Bzb increases free and active forms of β-catenin in mouse and human OB progenitor cells (OB) and primary human MSCs. OB lines C2C12, C3H10T1/2, Saos-2, and MG63 and primary MSCs from 2 healthy donors (designated as #hMSC1 and #hMSC2) and 2 patients with MM (designated P#1 and P#2) were grown in the absence or presence of 500 nM of Bzb for indicated times (A,F,G) or with indicated serial concentrations of Bzb (B-D) for 6 hours. Saos-2 and MG63 cells were cultured in the absence or presence of 500 nM MG132 for indicated times (E). MSCs from 6 patients with MM (designated P#3 to P#8 in panel H) were grown in the absence or presence of 500 nM Bzb for 5 hours. Protein lysates (500 mg) subjected to GST-E-cadherin pull down were immunoblotted using antibodies to β-catenin (A-C,E-H) or active (nonphosphorylated) β-catenin (C). Arrows indicate ubiquitinated β-catenin. (D) The images of antiactive β-catenin antibody staining in C2C12, MG63, and Saos-2 lysates were analyzed with Photoshop software and quantified by National Institutes of Health Image 1.16 software as described in “Immunoblotting.” Results are mean plus or minus SD (n = 3). **P < .001, ***P < .001, ***P < .001 vs controls. (I) The MSCs from 2 patients with MM were treated with indicated concentrations of Bzb for 6 hours. Free β-catenin protein in the lysates separated using GST-E-cadherin pull down was determined by immunoblotting analysis as described in Figure 1A.