Fig. 2

PBK knockdown with lentiviral shRNAs led to reduction in viability and sphere forming capacity in GICs. a Western blot analyses demonstrated down-regulation of PBK protein level in T65, T08 and T59 shRNAs 1, 2 and 3 cell lines compared to non-silencing cells and the original tumor. ACTB was used as a control. b Immunofluorescence analysis of cryosections of GIC spheres transduced with shRNA 2 and 3 compared to the control, shows the loss of PBK protein in the cells present in the spheres. This loss of PBK directly correlated to the loss of GFP. Scale bar = 20 μm. c Quantitative analyses of PBK knockdown cell lines of different GICs showed a significant decrease (n = 5 in each assessment) in viability of knockdown cells compared to the non-silencing controls. d, e Analysis of sphere formation capacity - Sphere numbers and sphere areas of the PBK knockdown cell lines were also diminished as compared to the non-silencing controls. PBK knockdown led to an efficient decrease in the sphere formation capacity of GICs. Error bars = SD; n = 5. Asterisks correspond to p values and indicate level of significance: * = (p ≈ 0.01-0.05), ** = (p ≈ 0.001-0.01) and **** = (p < 0.0001)