Fig. 4.

Colocalization of PKC-θ and NF-κB in T cells. (A) Representative immunoblots of PKC-θ, p50 and p65 protein levels in the nuclear extracts of activated Jurkat T cells (0, 2, 4 h) during primary (1°), and secondary (2°) stimulation with Sp1 as the loading control (representative graph of three independent repeats). The corresponding p65 and PKC-θ (normalized to Sp-1) densitometry is shown below (mean±s.e.m., n=3). *P≤0.05; ns, not significant (two-way ANOVA). (B) NF-κB (p65) activity detected in nuclear extracts from Jurkat T cells during day 0, primary, and day 9, secondary, activation (0, 2 and 4 h) with PMA and Ca²⁺ ionophore (+P/I). Representative graph of three independent replicates, error bars show s.e.m. of technical replicates. (C) The percentage of CD45RO⁻ (naïve) and CD45RO⁺ (memory) ex-vivo-derived human CD4⁺ T cells with p50 and p65 staining as detected by using flow cytometry with (+P/I) or without (NS) PMA and Ca²⁺ ionophore (median±quartiles, n=6). The whiskers represent the first and fourth quartiles. **P≤0.01 (Wilcoxon matched-pairs signed rank test was used to compare groups). (D) p50 and p65 staining levels were calculated as fold-changes between human CD45RO⁻ (naïve) and CD45RO⁺ (memory) T cells with (+P/I) or without (NS) PMA and Ca²⁺ ionophore (mean±s.e.m., n=6).