Expression of miR-20a and miR-20a precursors in human naïve CD4⁺ T cells.

<p>Human naïve CD4⁺ T cells were stimulated with plate-bound CD3 and CD28 mAbs for the indicated time periods. Quantification of <b>(A)</b> miR-20a and <b>(B)</b> miR-20a precursors (both pri- and pre-miRNAs) was performed using RT qPCR. <b>(C)</b> Naïve CD4⁺ T cells were also preincubated with DMSO, U0126 (10 μM), IKK VII (200 nM) and EGTA (10 mM) for 30 minutes. Subsequently, samples were either left unstimulated or stimulated with plate-bound CD3 for 4 hours at 37°C. Expression of miR-20a precursors was quantified using RT qPCR. <b>(D)</b> Quantification of miR-20a level upon overexpression. Human naïve CD4⁺ T cells were transfected with plasmids encoding either miR-20a or miR-control (miR-Ctrl). Cells were either left unstimulated or stimulated for 2 hours with CD3xCD28 mAbs. Quantification of miR-20a in both resting and activated cells was performed using real-time PCR. Unless otherwise mentioned, all the data was normalized to either resting cells or unstimulated control transfected cells. Data represent arbitrary units ± SEM of at least 3 (A, C, D) and 2 (B) independent experiments. Significant P values were done by using Student’s t Test. (*, P < 0.05; **, P < 0.01; ***, P < 0.001).