Neutralizing IL-2 reduces the survival of CD8⁺ but not CD4⁺ T cells.

<p>CD4⁺CD25<b>⁻</b> and CD8⁺CD25<b>⁻</b> T cells were purified from wild-type B6 mice and stimulated with anti-CD3/CD28 mAbs in the presence or absence of neutralizing IL-2 mAb at two different concentrations as indicated. Some cells were left unstimulated (no anti-CD3/CD28) and were used as negative controls. A total of 5×10⁴ cells per well were cultured in flat-bottom 96-well plates and harvested for analysis at the indicated time points. (<b>A</b>) Harvested cells were stained with Propidium Iodide (PI) to measure cell survival by flow cytometry. Bar graphs show the percentage of PI negative cells in each group (n = 3). (<b>B</b>) Supernatants were harvested from the cultures at the indicated time points which were then analyzed for levels of IL-2 using ELISA. Bar graphs show the concentration of IL-2 in the culture supernatants. (<b>C</b>) Histograms show the expression of the anti-apoptotic protein Bcl-2 measured by flow cytometry in the cultured cells at the indicated time points. (<b>D</b>) Histograms represent the mitochondrial membrane potential measured by flow cytometry of harvested cells (with higher fluorescence depicting a hyper-polarized state of mitochondria) at the indicated time points. The figure shows results from one representative experiment which was repeated three times with similar results. Bars indicate average value ± standard deviation (SD) obtained from three culture wells (technical triplicates) in each experimental group (*denotes <i>p</i><0.05 and **denotes <i>p</i><0.005).