TGF-beta/atRA-induced Tregs express a selected set of microRNAs involved in the repression of transcripts related to Th17 differentiation

Regulatory T cells (Tregs) are essential regulators of immune tolerance. atRA and TGF-beta can inhibit the polarization of naive T cells into inflammatory Th17 cells, favoring the generation of stable iTregs, however the regulatory mechanisms involved are not fully understood. In this context, the roles of individual microRNAs in Tregs are largely unexplored. Naive T cells were immunomagnetically isolated from umbilical cord blood and activated with anti- human CD2/CD3/CD28 beads in the presence of IL-2 alone (CD4(Med)) or with the addition of TGF-beta and atRA (CD4(TGF/atRA)). As compared to CD4(Med), the CD4(TGF/atRA) condition allowed the generation of highly suppressive CD4+CD25(hi)CD127-FOXP3(hi) iTregs. Microarray profiling allowed the identification of a set of microRNAs that are exclusively expressed upon TGF-beta/atRA treatment and that are predicted to target a set of transcripts concordantly downregulated. This set of predicted targets were enriched for central components of IL-6/JAK/STAT and AKT-mTOR signaling, whose inhibition is known to play important roles in the generation and function of regulatory lymphocytes. Finally, we show that mimics of exclusively expressed miRs (namely miR-1299 and miR-30a-5p) can reduce the levels of its target transcripts, IL6R and IL6ST (GP130), and increase the percentage of FoxP3(+) cells among CD4(+) CD25(+/hi) cells.