KSHV Replication and Inhibition Assay

BCBL-1 cells were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS, Gibco-BRL, Gaithersburg, MD, USA). For the lytic replication of KSHV, BCBL-1 cells were induced with 20 ng/mL 12-O-tetradecanoyl-phorbol-13-acetate (PMA/TPA) (Beyotime Institute of Biotechnology, Wuhan, China). Human iSLK.219 cells, derived from iSLK cells, were latently infected with a recombinant rKSHV.219 virus and also contained a doxycycline-inducible RTA [26 (link)]. The rKSHV.219 expresses the red fluorescent protein (RFP) from the KSHV lytic PAN promoter, the green fluorescent protein (GFP) from the EF-1α promoter [27 (link)]. Human iSLK.219 cells were cultured in DMEM supplemented with 10% FBS. The cells were reactivated with 1 µg/mL doxycycline (Dox) and 1.2 mM sodium butyrate (NaB). The chemicals used for cell culture were purchased from Invivogen (San Diego, CA, USA). Celecoxib was purchased from Sigma-Aldrich (St. Louis, MO, USA), NS-398, SP600125 and SB203580 were purchased from Beyotime Institute of Biotechnology, and nimesulide and cidofovir (CDV) were obtained from Selleck Chemicals (Shanghai, China). All drugs were dissolved in dimethylsulfoxide (DMSO).