Figure 2. Ortho IL-2 selectively expands oIL-2Rβ–transduced Tregs without affecting cocultured naive T cells or reducing Treg functions.

Naive Thy1.1⁺ Tcons were cocultured with Thy1.2⁺Foxp3^GFP+ Tregs that were transduced with oIL-2Rβ (oTregs; transduction efficiency was ~30%). Flow cytometry analyses performed after 4-day coculture. (A) Representative histograms show expression levels of CD25 and ICOS and dilution of proliferation dye on Thy1.1⁺ Tcons. (B) Representative pseudocolor plots show the proportion of Foxp3^GFP+ cells (oTreg: upper gate) and CD25^hiThy1.1⁺ Tcons (lower gate). Gray, no IL-2; orange, WT IL-2 (1 × 10³ IU/mL); red, ortho IL-2 (1 × 10⁵ IU/mL). Box plots showing CD25 MFI (C) and percentage of proliferation (D) in Thy1.1⁺ Tcons. Thy1.1⁺ Tcon alone is shown as no suppression control (Treg [–], white box). P values in yellow/red on the columns were calculated between Tcon alone control and coculture at each IL-2 concentration. (E) Box plots showing CD25 MFI in oTregs. (F) Cell number per well relative to no IL-2 control in Thy1.1⁺ Tcons (black line) or oTregs (green line) with mean plus 95% confidence intervals. P values calculated between Thy1.1⁺ Tcons and oTreg at each IL-2 concentration are shown. (G) Box plots showing percentage of oTregs per well. P values were calculated between no IL-2 control and each IL-2 concentration. Quantification of triplicate wells in 1 representative experiment of 3 independent experiments. **P < 0.01; ***P < 0.001, Welch’s 2-sample t test. WT and ortho IL-2: 1 IU = 312.5 ng.