Fig. 5

PRV7S can infect and reproduce in human cancer stem cell. (a) Schematic illustration of experiments performed on AML-M5 human induced pluripotent stem cell (hiPSC), also known as human cancer stem cell. AML-M5 hiPSC were seeded into multi-well plates and subsequently infected by PRV7S. Cell viability, caspase activity, viral titre and viral RNA from infected cells were measured over time. Created with BioRender.com (https://BioRender.com/r23t298). (b) Micrographs of non-infected, healthy colonies of AML-M5 hiPSCs seeded on mitomycin C-treated primary mouse embryonic fibroblasts (MEFs). Scale bars: 400 μm (left), 200 μm (right). (c) hiPSCs infected with PRV7S at a multiplicity of infection (MOI) of 0.1 over 7 days. Pronounced cytopathic effects (CPE) and plaques (clear zones) were observed from 3 days post-infection (dpi) onwards. Scale bars: 400 μm. (d) Hoechst 33342 and propidium iodide (PI) staining was performed on negative control hiPSC colonies (left) and PRV7S-infected colonies (right). The PRV7S-infected colony exhibited more intense PI staining compared to the negative control. Scale bars: 100 μm (left and right). (e) Cell viability of PRV7S-infected hiPSCs was assessed using the MTT assay over 5 days. A statistically significant decrease in cell viability was observed from 3 dpi onwards. (f,g) Viral kinetics of PRV7S propagation in hiPSCs were determined using the TCID50 assay (f) and qPCR (g). Viral titre and viral RNA increased from 1 to 5 dpi. (h) Caspase-3/7 activity in PRV7S-infected hiPSCs increased slightly by more than 1.2-fold at 5 dpi. The red dashed line indicates a 1.2-fold change threshold. (e–h) Data are presented as mean ± SD. Three independent experiments were performed, each with four replicates. ANOVA or t-test were conducted. *p < 0.05; ***p < 0.001; ****p < 0.0001.