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Lymphocyte microRNAs in health and disease: Understanding lymphocyte functions through the identification of microRNA target genes and exploiting serum microRNA signatures to monitor immune responses

Final Report Summary - I-MIRNOME (Lymphocyte microRNAs in health and disease: Understanding lymphocyte functions through the identification of microRNA target genes and exploiting serum microRNA signatures to monitor immune responses)

CD4+ T cell subsets are not only specialised to fight different classes of pathogens, but are also major drivers of immune mediated diseases. Differentiation of CD4+ T cells subsets has been extensively investigated at the gene expression level, but little is known on the post-transcriptional control exerted by microRNAs, small RNAs that bind mRNAs affecting their stability and/or translation. We have therefore decided to decipher the role of microRNAs in T cell differentiation in CD4+ T cell subsets purified from peripheral blood and from inflamed organs (liver and intestine). We will exploit this new knowledge to profile signatures of in-vivo activated T cell subsets and to map genes key to T cell differentiation that could provide a better understanding of T cell commitment and will provide functional evidence of key genes targeted by microRNAs which could therefore be the target of new immunomodulatory drugs. At the same time, we will develop quantitative assays to monitor microRNAs signatures in the serum of HCV-infected individuals and Crohn’s disease patients that could provide valuable non-invasive biomarkers, useful for the assessment and monitoring of the inflammation process.
We have described an “atlas” of the microRNAs present in human resting primary cells of seventeen lymphocyte subsets that circulate in the peripheral blood of healthy donors, and we have shown that there are specific signatures of microRNAs that mark those resting peripheral CD4+ T cell subsets, i.e. naïve, Th1, Th2 and Th17. Furthermore, we defined miR-125b as a CD4+ naïve T cells specific miRNA, that is involved in the maintenance of the naïve state. Moreover we have already profiled by RNA-seq the transcriptome of different human CD4+ T cell subsets purified by cell sorting from peripheral blood mononuclear cells (CD4+ naïve; Th1; Th2; Th17; Tregs) To analyze the RNA sequencing data we have improved the RNA-seq analysis pipeline integrating already existing tools and tools we have developed in a new more straightforward analysis workflow.
In recent years, it has been found that most cells release microRNAs (miRNAs) in the extra-cellular environment and this phenomenon has been exploited to assess cell functions at distance via measurement of serum miRNAs. However, we have found that the release of miRNA through exosomes is not a passive phenomenon, but actually a regulated process. Indeed, the exosomal RNA content is not at all a mere representation of the intracellular one. W have now identified 17 exosome-associated-miRNAs that are signature of of different CD4+ Th-cell subsets (e.g. Th1, Th2, Th17).
These studies will potentially have two different and equally relevant impacts on the field. On the one side, the analysis of lymphocyte extracellular signatures may shed light on the role of miRNA disposal during activation, as this type of release may represent an additional layer of post-transcriptional regulation for miRNAs with very rapid effects on target genes of the discarded miRNAs. On the other side, these same studies will pave the way for the identification of powerful biomarkers of lymphocyte activation. Indeed, upon the identification of miRNAs that are differentially released by different lymphocyte effector T cells, the assessment of their modulation in serum may render possible to mark the elicitation of these cells, which occurs in lymphoid tissues or damaged organs and would provide pivotal information about the nature of the immune responses occurring in both health and disease.