Effect of PM2.5 on MicroRNA Expression and Function in Nasal Mucosa of Rats With Allergic Rhinitis

2020 ◽  
Vol 34 (4) ◽  
pp. 543-553
Author(s):  
Yu Huang ◽  
Zhi-Qiang Guo ◽  
Ru-Xin Zhang ◽  
Ren-Wu Zhao ◽  
Wei-Yang Dong ◽  
...  
Keyword(s):  
Allergic Rhinitis ◽  
Nasal Mucosa ◽  
Mirna Expression ◽  
Expression Profiles ◽  
Mirna Gene ◽  
Ar Model ◽  
Qrt Pcr ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas ◽  
And Function

Background Particulate matter 2.5 (PM2.5) refers to particulate matter with aerodynamic equivalent diameter less than or equal to 2.5 µm, which is an important component of air pollution. PM2.5 aggravates allergic rhinitis (AR) and promotes AR nasal mucosa inflammation. Therefore, the influence of PM2.5 inhalation exposure on microRNA (miRNA) expression profiles and function in the nasal mucosa of AR rats was investigated. Methods Female Sprague Dawley rats were distributed randomly to 2 groups: AR model PM2.5 exposure group (ARE group) and AR model PM2.5-unexposed control group (ARC group). The rats of ARE group were made to inhale PM2.5 at a concentration of 200 µg/m3, 3 h/day, for 30 days. miRNA expression profiles of the nasal mucosa from both groups were determined using an miRNA gene chip and were verified by quantitative real-time PCR (qRT-PCR). Gene function enrichment analysis was performed using bioinformatics analysis. Results The ARE group revealed 20 significantly differentially expressed miRNAs, including 4 upregulated and 16 downregulated miRNAs (fold change > 1.5 or < 0.66, P < .05). Of these, 9 selected miRNAs were verified by qRT-PCR, and the results of 8 miRNAs were in accordance with the miRNA gene chip results, with highly positive correlation ( r = .8583, P = .0031). Numerous target genes of differentially expressed miRNAs were functionally enriched in high-affinity immunoglobulin E receptor signaling, ErbB signaling, mucin O-glycans biosynthesis, transforming growth factor β signaling, mitogen-activated protein kinase signal transduction, phosphatidylinositol signaling, mucopolysaccharide biosynthesis, mammalian target of rapamycin signaling, T cell receptor signaling, Wnt signaling, chemokine signal transduction, and natural killer cell-mediated cytotoxicity pathways. Conclusions PM2.5 causes significant changes in miRNA expression in the nasal mucosa of AR rats. miRNA plays an important role in regulating PM2.5 effects in AR rat biological behavior and mucosal inflammation. This study provides a theoretical basis for the prevention and treatment of AR from the effects of environmental pollution on the gene regulation mechanism.

scholarly journals Chinese Herbal Medicine Formula Shenling Baizhu San Ameliorates High-Fat Diet-Induced NAFLD in Rats by Modulating Hepatic MicroRNA Expression Profiles

2019 ◽  
Vol 2019 ◽  
pp. 1-14
Author(s):  
Maoxing Pan ◽  
Yuanjun Deng ◽  
Chuiyang Zheng ◽  
Huan Nie ◽  
Kairui Tang ◽  
...  
Keyword(s):  
Mirna Expression ◽  
High Fat Diet ◽  
Target Genes ◽  
Expression Profiles ◽  
Differentially Expressed ◽  
High Fat ◽  
Expression Levels ◽  
Qrt Pcr ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas

Objective. The purpose of present study was to investigate the potential mechanism underlying the protective effect of Shenling Baizhu San (SLBZS) on nonalcoholic fatty liver disease (NAFLD) by microRNA (miRNA) sequencing. Methods. Thirty male Wistar rats were randomly divided into a normal control (NC) group, a high-fat diet (HFD) group, and an SLBZS group. After 12 weeks, the biochemical parameters and liver histologies of the rats were assessed. The Illumina HiSeq 2500 sequencing platform was used to analyse the hepatic miRNA expression profiles. Representative differentially expressed miRNAs were further validated by qRT-PCR. The functions of the differentially expressed miRNAs were analysed by bioinformatics. Results. Our results identified 102 miRNAs that were differentially expressed in the HFD group compared with the NC group. Among those differentially expressed miRNAs, the expression levels of 28 miRNAs were reversed by SLBZS administration, suggesting the modulation effect of SLBZS on hepatic miRNA expression profiles. The qRT-PCR results confirmed that the expression levels of miR-155-5p, miR-146b-5p, miR-132-3p, and miR-34a-5p were consistent with those detected by sequencing. Bioinformatics analyses indicated that the target genes of the differentially expressed miRNAs reversed by SLBZS were mainly related to metabolic pathways. Conclusion. This study provides novel insights into the mechanism of SLBZS in protecting against NAFLD; this mechanism may be partly related to the modulation of hepatic miRNA expression and their target pathways.

scholarly journals Baicalein, a Natural Anti-Cancer Compound, Alters MicroRNA Expression Profiles in Bel-7402 Human Hepatocellular Carcinoma Cells

2017 ◽  
Vol 41 (4) ◽  
pp. 1519-1531 ◽  
Author(s):  
Beibei Bie ◽  
Jin Sun ◽  
Jun Li ◽  
Ying Guo ◽  
Wei Jiang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Cell Lines ◽  
Mirna Expression ◽  
Expression Profiles ◽  
Akt Pathway ◽  
Qrt Pcr ◽  
Cycle Arrest ◽  
Mirna Expression Profiles

Background/Aims: Baicalein has been shown to possess significant anti-hepatoma activity by inhibiting cell proliferation. Whether the anti-proliferative effect of baicalein is related to its modulation of miRNA expression in hepatocellular carcinoma (HCC) is still unknown. Methods: The anti-proliferative effects of baicalein on HCC cell line Bel-7402 was assessed by detecting the proliferation activity, cell cycle distribution, expression changes of p21/CDKN1A, P27/CDKN1B, total Akt and phosphoryted AKT. Microarray analysis was conducted to determine the miRNA expression profiles in baicalein-treated or untreated Bel-7402 cells and then validated by qRT-PCR in two HCC cell lines (Bel-7402 and Hep3B). The gain-of-function of miR-3127-5p was performed by detecting anti-proliferative effects after transfecting miRNA mimics in cells. Finally, the expression level of miR-3127-5p in different HCC cell lines was determined by qRT-PCR. Results: Baicalein was able to inhibit the proliferation of Bel-7402 cells by inducing cell cycle arrest at the S and G2/M phase via up-regulating the expression of p21/CDKN1A and P27/CDKN1B and suppressing the PI3K/Akt pathway. Baicalein could alter the miRNA expression profiles in Bel-7402 cells. Putative target genes for differentially expressed miRNAs could be enriched in terms of cell proliferation regulation, cell cycle arrest and were mainly involved in MAPK, PI3K-Akt, Wnt, Hippo and mTOR signaling pathways. MiR- 3127-5p, one of up-regulated miRNAs, exhibits low expression level in several HCC cell lines and its overexpression could inhibit cell growth of Bel-7402 and Hep3B cell lines by inducing S phase arrest by up-regulating the expression of p21and P27 and repressing the PI3K/Akt pathway. Conclusions: Modulation of miRNA expression may be an important mechanism underlying the anti-hepatoma effects of baicalein.

scholarly journals MicroRNA expression patterns associated with hyperfunctioning and non-hyperfunctioning phenotypes in adrenocortical adenomas

2014 ◽  
Vol 170 (4) ◽  
pp. 583-591 ◽  
Author(s):  
David Velázquez-Fernández ◽  
Stefano Caramuta ◽  
Deniz M Özata ◽  
Ming Lu ◽  
Anders Höög ◽  
...  
Keyword(s):  
Mirna Expression ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Gene Mutations ◽  
Mirna Expression Profile ◽  
Differentially Expressed ◽  
Adrenocortical Adenoma ◽  
Tumor Subtypes ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas

BackgroundThe adrenocortical adenoma (ACA) entity includes aldosterone-producing adenoma (APA), cortisol-producing adenoma (CPA), and non-hyperfunctioning adenoma (NHFA) phenotypes. While gene mutations and mRNA expression profiles have been partly characterized, less is known about the alterations involving microRNA (miRNA) expression.AimTo characterize miRNA expression profile in relation to the subtypes of ACAs.Subjects and methodsmiRNA expression profiles were determined in 26 ACAs (nine APAs, ten CPAs, and seven NHFAs) and four adrenal references using microarray-based screening. Significance analysis of microarrays (SAM) was carried out to identify differentially expressed miRNAs between ACA and adrenal cortices or between tumor subtypes. Selected differentially expressed miRNAs were validated in an extended series of 43 ACAs and ten adrenal references by quantitative RT-PCR.ResultsAn hierarchical clustering revealed separate clusters for APAs and CPAs, while the NHFAs were found spread out within the APA/CPA clusters. When NHFA was excluded, the clustering analysis showed a better separation between APA and CPA. SAM analysis identified 40 over-expressed and three under-expressed miRNAs in the adenomas as compared with adrenal references. Fourteen miRNAs were common among the three ACA subtypes. Furthermore, we found specific miRNAs associated with different tumor phenotypes.ConclusionThe results suggest that miRNA expression profiles can distinguish different subtypes of ACA, which may contribute to a deeper understanding of ACA development and potential therapeutics.

scholarly journals Renal Tissue miRNA Expression Profiles in ANCA-Associated Vasculitis—A Comparative Analysis

2021 ◽  
Vol 23 (1) ◽  
pp. 105
Author(s):  
Matic Bošnjak ◽  
Željka Večerić-Haler ◽  
Emanuela Boštjančič ◽  
Nika Kojc
Keyword(s):  
Mirna Expression ◽  
Expression Profiles ◽  
Target Organ Damage ◽  
Organ Damage ◽  
Renal Tissue ◽  
Differentially Expressed ◽  
Tissue Samples ◽  
Anca Associated Vasculitis ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas

Anti-neutrophil cytoplasm antibody (ANCA)-associated vasculitis (AAV) comprises autoimmune disease entities that cause target organ damage due to relapsing-remitting small vessel necrotizing vasculitis, and which affects various vascular beds. The pathogenesis of AAV is incompletely understood, which translates to considerable disease- and treatment-related morbidity and mortality. Recent advances have implicated microRNAs (miRNAs) in AAV; however, their accurate characterization in renal tissue is lacking. The goal of this study was to identify the intrarenal miRNA expression profile in AAV relative to healthy, non-inflammatory and inflammatory controls to identify candidate-specific miRNAs. Formalin-fixed, paraffin-embedded renal biopsy tissue samples from 85 patients were obtained. Comprehensive miRNA expression profiles were performed using panels with 752 miRNAs and revealed 17 miRNA that differentiated AAV from both controls. Identified miRNAs were annotated to characterize their involvement in pathways and to define their targets. A considerable subset of differentially expressed miRNAs was related to macrophage and lymphocyte polarization and cytokines previously deemed important in AAV pathogenesis, lending credence to the obtained results. Interestingly, several members of the miR-30 family were detected. However, a validation study of these differentially expressed miRNAs in an independent, larger sample cohort is needed to establish their potential diagnostic utility.

scholarly journals Detection of Differentially Expressed MicroRNAs in Rheumatic Heart Disease: miR-1183 and miR-1299 as Potential Diagnostic Biomarkers

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Ni Li ◽  
Jiangfang Lian ◽  
Sheng Zhao ◽  
Dawei Zheng ◽  
Xi Yang ◽  
...  
Keyword(s):  
Heart Disease ◽  
Pulmonary Artery ◽  
Rheumatic Heart Disease ◽  
Mirna Expression ◽  
Expression Profiles ◽  
Differentially Expressed ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas ◽  
Secondary Pulmonary Hypertension ◽  
Rheumatic Heart

This study compared microRNA (miRNA) expression profiles between rheumatic heart disease (RHD) patients and healthy controls to investigate their differential expression and help elucidate their mechanisms of action. Microarray analysis was used to measure miRNA expression, and a total of 133 miRNAs were shown to be significantly upregulated in RHD patients compared with controls, including miR-1183 and miR-1299. A total of 137 miRNAs, including miR-4423-3p and miR-218-1-3p, were significantly downregulated in RHD patients. Quantitative real-time-PCR confirmed microarray findings for miR-1183 and miR-1299 in both tissue and plasma. Bioinformatic predictions were also made of differentially expressed miRNAs as biomarkers in RHD by databases and GO/pathway analysis. Furthermore, we investigated miR-1183 and miR-1299 expression in RHD patients with secondary pulmonary hypertension (PAH). Our findings identified an important role for miR-1299 as a direct regulator of RHD, while the observed difference in expression of miR-1183 between RHD-PAH patients with high or low pulmonary artery pressure suggests that miR-1183 overexpression may reflect pulmonary artery remodeling. miR-1183 and miR-1299 appear to play distinct roles in RHD pathogenesis accompanied by secondary PAH and could be used as potential biological markers for disease development.

scholarly journals Genome-wide profiling of miRNA expression patterns in tubal endometriosis

Reproduction ◽  
2019 ◽  
Vol 157 (6) ◽  
pp. 525-534 ◽  
Author(s):  
Hang Qi ◽  
Guiling Liang ◽  
Jin Yu ◽  
Xiaofeng Wang ◽  
Yan Liang ◽  
...  
Keyword(s):  
Pathway Analysis ◽  
Mirna Expression ◽  
Gene Networks ◽  
Target Genes ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Mirna Gene ◽  
Differentially Expressed ◽  
Signal Pathways ◽  
Differentially Expressed Mirnas

MicroRNA (miRNA) expression profiles in tubal endometriosis (EM) are still poorly understood. In this study, we analyzed the differential expression of miRNAs and the related gene networks and signaling pathways in tubal EM. Four tubal epithelium samples from tubal EM patients and five normal tubal epithelium samples from uterine leiomyoma patients were collected for miRNA microarray. Bioinformatics analyses, including Ingenuity Pathway Analysis (IPA), Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, were performed. Quantitative real-time polymerase chain reaction (qRT-PCR) validation of five miRNAs was performed in six tubal epithelium samples from tubal EM and six from control. A total of 17 significantly differentially expressed miRNAs and 4343 potential miRNA-target genes involved in tubal EM were identified (fold change >1.5 and FDR-adjustedPvalue <0.05). IPA indicated connections between miRNAs, target genes and other gynecological diseases like endometrial carcinoma. GO and KEGG analysis revealed that most of the identified genes were involved in the mTOR signaling pathway, SNARE interactions in vesicular transport and endocytosis. We constructed an miRNA-gene-disease network using target gene prediction. Functional analysis showed that the mTOR pathway was connected closely to tubal EM. Our results demonstrate for the first time the differentially expressed miRNAs and the related signal pathways involved in the pathogenesis of tubal EM which contribute to elucidating the pathogenic mechanism of tubal EM-related infertility.

scholarly journals Putative Role of MicroRNA-Regulated Pathways in Comorbid Neurological and Cardiovascular Disorders

2009 ◽  
Vol 2009 ◽  
pp. 1-5 ◽  
Author(s):  
Sébastien S. Hébert
Keyword(s):  
Mirna Expression ◽  
Molecular Mechanisms ◽  
Expression Profiles ◽  
Heart Cell ◽  
Cardiovascular Disorders ◽  
Increased Risk ◽  
Mirna Expression Profiles ◽  
And Function ◽  
The Brain

Background. The conserved noncoding microRNAs (miRNAs) that function to regulate gene expression are essential for the development and function of the brain and heart. Changes in miRNA expression profiles are associated with an increased risk for developing neurodegenerative disorders as well as heart failure. Here, the hypothesis of how miRNA-regulated pathways could contribute to comorbid neurological and cardiovascular disorders will be discussed. Presentation. Changes in miRNA expression occurring in the brain and heart could have an impact on coexisting neurological and cardiovascular characteristics by (1) modulating organ function, (2) accentuating cellular stress, and (3) impinging on neuronal and/or heart cell survival. Testing. Evaluation of miRNA expression profiles in the brain and heart tissues from individuals with comorbid neurodegenerative and cardiovascular disorders will be of great importance and relevance. Implications. Careful experimental design will shed light to the deeper understanding of the molecular mechanisms tying up those different but yet somehow connected diseases.

miRNA Profiling in Human Granulopoiesis and in Activated Neutrophils From Skin Window.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2136-2136
Author(s):  
Maria Torp Larsen ◽  
Christoffer Hother ◽  
Mattias Hager ◽  
Corinna Cavan Pedersen ◽  
Lars Jacobsen ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Inflammatory Response ◽  
Mirna Expression ◽  
Peripheral Blood ◽  
Expression Profiles ◽  
Cycle Arrest ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas ◽  
Activated Neutrophils

Abstract Abstract 2136 Formation of polymorphonuclear neutrophils (PMN) is a tightly regulated process where the myeloid progenitor cells, myeloblasts (MBs), divide and mature in the bone marrow, along a well defined path. The cells pass through six well defined stages in differentiation ending up with the release of mature PMNs to peripheral blood (granulopoiesis). Expression of essential transcription factors such as RUNX1, C/EBP-a, and C/EBP-e during granulopoiesis has been shown to have great importance for correct neutrophil development. microRNAs (miRNAs) could be important players in the fine-tuning of transcription factor expression due to their ability to regulate protein synthesis. The function of neutrophils is to detect and destroy invading microorganisms. This involves activation of the PMNs in the blood stream causing a release of secretory vesicles and up-regulation of extracellular adhesion molecules followed by migration in the tissue towards the focus of inflammation. Expression of miRNAs might also be regulated during activation and diapedesis of the neutrophils in order to adapt the neutrophil to its new environment and function. A regulatory role for miRNAs has been demonstrated for several biological processes, such as proliferation, differentiation, inflammation and cancer, and dysregulation of miRNA expression has been shown to contribute to disease development. The purpose of this study was to determine the miRNA expression profiles during normal human granulopoiesis starting with the first identifiable granulocytic precursor cell (MB) and ending with activated neutrophils that have migrated into the tissue using an Affymetrix 2,0 miRNA microarray platform. We isolated four populations of cells: Myeloblasts (MB) and promyelocytes (PM), myelocytes (MC) and metamyelocytes (MM), and band cells (BC) and segmented cells (SC) from the bone marrow and PMNs from peripheral blood from three different donors. We found 135 differentially expressed miRNAs in granulopoiesis, which could be divided into six clusters according to their expression pattern. 87% of the 135 miRNAs were differentially regulated between the MB/PM (dividing cells) and the MC/MM stages (cessation of cell proliferation and initiation of terminal differentiation) and could imply a need for miRNA-mediated regulation of the many proteins involved in regulating this process. Interestingly, we also found two distinct clusters of miRNAs that were either up- or down-regulated only in the MC/MM population, indicating the importance of a specific temporary regulation of some proteins during neutrophil development. To determine miRNA expression profiles in activated granulocytes, we examined PMNs and activated neutrophils from skin window (i.e. PMNs migrated to a site of inflammation). We found seven differentially expressed miRNAs, - all of them up regulated in the activated neutrophils. Using microRNA target-prediction software, we found that miRNAs 155, 146a and 130a, all of which are strongly up-regulated in the MB/PM stage, have several targets in the IL1-receptor signalling cascade, indicating the importance of miRNA of dampening an innate immune response in immature neutrophil precursors. miR-146a, 155 and 130a also have predicted targets in either the TGF-βI or the TGF-βII receptor which inhibits proliferation when binding to TGF-β. This finding supports the proliferating profile for the MB/PM cells, and the shift towards cell cycle arrest when the cells differentiate to the next stage, where expression of these three miRNAs is low. miRNA-34c-3p is highly expressed only in the MC/MM stage and has verified targets in many different mRNAs involved in the regulation of cell cycle arrest. All the miRNAs that were up-regulated in the activated neutrophils have several predicted targets in the IL1R pathway, and some of them (miR-212, −132 and −297) have previously been shown to be important in regulating the inflammatory response. The study indicates that several different miRNAs have important roles in the regulation of normal granulopoiesis, and that miRNAs also might be part of a possible negative feed back loop in the inflammatory response in activated neutrophils. Grant acknowledgments: The Danish Cancer Society, Lundbeck foundation, Danish Medical Research Council, Brøchner Mortensen foundation Disclosures: No relevant conflicts of interest to declare.

Comparative Analysis of miRNA Expression Profiles of Multiple Myeloma with 1q21 Gains and Normal FISH

2018 ◽  
Vol 139 (2) ◽  
pp. 96-100
Author(s):  
Yushan Cui ◽  
Yang Liu ◽  
Danyang Wang ◽  
Yuzhang Liu ◽  
Lina Liu ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Mirna Expression ◽  
Expression Profiles ◽  
In Situ Hybridisation ◽  
P Value ◽  
Fluorescence In Situ Hybridisation ◽  
Expression Array ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas ◽  
Normal Fish

Background: Multiple myeloma (MM) with 1q21 gains invariably has a poor prognosis. Many recent studies have reported the relationship between micro (mi)RNA expression and MM prognosis. However, there is little information on the association between miRNA alterations and 1q21 gains. Methods: We compared the miRNA expression profiles of MM with 1q21 gains and MM with normal fluorescence in situ hybridisation (FISH) by gene expression array. Differentially expressed miRNAs were identified using Affymetrix TAC software. Thresholds were defined as a false discovery rate <0.05, p value <0.05, and n-fold change >2. Results: Six miRNAs (let-7f-5p and -7g-5p, and miR-29a-3p, -29b-1-5p, -331-3p, and -223-3p) were downregulated and 4 (miR-30e-5p, -17-3p, -18b-5p, and -19a-3p) were upregulated in MM with 1q21 gains relative to MM with normal FISH. Conclusions: The identified set of miRNAs can serve as biomarkers for distinguishing MM with 1q21 gains from MM with normal FISH.

miRNAs Function As Biomarker in Pediatric AML and Regulate Gene-Silencing in AML-Relevant Pathways

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 570-570
Author(s):  
Svenja Daschkey ◽  
Silja Röttgers ◽  
Jutta Bradtke ◽  
Andrea Teigler-Schlegel ◽  
Arndt Borkhardt ◽  
...  
Keyword(s):  
Cell Growth ◽  
Mirna Expression ◽  
Expression Profiles ◽  
Statistical Testing ◽  
Differentially Expressed ◽  
Mirna Sequence ◽  
Argonaute Proteins ◽  
Mirna Expression Profiles ◽  
Differentially Expressed Mirnas ◽  
Pediatric Aml

Abstract Abstract 570 microRNAs (miRNAs) are small (21-24 nt), non-coding and highly conserved molecules, which are involved in several important regulatory processes like cell growth, proliferation, differentiation, immune response and apoptosis. Thus, their involvement in the pathogenesis of several diseases, including acute myeloid leukemia (AML) is not surprising. Several studies address the miRNA expression changes in adulthood AML, however, comprehensive studies in AML of children and adolescents are missing so far. We investigated the miRNA expression profiles of different AML subtypes from pediatric patients, in order to identify differentially expressed miRNAs. Subsequently, appropriate cell line models were used for global biochemical identification of miRNA targeting structures. miRNA expression profiles of 102 pediatric AML patient samples were identified using microarray technology, and analyzed by unsupervised hierarchical cluster analysis and statistical testing. AML subtypes with translocations t(8;21) and t(15;17) can be separated from each other, solely based on their miRNA expression profile, while other translocations involving mixed-lineage leukemia (MLL) rearrangements are interspersed and lack a characteristic miRNA signature. Only six and seven miRNAs are differentially expressed between AML samples with translocations t(8;21) and t(15;17), respectively, and all other AML subtypes. This is surprising, since patients of different AML subtypes, investigated in this study, differ greatly in their clinical presentation. Differentially expressed miRNAs contain lineage specific miRNAs (miR-223), oncogenic miRNAs (miR-21) and more ubiquitously expressed miRNAs (let-7b/c, miR-100, −125b and −181a/b) with no designated characteristics. Furthermore, these differentially expressed miRNAs were not described as abundant in adult AML patients. To gain further insights into the function of differentially expressed miRNAs, we established a modified PAR-CLIP method termed PAR-CLIP-Array (Photo-activatable-Ribonucleoside-Enhanced Crosslinking-Immunoprecipitation and Microarray Hybridization) for global identification of Ago-associated miRNAs and their mRNA-targets. On average 25% of mRNAs in AML cell lines bearing the AML1/ETO or PML/RARα translocation were identified in Argonaute complexes and carry at least one miRNA binding site and thus are under miRNA control. 60% and 27% of miRNAs and mRNAs, respectively, overlap between the four analyzed Argonaute proteins, while 50% and 52% (46 miRNAs and 241 mRNAs) were associated with one Argonaute protein specifically. However, pathway classification of Ago-associated target-mRNAs indicate more than 90% overlap between the Argonaute proteins and thus are indicative of a concerted action of these four proteins in 150 pathways identified. Moreover, miR-181a/b, up-regulated in t(15;17)-positive AML patients, were detected in association with the four human Argonaute proteins in NB4 cells and show binding sites for the protein kinase PDPK1 potentially leading to inhibition of AKT, whereas eight other Ago-associated miRNA sequence families (seqgrp-miR-98, −130a, −19a, −25, −27a, −301a, −361 and −320) in association with Ago3 are able to repress the upstream tumor suppressor TSC1 leading to activation of the mTOR pathway and increased cell growth. In addition, the repression of the MAP kinase phosphatase DUSP6 by four Ago-associated miRNA sequence families (seqgrp-miR-29a, −17, −125a and −98) leads to activation of proliferative genes in the MAPK pathway of both, t(8;21)- and t(15;17)-positive AML. In summary, miRNAs represent suitable biomarkers for differentiation of AML subtypes of pediatric AML patients. Furthermore, our studies show that the four human Argonaute proteins cooperate in the regulation of AML-relevant signaling pathways providing new insights into AML biology and may present the starting point for novel therapeutic interventions. Disclosures: No relevant conflicts of interest to declare.

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