ETS1 and FLI1 affect proliferation and viability in OCI-Ly7. (A) ETS1 and FLI1 protein levels in 22 DLBCL cell lines. Western blot analysis was carried out on whole-cell lysates with antibodies against ETS1, FLI1, and GAPDH as loading control. (B) ETS1 and FLI1 down-regulation in OCI-Ly7 cell line. Real-time PCR and western blot analysis of ETS1 (left) and FLI1 (right) levels after shRNA (day 5). As a control, β2-microglobulin mRNA and GAPDH protein expression were documented. (C) For the growth curve, OCI-Ly7 cells infected with control (shGFP) lentivirus or a lentivirus expressing shRNA for ETS1 and FLI1 (shETS1 and shFLI1, respectively), were seeded on 24-well plates at a density of 3 × 105 cells/well. Cultures were harvested every day, and the number of cells was determined. The numbers refer to mean values of triplicate determinations. (D) At day 3 of the growth curve, levels of EdU incorporation were determined by fluorescence-activated cell sorter analysis. (E) ETS1 and FLI1 down-regulation causes proliferation impairment. Y-axis: percentage of GFP positive and negative cells. X-axis: days. Histograms are representative of 1 experiment of 2. (F) ETS1 (upper) and FLI1 (lower) down-regulation induces apoptosis. Percentage of apoptotic cells was determined by ANNEXINV staining at day 5. Dot plots are representative of 1 experiment; histogram graphs represent the average of at least 3 independent experiments. *P value < .05.
