allergy pathways in human. SBP and SBP-derived microRNA induced NF-κB-mediated proinflammatory response in human lung cells as judged by increased levels of NF-κB p65, IL-8 and TNFα. NSUN2 and NSUN5 were involved in pollen-derived microRNA processing. Pollen-derived microRNA also modulated autophagic pathway by changes in the pools of LC3B and p62 that may affect autophagy-based adaptive responses during allergic lung inflammation. We postulate that SBP-derived microRNAs can be considered as novel proinflammatory environmental agents.

MicroRNAs Involved in the Therapeutic Functions of Noni (Morinda citrifolia L.) Fruit Juice in the Treatment of Acute Gouty Arthritis in Mice Induced with Monosodium Urate.

We investigated the functions of microRNAs in the therapeutic effects of noni (Morinda citrifolia L.) fruit juice on mouse models of acute gouty arthritis induced with monosodium urate (MSU). Compared with the model group (treated with MSU), mice in both the positive control group (treated with both MSU and colchicine) and noni fruit juice group (treated with MSU and noni fruit juice) showed a significantly decreased degree of paw swelling in 5 days, as well as the contents of two types of proinflammatory cytokines (i.e., NALP3 and TNF-α). Based on the next-generation sequencing technology, a total of 3896 microRNAs (234 known and 3662 novel) were identified in mice treated with noni fruit juice. A large amount of differentially expressed miRNAs were identified in the noni fruit juice group, suggesting the significant effects of noni fruit juice on the mice with acute gouty arthritis, while the different patterns of change in the numbers of both upregulated and downregulated miRNAs in both noni fruit juice and positive control groups indicated that the mice of acute gouty arthritis may be regulated by differential mechanisms between the treatments of noni fruit juice and colchicine. The target genes of microRNAs involved in the pathogenesis and pathology of acute gouty arthritis in mice were identified and further annotated by both Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Our results revealed the therapeutic effects of noni fruit juice on acute gouty arthritis in mice with a group of microRNAs involved in the pharmacological mechanisms of noni fruit juice, providing scientific evidence to support both the agricultural cultivation and pharmacological significance of noni plants.

Human Prostate Tissue MicroRNAs and Their Predicted Target Pathways Linked to Prostate Cancer Risk Factors.

Simple SummaryProstate cancer is a major medical issue in men worldwide. In 2018, prostate cancer was the most frequently diagnosed cancer among men in developed countries. MicroRNAs seem to be important regulators of prostate cancer, but better understanding of their role in this disease is still needed to develop novel diagnostic tools and treatments. In this study, we aim to achieve insight on the microRNA profile of prostatic tissue environment in men with prostate cancer and to correlate this microRNA expression profile with risk factors of prostate cancer. We also examined the effects of cholesterol-lowering atorvastatin on the prostatic tissue microRNA profile. Our results provide new evidence from the effects of prostate cancer-related factors on the expression of prostate tissue microRNAs. Notably, this can reveal new pathways that may link risk factors to prostate cancer and pinpoint new therapeutic possibilities.MicroRNAs are important in prostate cancer development, progression and metastasis. The aim of this study was to test microRNA expression profile in prostate tissue obtained from prostate cancer patients for associations with various prostate cancer related factors and to pinpoint the predicted target pathways for these microRNAs. Prostate tissue samples were obtained at prostatectomy from patients participating in a trial evaluating impact of pre-operative atorvastatin on serum prostate specific antigen (PSA) and Ki-67 expression in prostate tissue. Prostate tissue microRNA expression profiles were analyzed using OpenArray® MicroRNA Panel. Pathway enrichment analyses were conducted for predicted target genes of microRNAs that correlated significantly with studied factors. Eight microRNAs correlated significantly with studied factors of patients after Bonferroni multiple testing correction. MiR-485-3p correlated with serum HDL-cholesterol levels. In atorvastatin-treated subjects, miR-34c-5p correlated with a change in serum PSA and miR-138-3p with a change in total cholesterol. In the placebo arm, both miR-576-3p and miR-550-3p correlated with HDL-cholesterol and miR-627 with PSA. In pathway analysis, these eight microRNAs related significantly to several pathways relevant to prostate cancer. This study brings new evidence from the expression of prostate tissue microRNAs and related pathways that may link risk factors to prostate cancer and pinpoint new therapeutic possibilities.

Long noncoding RNA LINC01287 promotes proliferation and inhibits apoptosis of lung adenocarcinoma cells via the miR‐3529‐5p/RNASEH2A axis under the competitive endogenous RNA pattern

Lung adenocarcinoma (LUAD) is regarded as the most common type of lung cancer. The molecular targeted therapies for LUAD have being extensively studied. Ribonuclease H2 subunit A (RNASEH2A) is a nucleotide degrading enzyme gene that exerts great influence on cell proliferation, DNA replication and genomic stability. According to bioinformatics analysis, RNASEH2A expression in LUAD tissues is predicted to be upregulated and high expression of RNASEH2A might be related to lower survival rate in LUAD patients. In the present study, we investigated functions of RNASEH2A in LUAD. The mRNA RNASEH2A showed high expression in LUAD cells, and its knockdown inhibited proliferation and induced apoptosis in LUAD cells. RNASEH2A was found to be a target gene of microRNA miR-3529-5p after their expression levels and interaction being examined. Long noncoding RNA LINC01287 upregulated RNASEH2A expression in LUAD cells by combining with miR-3529-5p in a competitive way. Rescue assays revealed that the overexpression of RNASEH2A reversed the suppression of cell proliferation and the promotion of cell apoptosis induced by miR-3529-5p overexpression or LINC01287 knockdown. Finally, forkhead box A1 (FOXA1) interacted with RNASEH2A promoter and LINC01287 promoter to upregulate the expression levels of RNASEH2A and LINC01287 in LUAD cells. Overall, FOXA1-induced LINC01287 serves as a competing endogenous RNA to promote proliferation and inhibit apoptosis of LUAD cells via upregulation of RNASEH2A expression at the posttranscriptional level by competitively combining with miR-3529-5p.

Knockdown of Myeloid Cell Leukemia-1 by MicroRNA-101 Increases Sensitivity of A549 Lung Cancer Cells to Etoposide.

Background: Studies have shown that myeloid cell leukemia-1 (Mcl-1) is the target gene for microRNA -101 (miRNA-101), and decreased levels of miRNA-101 are associated with elevated levels of Mcl-1 and lung cancer survival. The objective of the present study was to investigate the effect of miRNA-101 on the sensitivity of A549 lung cancer cells to etoposide. Methods: The study was conducted during 2018 and 2019 at Arak University of Medical Sciences, Arak, Iran. The effect of miRNA-101 on Mcl-1 expression was assessed using reverse transcription-quantitative polymerase chain reaction 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), and trypan blue exclusion assays were performed to determine the effect of treatments on cell survival and proliferation, respectively. The interaction between miRNA-101 and etoposide was evaluated using the combination index analysis of Chou-Talalay. Apoptosis was quantified using ELISA cell death assay. ANOVA and Bonferroni’s tests were used to determine statistical differences between the groups (P<0.05). GraphPad Prism software (version 6.01) was used for data analysis.Results: The results showed that miRNA-101 clearly inhibited the expression of Mcl-1 and reduced the growth of A549 cells, relative to blank control and negative control miRNA (P<0.05). Transfection of miRNA-101 synergistically enhanced the sensitivity of the A549 cells to etoposide. Apoptosis assay data also showed that miRNA-101 triggered apoptosis and augmented the etoposide-mediated apoptosis. Conclusion: Up-regulation of miRNA-101 inhibited cell survival and proliferation, and sensitized A549 cells to etoposide by suppressing Mcl-1 expression. miRNA-101 replacement therapy can be considered as an effective therapeutic strategy in non-small cell lung cancer.

Exploring microRNA target genes and identifying hub genes in bladder cancer based on bioinformatic analysis

BackgroundBladder cancer (BC) is the second most frequent malignancy of the urinary system. The aim of this study was to identify key microRNAs (miRNAs) and hub genes associated with BC as well as analyse their targeted relationships.MethodsAccording to the microRNA dataset GSE112264 and gene microarray dataset GSE52519, differentially expressed microRNAs (DEMs) and differentially expressed genes (DEGs) were obtained using the R limma software package. The FunRich software database was used to predict the miRNA-targeted genes. The overlapping common genes (OCGs) between miRNA-targeted genes and DEGs were screened to construct the PPI network. Then, gene ontology (GO) analysis was performed through the “cluster Profiler” and “org.Hs.eg.db” R packages. The differential expression analysis and hierarchical clustering of these hub genes were analysed through the GEPIA and UCSC Cancer Genomics Browser databases, respectively. KEGG pathway enrichment analyses of hub genes were performed through gene set enrichment analysis (GSEA).ResultsA total of 12 DEMs and 10 hub genes were identified. Differential expression analysis of the hub genes using the GEPIA database was consistent with the results for the UCSC Cancer Genomics Browser database. The results indicated that these hub genes were oncogenes, but VCL, TPM2, and TPM1 were tumour suppressor genes. The GSEA also showed that hub genes were most enriched in those pathways that were closely associated with tumour proliferation and apoptosis.ConclusionsIn this study, we built a miRNA-mRNA regulatory targeted network, which explores an understanding of the pathogenesis of cancer development and provides key evidence for novel targeted treatments for BC.

Long non‑coding RNA SNHG3 promotes the development of non‑small cell lung cancer via the miR‑1343‑3p/NFIX pathway.

The aim of the present study was to identify the function of long non-coding RNA (lncRNA) small nucleolar RNA host gene 3 (SNHG3) and examine its effects on non-small cell lung cancer (NSCLC). A series of in vitro experiments were employed to evaluate the effects of SNHG3 on the progression of NSCLC, including Cell Counting Kit-8, 5-Ethynyl-2′-deoxyuridine, flow cytometry, wound healing, Transwell, western blotting and reverse transcription-quantitative PCR assays. Bioinformatics analyses and a luciferase reporter assay were performed to identify the target gene of SNHG3 and microRNA (miR)-1343-3p. Finally, recuse experiments were conducted to verify the effect of SNHG3 and its target gene on proliferation, apoptosis, migration and invasion. The findings indicated that lncRNA SNHG3 was highly expressed in NSCLC tissues and cell lines. Knockdown of lncRNA SNHG3 inhibited cell proliferation, migration and invasion, and accelerated cell apoptosis in NSCLC cell lines. The results of the bioinformatics analysis and the luciferase reporter assay indicated that lncRNA SNHG3 directly bound to miR-1343-3p and that it could downregulate the expression levels of miR-1343-3p to promote the progression of NSCLC. Rescue experiments indicated that lncRNA SNHG3 increased nuclear factor IX (NFIX) expression by sequestering miR-1343-3p in NSCLC. These results suggested that the SNHG3/miR-1343-3p/NFIX axis may serve as a novel prognostic biomarker and therapeutic target for NSCLC.

Construction of a Myc-associated ceRNA network reveals a prognostic signature in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) remains an extremely lethal disease worldwide. High-throughput methods have revealed global transcriptome dysregulation; however, a comprehensive investigation of the complexity and behavioral characteristics of the competing endogenous RNA (ceRNA) network in HCC is lacking. In this study, we extracted the transcriptome (RNA) sequencing data of 371 HCC patients from The Cancer Genome Atlas platform. With the comparison of the high Myc expression (Mychigh) tumor and low Myc expression (Myclow) tumor groups in HCC, we identified 1,125 differentially expressed (DE) mRNAs, 589 long non-coding RNAs (lncRNAs), and 93 microRNAs (miRNAs). DE RNAs predicted the interactions necessary to construct an associated Myc ceRNA network, including 19 DE lncRNAs, 5 miRNAs, and 72 mRNAs. We identified a significant signature (long intergenic non-protein-coding [LINC] RNA 2691 [LINC02691] and LINC02499) that effectively predicted overall survival and had protective effects. The target genes of microRNA (miR)-212-3p predicted to intersect with DE mRNAs included SEC14-like protein 2 (SEC14L2) and solute carrier family 6 member 1 (SLC6A1), which were strongly correlated with survival and prognosis. With the use of the lncRNA-miRNA-mRNA axis, we constructed a ceRNA network containing four lncRNAs (LINC02691, LINC02499, LINC01354, and NAV2 antisense RNA 4), one miRNA (miR-212-3p), and two mRNAs (SEC14L2 and SLC6A1). Overall, we successfully constructed a mutually regulated ceRNA network and identified potential precision-targeted therapies and prognostic biomarkers.

Topic Evolution Analysis for Omics Data Integration in Cancers.

One of the vital challenges for cancer diseases is efficient biomarkers monitoring formation and development are limited. Omics data integration plays a crucial role in the mining of biomarkers in the human condition. As the link between omics study on biomarkers discovery and cancer diseases is deepened, defining the principal technologies applied in the field is a must not only for the current period but also for the future. We utilize topic modeling to extract topics (or themes) as a probabilistic distribution of latent topics from the dataset. To predict the future trend of related cases, we utilize the Prophet neural network to perform a prediction correction model for existing topics. A total of 2,318 pieces of literature (from 2006 to 2020) were retrieved from MEDLINE with the query on “omics” and “cancer.” Our study found 20 topics covering current research types. The topic extraction results indicate that, with the rapid development of omics data integration research, multi-omics analysis (Topic 11) and genomics of colorectal cancer (Topic 10) have more studies reported last 15 years. From the topic prediction view, research findings in multi-omics data processing and novel biomarker discovery for cancer prediction (Topic 2, 3, 10, 11) will be heavily focused in the future. From the topic visuallization and evolution trends, metabolomics of breast cancer (Topic 9), pharmacogenomics (Topic 15), genome-guided therapy regimens (Topic 16), and microRNAs target genes (Topic 17) could have more rapidly developed in the study of cancer treatment effect and recurrence prediction.

Differential expression analysis and profiling of hepatic miRNA and isomiRNA in dengue hemorrhagic fever

Dengue virus causes dengue hemorrhagic fever (DHF) and has been associated to fatal cases worldwide. The liver is one of the most important target tissues in severe cases, due to its intense viral replication and metabolic role. microRNAs role during infection is crucial to understand the regulatory mechanisms of DENV infection and can help in diagnostic and anti-viral therapies development. We sequenced the miRNome of six fatal cases and compared to five controls, to characterize the human microRNAs expression profile in the liver tissue during DHF. Eight microRNAs were differentially expressed, including miR-126-5p, a regulatory molecule of endothelial cells, miR-122-5p, a liver specific homeostasis regulator, and miR-146a-5p, an interferon-regulator. Enrichment analysis with predicted target genes of microRNAs revealed regulatory pathways of apoptosis, involving MAPK, RAS, CDK and FAS. Immune response pathways were related to NF- kB, CC and CX families, IL and TLR. This is the first description of the human microRNA and isomicroRNA profile in liver tissues from DHF cases. The results demonstrated the association of miR-126-5p, miR-122-5p and miR-146a-5p with DHF liver pathogenesis, involving endothelial repair and vascular permeability regulation, control of homeostasis and expression of inflammatory cytokines.

Combined De-Repression of Chemoresistance Associated Mitogen-Activated Protein Kinase 14 and Activating Transcription Factor 2 by Loss of microRNA-622 in Hepatocellular Carcinoma.

Simple SummaryNovel approaches to target RAS-signaling have led to the so called “RAS renaissance”. In HCC, it was demonstrated that wild-type RAS-RAF-ERK-signaling strongly contributes to HCC progression and drug resistance. In this context, microRNA-622 has emerged as one of the most promising tumorsuppressive microRNAs, targeting RAS-signaling in HCC. However, the majority of microRNA-622 target genes remained elusive. Moreover, limited chemoresponse of HCC was demonstrated by activation of MAPK14-ATF2-signaling which represents a RAS-collateral-pathway, but the expression and regulation of both MAPK14 and ATF2 remained to be clarified. In this study, we demonstrated co-overexpression of MAPK14 and ATF2 in HCC in vitro and in vivo. Moreover, in contrast to “one-microRNA-one-target” interactions, we identified common binding sites for microRNA-622 in both synergistic drug resistance-associated genes–MAPK14 and ATF2. Our data revealed that microRNA-622 functions as a superior pathway regulator inducing de-repression of the MAPK14-ATF2-axis in HCC.Chemoresistance is a major hallmark driving the progression and poor prognosis of hepatocellular carcinoma (HCC). Limited chemoresponse of HCC was demonstrated to be mediated by mitogen-activated protein kinase 14 (MAPK14) and activating transcription factor 2 (ATF2). Recently, we have demonstrated loss of control of RAS-RAF-ERK-signaling as a consequence of miR-622 downregulation in HCC. However, the majority of target genes of this potent tumorsuppressive microRNA had remained elusive. The MAPK14-ATF2-axis represents a collateral pathway ensuring persisting ERK-activation in the presence of sorafenib-mediated RAF-inhibition. In contrast to the function of the MAPK14-ATF2-axis, both the expression and regulation of MAPK14 and ATF2 in human HCC remained to be clarified. We found combined overexpression of MAPK14 and ATF2 in human HCC cells, tissues and in sorafenib resistant cell lines. High expression of MAPK14 and ATF2 was associated with reduced overall survival in HCC patients. Deciphering the molecular mechanism promoting combined upregulation of MAPK14 and ATF2 in HCC, we revealed that miR-622 directly targets both genes, resulting in combined de-repression of the MAPK14-ATF2-axis. Together, miR-622 represents a superior regulator of both RAS-RAF-ERK as well as MAPK14-ATF2-signaling pathways in liver cancer.

Extracellular Vesicle-Derived microRNAs of Human Wharton's Jelly Mesenchymal Stromal Cells May Activate Endogenous VEGF-A to Promote Angiogenesis.

Despite low levels of vascular endothelial growth factor (VEGF)-A, the secretome of human Wharton’s jelly (WJ) mesenchymal stromal cells (MSCs) effectively promoted proangiogenic responses in vitro, which were impaired upon the depletion of small (~140 nm) extracellular vesicles (EVs). The isolated EVs shared the low VEGF-A profile of the secretome and expressed five microRNAs, which were upregulated compared to fetal dermal MSC-derived EVs. These upregulated microRNAs exclusively targeted the VEGF-A gene within 54 Gene Ontology (GO) biological processes, 18 of which are associated with angiogenesis. Moreover, 15 microRNAs of WJ-MSC-derived EVs were highly expressed (Ct value ≤ 26) and exclusively targeted the thrombospondin 1 (THBS1) gene within 75 GO biological processes, 30 of which are associated with the regulation of tissue repair. The relationship between predicted microRNA target genes and WJ-MSC-derived EVs was shown by treating human umbilical-vein endothelial cells (HUVECs) with appropriate doses of EVs. The exposure of HUVECs to EVs for 72 h significantly enhanced the release of VEGF-A and THBS1 protein expression compared to untreated control cells. Finally, WJ-MSC-derived EVs stimulated in vitro tube formation along with the migration and proliferation of HUVECs. Our findings can contribute to a better understanding of the molecular mechanisms underlying the proangiogenic responses induced by human umbilical cord-derived MSCs, suggesting a key regulatory role for microRNAs delivered by EVs.

MiR-146b-3p regulates proliferation of pancreatic cancer cells with stem cell-like properties by targeting MAP3K10.

Purpose: Cancer stem cells (CSCs) initiate and maintain tumorigenesis due to their unique pluripotency. However, pancreatic stem cell gene signatures are not completely revealed yet. Here, we isolated pancreatic cancer stem cells (P-CSCs) and exploited their distinct genome-wide mRNA and miRNA expression profiles using microarrays.Methods: CD24+ CD44+ ESA+ cells were isolated from two pancreatic xenograft cells by the flow cytometry and identified the stem cell-like properties by the tumor formation, self-renew and chemoresistance. Microarrays and qRT-PCR were used to exploit their distinct Genome-wide mRNA and miRNA expression profiles. The function and candidate target genes of key microRNA were detected after Ectopic restoration in the pancreatic cancer cell lines MIA Paca-2 (CSChigh) and BxPC-3 (CSClow).Results: In this study, we isolated P-CSCs from two xenografts cells. Genome-wide profiling experiments showed 479 genes and 15 microRNAs specifically expressed in the P-CSCs, including genes involved in TGF-β and p53 signaling pathways and particularly miR-146b-3p as the most significantly downregulated miRNA. We confirmed miR-146b-3p as a downregulated signature in pancreatic cancer tissues and cell line MIA Paca-2 (CSChigh) cells. Ectopic restoration of miR-146b-3p expression with pre-miR reduced cell proliferation, induced apoptosis, increased G1 phase and reduced S phase in cell cycle in MIA Paca-2 (CSChigh), but not in BxPC-3 (CSClow). Re-expression of miR-146b-3p with lentivirus significantly inhibited tumorigenicity in vivo in MIA Paca-2, but slightly in BxPC-3. Furthermore, we demonstrated that miR-146b-3p directly targeted MAP3K10 and might activate Hedgehog pathway as well through DYRK2 and GLI2.Conclusions: These results suggest that P-CSCs have distinct gene expression profiles. MiR-146b-3p inhibits proliferation and induced apoptosis in P-CSCs high cells lines by targeting MAP3K10. Targeting P-CSCs specific genes may provide novel strategies for therapeutic purposes.

SRNATargetDigger: A bioinformatics software for bidirectional identification of sRNA-target pairs with co-regulatory sRNAs information.

Identification of the target genes of microRNAs (miRNAs), trans-acting small interfering RNAs (ta-siRNAs), and small interfering RNAs (siRNAs) is an important step for understanding their regulatory roles in plants. In recent years, many bioinformatics software packages based on small RNA (sRNA) high-throughput sequencing (HTS) and degradome sequencing data analysis have provided strong technical support for large-scale mining of sRNA-target pairs. However, sRNA-target regulation is achieved using a complex network of interactions since one transcript might be co-regulated by multiple sRNAs and one sRNA may also affect multiple targets. Currently used mining software can realize the mining of multiple unknown targets using known sRNA, but it cannot rule out the possibility of co-regulation of the same target by other unknown sRNAs. Hence, the obtained regulatory network may be incomplete. We have developed a new mining software, sRNATargetDigger, that includes two function modules, “Forward Digger” and “Reverse Digger”, which can identify regulatory sRNA-target pairs bidirectionally. Moreover, it has the ability to identify unknown sRNAs co-regulating the same target, in order to obtain a more authentic and reliable sRNA-target regulatory network. Upon re-examination of the published sRNA-target pairs in Arabidopsis thaliana, sRNATargetDigger found 170 novel co-regulatory sRNA-target pairs. This software can be downloaded from http://www.bioinfolab.cn/sRNATD.html.

LBX2-AS1 promotes ovarian cancer progression by facilitating E2F2 gene expression via miR-455-5p and miR-491-5p sponging.

LBX2‐AS1 is a long non‐coding RNA that facilitates the development of gastrointestinal cancers and lung cancer, but its participation in ovarian cancer development remained uninvestigated. Clinical data retrieved from TCGA ovarian cancer database and the clinography of 60 ovarian cancer patients who received anti‐cancer treatment in our facility were analysed. The overall cell growth, colony formation, migration, invasion, apoptosis and tumour formation on nude mice of ovarian cancer cells were evaluated before and after lentiviral‐based LBX2‐AS1 knockdown. ENCORI platform was used to explore LBX2‐AS1‐interacting microRNAs and target genes of the candidate microRNAs. Luciferase reporter gene assay and RNA pulldown assay were used to verify the putative miRNA‐RNA interactions. Ovarian cancer tissue specimens showed significant higher LBX2‐AS1 expression levels that non‐cancerous counterparts. High expression level of LBX2‐AS1 was significantly associated with reduced overall survival of patients. LBX2‐AS1 knockdown significantly down‐regulated the cell growth, colony formation, migration, invasion and tumour formation capacity of ovarian cancer cells and increased their apoptosis in vitro. LBX2‐AS1 interacts with and thus inhibits the function of miR‐455‐5p and miR‐491‐5p, both of which restrained the expression of E2F2 gene in ovarian cancer cells via mRNA targeting. Transfection of miRNA inhibitors of these two miRNAs or forced expression of E2F2 counteracted the effect of LBX2‐AS1 knockdown on ovarian cancer cells. LBX2‐AS1 was a novel cancer‐promoting lncRNA in ovarian cancer. This lncRNA increased the cell growth, survival, migration, invasion and tumour formation of ovarian cancer cells by inhibiting miR‐455‐5p and miR‐491‐5p, thus liberating the expression of E2F2 cancer‐promoting gene.