Reduced p-PLCγ2 and β-catenin expression levels are observed in MDSCs during tumor progression in mice and humans. (A) Western blot analyses show p-PLCγ2 and β-catenin protein levels in MDSCs from tumor-free (No Tumor) and LLC or B16 tumor-bearing C57BL/6 WT mice. β-actin was used as loading control. Mean ± SD of three tumor-free mice (No Tumor) and three tumor-bearing mice (Tumor). **, P < 0.01; ***, P < 0.001. (B) 10⁴ B16-Fl cells were i.v. injected in WT mice to allow tumor dissemination to bone. When bone metastases (Met.) were established, animals were sacrificed and bone marrow and spleen were analyzed by FACS using anti–Gr-1 and -CD11b antibodies to measure the percentage of MDSCs. Mice receiving saline i.v. injection were used as controls (No Tumor). Mean ± SD (n = 4) is shown. Data are reported from one of two similar independent experiments. *, P < 0.05. Western blot analysis of p-PLCγ2, β-catenin and β-actin (loading control) levels in MDSCs from spleen of mice with bone metastases (Met.) or tumor-free controls (No Tumor). One representative Western blot from three different mouse samples is shown. (C) MDSCs isolated from healthy donors (HD) or pancreatic cancer patients (Pt) were analyzed for β-catenin and phosphorylation levels of PLCγ2, PKC, and GSK3β. β-actin was used as loading control. Graph shows semiquantitative analysis of protein levels from Western blots of all samples after normalization to total protein loaded into the gel (measured by β-actin). Three representative patients out of five are shown. *, P < 0.05.