Fig. 5

PD-SK induces stronger apoptosis in Colo320 cells than PD-H. (A) Determination of CVB3 VP1 and cellular proteins involved in apoptosis induction. Left images, Colo320 cells were infected with 0.1 MOI of PD-SK or PD-H and analyzed 24 h later for expression of eIF4G, cleaved eIF4G, PARP, cleaved PARP and cleaved Caspase 3, Caspase 9, cleaved caspase 9, caspase 8 and cleaved caspase 8 by western blotting. The γ-tubulin was used as internal loading control. Right diagrams: Quantification of the expression of indicated proteins was carried out relative to the expression of γ-tubulin by densitometric analysis using the ImageJ densitometry software. (B) Immunofluorescence staining of Colo320 cells for cleaved caspase 3. Cells were seeded in 24 well plates and infected with PD-H and PD-SK at MOI 0.1. Twenty-four h after infection cells where fixed with 4% formaldehyde and stained for cleaved caspase 3. Scale bar 100 μm. (C) Dapi staining of Colo320 cells. Cells were treated as in B and stained with Dapi. Left diagram, relative amount of living and apoptotic cells of 100 random counted cells. Right, images after staining with Dapi. Arrows indicate apoptotic cells. Scale bar 20 μm. (D) Relative caspase 3/7 activity in Colo320 cells 24 h after infection with PD-H, PD-K2A, PD-K2B, PD-K2AB and PD-SK at indicated MOIs. Shown are mean values ± SEM. Significance compared to PD-H, * p < 0.05; ** p < 0.01. Significance compared to PD-K2B: # p < 0.05