Different Ca²⁺ responses of CB and adult CD31⁺ naive T cells.

Abstract

<p>(A-B) Ca²⁺ mobilization in CD4⁺CD45RA⁺CD31⁺ T cells of one healthy donor (representative of at least eight healthy individuals) of PBMCs (A, adult) or CB (B) were performed in response to 0.05 μg/ml anti-CD3 Ab plus 0.5 μg/ml soluble anti-CD28 Ab (red curve) or only 0.05 μg/ml of anti-CD3 Ab alone (black curve, with anti-CD28 isotype Ab) in combination with GAMIg. The blue dotted line displays the maximum Ca²⁺ response of adult CD31⁺ naive T cells for anti-CD3 Ab stimulation alone. (C) Box plot with scatter plots representing means and SD of Ca²⁺ influx response normalized by maximal Ca²⁺ influx response to ionomycin of adult (gray circle) and CB (black circle) and their dependency on anti-CD28 Ab costimulation (anti-CD3 Ab plus anti-CD28 Ab (red box) or anti-CD3 Ab with anti-CD28 Ab isotype (blue box)). Statistical significance between groups * <i>P<</i>0.05 was determined by two-tailed ANOVA with Tukey-Kramer post-hoc test. n = number of individuals. (D) Comparison of different CD4⁺ T cell subset stimulated with anti-CD3 Ab plus anti-CD28 Ab (red box) or with anti-CD3 Ab alone (blue box). Box plot with scatter plots representing means and standard deviations of Ca²⁺ influx response normalized by maximal Ca²⁺ influx response to ionomycin. Statistical significance of differences between anti-CD3/anti-CD28 Ab or anti-CD3 Ab stimulation at concentration 0.05 μg/ml of anti-CD3 Ab CD31⁺ between groups of different T cell subsets was determined by two-tailed ANOVA <i>P</i> = 0.7745, CD45RA⁺ <i>P</i> = 0.8195, CD4⁺ <i>P</i> = 0.9926. ns = not significant. n = number of individuals. (E) CD4⁺CD45RA⁺CD31⁺ T cells of CB (filled line), and adults (dashed line) treated with 0.5 μg/ml soluble anti-CD3 Ab and anti-CD28 Ab cross-linked with GAMIg in the presence (red) or absence (black) of 2 mM EGTA. One representative experiment out of three comparable experiments is shown. (F) STIM1 protein expression in CD4⁺ T cells of CB (black) and in naive CD31⁺ T cells from adults (gray) after stimulation using anti-CD3/anti-CD28 Ab. The densitometric analyses of the ratio of STIM1/α Tubulin are shown. Results are representative of at least two experiments.</p