T cells from miR-17-92 transgenic mice demonstrate enhanced Th1 phenotype. Splenic CD4+ T cells were immuno-magnetically isolated from miR-17-92 TG/TG or control animals. (A), miR-17-5p expression was analyzed in total RNA extracted from these freshly isolated cells. (B), Flow analysis was carried out on these freshly isolated cells for surface expression of CD49d, a subunit composing VLA-4. The grey-shaded region represents CD4+ T cells isolated from control wild type animals and the unshaded region with the solid line represents CD4+ T cells from miR-17-92 TG/TG mice. Dotted lines represent samples stained with isotype control Rat IgG2b. As the background staining with the isotype IgG2b was equally very low in the two cell types, the corresponding histograms are barely distinguishable each other. (C), Isolated cells were stimulated in Th1 skewing condition for 9 days and 5 × 106cells were then plated in fresh media for 24 hours, at which point supernatant was collected and analyzed for IFN-γ by ELISA. Both in (A), and (C), values in the two groups were statistically different with p < .01 using the student t test.