Activation of STAT1 in CLL cells upon lenalidomide or IL-10 treatment. (A) A total of 3 × 10⁷ PBMC from CLL patients were cultured in the presence or absence of 10 µM lenalidomide in 4 mL complete medium in 6-well plates for 3 days. Cell lysates were analyzed by immunoblotting using anti-STAT1, anti-phospho STAT1(Tyr701), and anti-tubulin antibodies. One representative result out of 3 independently performed experiments is shown. (B) A total of 3 × 10⁷ PBMC from CLL patients were cultured in the presence or absence of 50 ng/mL recombinant human (rh-)IL-10 in 4 mL complete medium in 6-well plates for 2 days. Immunoblot analysis was performed as described in panel A. One representative result out of 3 independently performed experiments is shown. (C) Corresponding to Figure 3A: Quantification of protein bands by ImageJ software revealed significantly stronger p-STAT1 signals in lenalidomide-treated samples (P = .037). (D) A total of 1 × 10⁷ CLL PBMC were cultured for 3 days in the presence of 10 µM lenalidomide or 50 ng/mL IL-10 in combination with either 10 µg/mL anti–IL-10 or isotype control antibodies. All supplements were added to the medium at days 0 and 2. After 3 days of culture, cells were harvested and immunoblot analysis was performed as described in panel A.