Fig. 3: P4HB maintains GSCs stemness via the Wnt/β-Catenin signaling pathway.

A Relative mRNA expression levels of the indicated signaling pathways in 83 GSCs infected with shCtrl or shP4HB-1 and shP4HB-2 lentiviral vectors. Data are presented as means ± SD, n = 3, t-test. B–D Western blot analyses of P4HB, β-catenin, and Cyclin D1 protein levels in X01, 448, 83 GSCs infected with shCtrl or shP4HB-1 and shP4HB-2 lentivirus, with α-tubulin used as a loading control. E Western blot analysis of P4HB and β-catenin in isolated nuclear or cytosolic lysates from 83 GSCs infected with shCtrl or shP4HB-1 and shP4HB-2 lentivirus. Lamin B1 and β-actin were used as markers for the nucleus and cytoplasm, respectively. F Heatmap analysis of different gene expression clusters in X01 shCtrl and shP4HB GSCs by RNAseq analysis. G Differentially expressed genes were mapped by fold change between the shCtrl and shP4HB groups. H Western blot analysis of P4HB and LRP6 protein levels in X01 GSCs infected with shCtrl or shP4HB-1 and shP4HB-2 lentivirus, Vinculin was used as a loading control. I Western blot analysis of β-catenin, CyclinD1, and LRP6, P-LRP6 protein levels 3 in X01 and 83 GSCs treated with 10 μM ICG001, 25 ng/ml recombinant P4HB protein (rP4HB). GAPDH was used as a loading control.