Autologous T-cell-mediated CLL cell proliferation is dependent on Rac. (A) Flow cytometric analysis of Ki-67-positive CLL cells after anti-CD3/CD28 bead-mediated activation with or without NSC-32766 (50 µM) pretreatment (n = 11). Data are presented in box plot format, whereas the 25th and 75th percentiles form the box, with the median marked as a line; the smallest and the largest observation form whiskers. (B) Ki-67-positive CLL cells after treatment with 10 µM Y-27632 (n = 7). (C) Relative Tiam1 gene expression in a time course experiment (left) and after 24 hours (n = 5) (right means ± SD). Dashed line, NSC-32766 treatment; solid line, control. (D) Relative c-Myc mRNA expression in a time course experiment (left) and after 24 hours (n = 5) (right means ± SD). Dashed line, NSC-32766 treatment; solid line, control. (E) Western blot analysis of Tiam1, Rac1, and c-Myc protein expression in activated and nonactivated CLL cells. Cyclophilin A (Peptidylprolyl isomerase A; PPIA) protein was used for normalization. A representative blot (left) and a densitometric analysis (right) of 3 independent experiments are shown (bars: c-Myc, light gray; Tiam1, dark gray; Rac1, black). (F) Relative mRNA expression of Tiam1 and c-Myc (left) or Rac1 and c-Myc (right) in activated CLL cells after cells were nucleofected with Tiam1-siRNA, Rac1-siRNA, or nontargeting control siRNA. One representative out of 3 independent experiments is shown.