Ca<sup>2+</sup> influx is a key event for IL-2 in adult CD31<sup>+</sup> naive T cells.


<p>(A) No Ca<sup>2+</sup> influx signal with GAMIg alone (black curve; anti-CD28 isotype Ab were substituted for anti-CD28 Ab) or anti-CD28 Ab plus GAMIg alone (red curve) in T cells stained for anti-CD4, anti-CD45, and anti-CD31. Results representative of at least 3 experiments are shown. (B-C) Ca<sup>2+</sup> influx into adult CD31<sup>+</sup> naive T cells stimulated using anti CD3 Ab and anti CD28 Ab. (B) A representative Ca<sup>2+</sup> influx experiment and (C) box plots for the different T cell subsets showing scatter plots of mean value and SD of Ca<sup>2+</sup> influx experiments by normalizing maximal Ca<sup>2+</sup> signals to the maximal Ca<sup>2+</sup> influx ionomycin are shown (* <i>P<</i>0.05; two-tailed ANOVA of differences). (D) Increased Ca<sup>2+</sup> influx in adult CD31<sup>+</sup> naive T cells compared with anti-CD3/anti-CD28 Ab (red) in response to anti-CD3 Ab alone (blue, with anti-CD28 isotype Ab) (* <i>P<</i>0.05; two-tailed ANOVA of differences) (E) Increased levels of NFATc2 protein expression in response to anti-CD3/anti-CD28 Ab stimulation in naive T cells. The immunoblot detection of the relative protein expression level by ratio of NFATc2 or pNFATc2 to αTubulin is shown for naive CD31<sup>+</sup> or CD31<sup>-</sup> T cells of adults stimulated with anti-CD3 Ab in combination with soluble anti-CD28 Ab (dark gray) or anti-CD28 Ab isotype (light gray). Laminin and αTubulin were used as loading controls. Results are representative of at least two experiments. unstim. = unstimulated. (F) IL-2 cytokine in supernatants does not differ in cultures under TCR/CD3 stimulation between subtypes of naive CD4 T cells (* <i>P<</i>0.05; two-tailed ANOVA of differences). ns = not significant.</p

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