Fig. 5
ALPL deficiency caused an imbalance in lineage differentiation of BMSCs through the Wnt/β-catenin pathway. a alpl+/− and shALP BMSCs showed significant downregulation of p-AKT, p-GSK3β, and active β-catenin compared with what was observed in WT BMSCs. β-actin was used as a protein loading control. b Lenti-alp/alpl+/− BMSCs showed significant upregulation of p-AKT, p-GSK3β, and active β-catenin compared with that of alpl+/− BMSCs. c alpl+/− BMSCs transfected with oeCaV1.2 or oeCaV1.3 showed significant upregulation of p-AKT, p-GSK3β, and active β-catenin. d alpl+/− BMSCs transfected with DN-Dyn1 showed significant upregulation of p-AKT, p-GSK3β, and active β-catenin. Alizarin red staining revealed that alpl+/− BMSCs treated with 10 μmol·L−1 sc79 or 10 mmol·L−1 LiCl had an increased capacity to form mineralized nodules when cultured under osteoinductive conditions (e), and they exhibited upregulation of the osteogenic-related proteins RUNX2 and Sp7 (f). alpl+/− BMSCs treated with sc79 or LiCl showed a decreased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions (g), and they exhibited downregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot (h). Alizarin red staining showed that alpl+/− BMSCs transfected with a plasmid overexpressing β-catenin (oeβ-cat/ALPL+/−) had an increased capacity to form mineralized nodules when cultured under osteoinductive conditions (i), and they exhibited an upregulation in the osteogenic-related proteins RUNX2 and Sp7 (j). alpl+/− BMSCs transfected with a plasmid overexpressing β-catenin (oeβ-cat/ALPL+/−) showed a decreased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions (k), and they exhibited downregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot (l). The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. *P < 0.05; **P < 0.01
