Double deletion of PDK2/4 inhibits intracellular Ca²⁺ concentration levels in PMA-stimulated neutrophils. (A) The intracellular Ca²⁺ concentration was measured in Fura-2AM–loaded WT or PDK2/4^−/− neutrophils that were stimulated with PMA (2 nM) in the presence of 1.3 mM CaCl₂. The representative curve for intracellular calcium Ca²⁺ concentration after stimulation with PMA is shown. The bar graphs show the peak Ca²⁺ level and the area under the curve (AUC). Values are mean ± SEM, n = 6 mice per group. Statistical analysis was performed using Mann-Whitney U test. (B) The Erk1/2 phosphorylation was measured in resting and PMA (50 nM)–stimulated WT and PDK2/4^−/− neutrophils. A representative western blot for phosphorylated Erk1/2 is shown. Total Erk1/2 was used as a loading control. The bar graphs show densitometry analysis of immunoblots. Values are mean ± SEM, n = 5 mice per group. Statistical analysis was performed using 2-way ANOVA followed by Tukey multiple comparisons test.