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Abstract Background MicroRNAs (miRNAs) are small (19-22-nt) single-stranded noncoding RNA molecules whose deregulation of expression can contribute to human disease including the multistep processes of carcinogenesis in human. Circulating miRNAs are emerging biomarkers in many diseases and cancers such as type 2 diabetes, pulmonary disease, colorectal cancer, and gastric cancer among others; however, defining a plasma miRNA signature in acute myeloblastic leukemia (AML) that could serve as a biomarker for diagnosis or in the follow-up has not been done yet. Methods TaqMan miRNA microarray was performed to identify deregulated miRNAs in the plasma of AML patients. Quantitative real-time RT-PCR was used to validate the results. Receiver-operator characteristic (ROC) curve analysis was conducted to evaluate the diagnostic accuracy of the highly and significantly identified deregulated miRNA(s) as potential candidate biomarker(s). Results The plasma expression level of let-7d, miR-150, miR-339, and miR-342 was down-regulated whilst that of let-7b, and miR-523 was up-regulated in the AML group at diagnosis compared to healthy controls. ROC curve analyses revealed an AUC (the areas under the ROC curve) of 0.835 (95% CI: 0.7119– 0.9581; P Conclusions Our findings indicated that plasma miR-150 and miR-342 are novel important promising biomarkers in the diagnosis of AML. These novel and promising markers warrant validation in larger prospective studies.

Abstract Background The differentiation of CD8+ T lymphocytes following priming of naïve cells is central in the establishment of the adaptive immune response. Yet, the molecular events underlying this process are not fully understood. MicroRNAs have been recently shown to play a key role in the regulation of haematopoiesis in mouse, but their implication in peripheral lymphocyte differentiation in humans remains largely unknown. Methods In order to explore the potential implication of microRNAs in CD8+ T cell differentiation in humans, microRNA expression profiles were analysed using microarrays and quantitative PCR in several human CD8+ T cell subsets defining the major steps of the T cell differentiation pathway. Results We found expression of a limited set of microRNAs, including the miR-17~92 cluster. Moreover, we reveal the existence of differentiation-associated regulation of specific microRNAs. When compared to naive cells, miR-21 and miR-155 were indeed found upregulated upon differentiation to effector cells, while expression of the miR-17~92 cluster tended to concomitantly decrease. Conclusions This study establishes for the first time in a large panel of individuals the existence of differentiation associated regulation of microRNA expression in human CD8+ T lymphocytes in vivo, which is likely to impact on specific cellular functions.

CD4+CD25+ FOXP3+ regulatory T cells (Tregs) represent a subpopulation of CD4+ T cells central for the suppression of physiological and pathological immune reactions. Although distinct cell surface antigens are expressed in regulatory T cells, those components are also present on the surface of activated CD4+CD25- FOXP3-T cells, thus making the discrimination between Tregs and conventional CD4+ T difficult and isolation of Tregs complex. Yet, the molecular components driving Tregs' function are still not fully characterized. Aiming at unraveling molecular components specifically marking Tregs, and upon using quantitative real-time PCR (qRT-PCR) followed by bioinformatics analysis, we identified, in this study, differential transcriptional profiles, in peripheral blood CD4 + CD25 + CD127low FOXP3+ Tregs versus CD4 + CD25-FOXP3- conventional T cells, for set of genes with distinct immunological roles. In conclusion, this study identifies some novel genes that appeared to be differentially transcribed in CD4+ Tregs versus conventional T cells. The identified genes could serve as novel molecular targets relevant to Tregs' function and isolation. [ABSTRACT FROM AUTHOR]

Regulatory T cells (Tregs) are central for maintaining immune balance and their dysfunction drives the expansion of critical immunologic disorders. During the past decade, microRNAs (miRNAs) have emerged as potent regulators of gene expression among which immune‐related genes and their immunomodulatory properties have been associated with different immune‐based diseases. The miRNA signature of human peripheral blood (PB) CD8+CD25 +CD127 low Tregs has not been described yet. We thus identified, using TaqMan low‐density array (TLDA) technique followed by individual quantitative real‐time polymerase chain reaction (qRT‐PCR) confirmation, 14 miRNAs, among which 12 were downregulated whereas two were upregulated in CD8 +CD25 +CD127 low Tregs in comparison to CD8 +CD25 − T cells. In the next step, microRNA Data Integration Portal (mirDIP) was used to identify potential miRNA target sites in the 3′‐untranslated region (3′‐UTR) of key Treg cell‐immunomodulatory genes with a special focus on interleukin 10 (IL‐10) and transforming growth factor β (TGF‐β). Having identified potential miR target sites in the 3′‐UTR of IL‐10 (miR‐27b‐3p and miR‐340‐5p) and TGF‐β (miR‐330‐3p), we showed through transfection and transduction assays that the overexpression of two underexpressed miRNAs, miR‐27b‐3p and miR‐340‐5p, downregulated IL‐10 expression upon targeting its 3′‐UTR. Similarly, overexpression of miR‐330‐3p negatively regulated TGF‐β expression. These results highlighted an important impact of the CD8 + Treg mirnome on the expression of genes with significant implication on immunosuppression. These observations could help in better understanding the mechanism(s) orchestrating Treg immunosuppressive function toward unraveling new targets for treating autoimmune pathologies and cancer. [ABSTRACT FROM AUTHOR]

Interleukin‐21 (IL‐21) is a cytokine with potent regulatory effects on different immune cells. Recently, IL‐21 has been contemplated for use in the treatment of cancers. However, the molecular mechanisms regulating human IL‐21 gene expression has not yet been described. In this study, we initially studied the promoter region and identified the transcription start site. We thereafter described the essential region upstream of the transcription start site and showed the in vivo binding of NFATc2 and SP1 transcription factors to this region, in addition to their positive role in IL‐21 expression. We also studied the role of microRNAs (miRNAs) in regulating IL‐21 expression. We, thus, established the miRNA profile of CD4+CD45RO+ versus CD4+CD45RA+ isolated from healthy volunteers and identified a signature composed of 12 differentially expressed miRNAs. We showed that miR‐302c is able to negatively regulate IL‐21 expression by binding directly to its target site in the 3′‐untranslated region. Moreover, after using fresh human CD4‐positive T cells, we observed the high acetylation level of histone H4, an observation well in line with the already described high expression of IL‐21 in CD4+CD45RO+ versus CD4+CD45RA+ T cells. Altogether, our data identified different molecular mechanisms regulating IL‐21 expression. We defined the promoter region of human interleukin‐21 (IL‐21) gene and identified different mechanisms, including Sp1 and NFATc2 transcription factors. Acetylation status of histones and miR‐302c is be involved in regulating the IL‐21 expression. These molecular mechanisms could, therefore, be targeted to manipulate IL‐21 expression in the context of disease treatment. [ABSTRACT FROM AUTHOR]

Background Recently, regulatory T (Treg) cells have gained interest in the fields of immunopathology, transplantation and oncoimmunology. Here, we investigated the microRNA expression profile of human natural CD8+CD25+ Treg cells and the impact of microRNAs on molecules associated with immune regulation. Methods We purified human natural CD8+ Treg cells and assessed the expression of FOXP3 and CTLA-4 by flow cytometry. We have also tested the ex vivo suppressive capacity of these cells in mixed leukocyte reactions. Using TaqMan low-density arrays and microRNA qPCR for validation, we could identify a microRNA 'signature' for CD8+CD25+FOXP3+CTLA-4+ natural Treg cells. We used the 'TargetScan' and 'miRBase' bioinformatics programs to identify potential target sites for these microRNAs in the 3'-UTR of important Treg cellassociated genes. Results The human CD8+CD25+ natural Treg cell microRNA signature includes 10 differentially expressed microRNAs. We demonstrated an impact of this signature on Treg cell biology by showing specific regulation of FOXP3, CTLA-4 and GARP gene expression by microRNA using site-directed mutagenesis and a dual-luciferase reporter assay. Furthermore, we used microRNA transduction experiments to demonstrate that these microRNAs impacted their target genes in human primary Treg cells ex vivo. Conclusions We are examining the biological relevance of this 'signature' by studying its impact on other important Treg cell-associated genes. These efforts could result in a better understanding of the regulation of Treg cell function and might reveal new targets for immunotherapy in immune disorders and cancer. [ABSTRACT FROM AUTHOR]

Background: The differentiation of CD8+T lymphocytes following priming of naïve cells is central in the establishment of the adaptive immune response. Yet, the molecular events underlying this process are not fully understood. MicroRNAs have been recently shown to play a key role in the regulation of haematopoiesis in mouse, but their implication in peripheral lymphocyte differentiation in humans remains largely unknown. Methods: In order to explore the potential implication of microRNAs in CD8+T cell differentiation in humans, microRNA expression profiles were analysed using microarrays and quantitative PCR in several human CD8+T cell subsets defining the major steps of the T cell differentiation pathway. Results: We found expression of a limited set of microRNAs, including the miR-17∼92 cluster. Moreover, we reveal the existence of differentiation-associated regulation of specific microRNAs. When compared to naive cells, miR-21 and miR-155 were indeed found upregulated upon differentiation to effector cells, while expression of the miR- 17∼92 cluster tended to concomitantly decrease. Conclusions: This study establishes for the first time in a large panel of individuals the existence of differentiation associated regulation of microRNA expression in human CD8+T lymphocytes in vivo, which is likely to impact on specific cellular functions. [ABSTRACT FROM AUTHOR]

CD4 + CD25 + FOXP3 + regulatory T cells (Tregs) represent a subpopulation of CD4 + T cells central for the suppression of physiological and pathological immune reactions. Although distinct cell surface antigens are expressed in regulatory T cells, those components are also present on the surface of activated CD4 + CD25 - FOXP3 - T cells, thus making the discrimination between Tregs and conventional CD4 + T difficult and isolation of Tregs complex. Yet, the molecular components driving Tregs' function are still not fully characterized. Aiming at unraveling molecular components specifically marking Tregs, and upon using quantitative real-time PCR (qRT-PCR) followed by bioinformatics analysis, we identified, in this study, differential transcriptional profiles, in peripheral blood CD4 + CD25 + CD127 low FOXP3+ Tregs versus CD4 + CD25-FOXP3- conventional T cells, for set of genes with distinct immunological roles. In conclusion, this study identifies some novel genes that appeared to be differentially transcribed in CD4+ Tregs versus conventional T cells. The identified genes could serve as novel molecular targets relevant to Tregs' function and isolation.