Induction of PRDM1 following treatment with IL-21 in CLL cells correlates with anergy. (A) Basal (ie, unstimulated) levels of PRDM1 protein in M-CLL and U-CLL samples (n = 5 each). A PCL sample was used as a positive control. n.s.: nonspecific band detected by the PRDM1 antibody. β-actin was analyzed as a loading control. (B) PRDM1 induction in representative “responsive” (R) and “non-responsive” (NR) CLL samples after 2 days in culture with the indicated concentrations of IL-21 (n = 3). (C) Correlation of PRDM1α protein induction in cells treated with 50 ng/mL IL-21 for 16 hours. The maximum percentage of cells that showed increased iCa²⁺ following anti-IgM stimulation: (i) shows linear correlation between these variables, and (ii) shows significant statistical difference in iCa²⁺ responsiveness between PRDM1 NR (≤twofold induction) and R (>twofold induction) CLL samples (n = 26). Fold induction of PRDM1α was calculated as a ratio by western blot densitometry between the IL-21–treated and untreated samples. The results of statistical evaluation are shown, (i) Pearson’s coefficient, and (ii) Mann-Whitney U test.