Figure 4

xCT inhibition induces intracellular ROS levels through upregulation of NADPH oxidases activities from KSHV-infected PEL cells. (A) BCP-1 and BCBL-1 were incubated with either MSG or SASP at the indicated concentrations for 24 h, then intracellular reactive oxygen species (ROS) were quantified using the ROS-specific dye CM-H2DCFDA and flow cytometry, and normalized to ROS levels for the vehicle-incubated cells. NF-κB inhibitor Bay11-7082 was used as a positive control. (B-C) BCP-1 and BCBL-1 were treated with either MSG (20 mM), SASP (0.5 mM) or vehicle for 24 h, then proteins expression were measured by immuoblots. NADPH oxidases activities were measured as described in Methods. (D) BCBL-1 were treated with either MSG (20 mM), SASP (0.5 mM) or vehicle in the presence or absence of the antioxidant N-acetylcysteine (NAC) 2.5 mM for 48 h, then cell apoptosis was assessed using Annexin V-PI staining and flow cytometry analysis (** vs MSG group; ## vs SASP group). Error bars represent the S.E.M. for 3 independent experiments. * = p < 0.05; **/## = p < 0.01.