Apoptosis induction and PI3K inhibition

Apoptosis was induced in LCLs of different B-cell donors 2–3 months after infection. Cells were treated with etoposide (4μg/ml, Sigma), staurosporine (4μg/ml, Sigma) or a DMSO solvent control for 20 hrs. We also used simvastatin (2mM, Calbiochem) or an ethanol solvent control for 5 days [46]. After treatment, cells were subjected to western blotting (PARP) or Annexin-V viability staining. We used a 10nM concentration of 17β-hydroxy wortmannin (Cayman Chemical) to inhibit the PI3K in EBV wild type-infected primary B lymphocytes 14 days post-infection. Cells were treated for 17 hrs and the cell cycle profile was determined using a BrdU incorporation assay as described above.