Activation of the canonical Wnt signaling in SP cells from DLBCL cell lines. (A) Increased expression of Wnt3a as well as reduced expression of SFRP4 was found in the SP vs non-SP cells of OCI Ly3 cells by quantitative reverse-transcription PCR (normalized to GAPDH; n = 10 OCI Ly3, n = 5 U2932, n = 3 SUDLH4; unpaired Student t test). (B) Western blot showed expression of Wnt3a and SFRP4 in all lymphoma cell lines tested, with variable expression levels. (C) Increased levels of Wnt3a and β-catenin vs lower levels of SFRP4 in whole-cell lysates of SP cells compared with non-SP cells of OCI Ly3. Densitometric quantification (right) of Wnt3a levels in SP and non-SP cells of OCI Ly3 (n = 4, unpaired Student t test, P = .0047). (D) Cellular β-catenin in OCI Ly3 and OCI Ly1 cells was enhanced by exogenic stimulation with recombinant Wnt3a (concentration as indicated for 8 hours). (E) β-catenin localized to the nuclei of both SP and non-SP cells from OCI Ly3, with higher frequency in SP cells (F; n = 200, OCI Ly3, P < .0001; paired 2-sided Student t test). (G) C-myc and axin2 were expressed at higher levels in SP compared with non-SP cells in OCI Ly3 (axin2: n = 5, P < .001; c-myc: n = 4, P < .01; 2-way ANOVA).