Figure 7

CLL-BC depends on STAT3 activity for their growth. (a) Immunoblot analysis of the indicated proteins in shRNA/GFP⁺ Raji B cells at day 8 post transduction. shL, shS3 and shS5 refer to control luciferase, STAT3 and STAT5 shRNA, respectively. Numbers at the bottom of each panel indicate STAT3/ β-actin and STAT5/β-actin ratio relative to shL cells, respectively. (b) FCM of pSTAT3Ser727 expression in CLL-BC at day 8 post transduction with the indicated shRNA/GFP encoding lentiviral vectors. B cells were co-cultured with MS5 stromal cells since day 1 post transduction. Vertical bars delineate specific fluorescent signals compared with control isotype immunoglobulins (ctl Ig, left panel). Results are expressed as mean fluorescence intensity (arbitrary unit, a.u.). Percentages of pSTAT3Ser₇₂₇/GFP⁺ cells are indicated inside each graph. (c–e) N-BC and CLL B cells were transduced with the indicated shRNA/GFP encoding lentiviral vectors. Apoptosis was analyzed at day 8 post transduction, on GFP⁺ and GFP^- B cells by FCM using Annexin V/7-AAD and TMRM/TOPRO3 labeling, as indicated. (c) Representative experiment (n=10), numbers indicate the percentages of cells in relevant quadrants. (d and e) Statistical analyses expressed as percentages of AnnexinV positive (d) and TMRM-probed MTP positive (MTP⁺) (e) cells. UT indicates untransduced cells (mean±S.E.M., CLL-BC, n=10, N-BC, n=5; **P<0.01; *P<0.05)