Fig. 4: EIF4G2KD increases CD133 and CD144 expression and viability of CD133+ and CD44+ populations following therapy in HEC-1A and RL95-2 cells.

Flow cytometry analysis of cell-surface expression of CD133 (A) and CD44 (B, C) in control and EIF4G2KD HEC-1A cells (A, B) and control and EIF4G2KD RL95-2 cells (C). Quantification of results is presented as mean ± SEM, n > 5 for both overall mean florescent intensity (MFI) and as percent of total population. P-vals were calculated using two-tailed t-test (*p < 0.05, **p < 0.01, ****p < 0.0001). A representative flow cytometry plot is shown for each. D Control and EIF4G2KD HEC-1A and RL95-2 cells were separated for CD133 and CD44 expression, respectively. Separated populations were treated with 2.5 nM Taxol or DMSO as control for 4 d and cell viability assayed by CellTiter-Glo. Luminescence was normalized to DMSO treated cells. Left graph, HEC-1A cells, right graph RL95-2 cells. E Cell viability of HEC-1A (left) and RL95-2 (right) CD133+ and CD133− or CD44+ and CD44- control and EIF4G2KD separated populations 4 d after exposure to 16 Gy (HEC-1A) or 8 Gy (RL95-2) X-ray irradiation. For D, E, results are presented as mean values ± SEM of n > 4 independent experiments. Significance was determined by two-way ANOVA. (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001). Total cell lysates form HEC-1A (F) and RL95-2 (G) CD133+ and CD133- or CD44+ and CD44- control and EIF4G2KD separated populations were subjected to western blot analysis for γH2AX and GAPDH, as loading control 4 d following irradiation. γH2AX signal was normalizedC to GAPDH and quantification results are presented as individual data points and also as mean values of 4 independent experiments. Statistical significance was determined for log₂ transformed mean values by two-way ANOVA (*p < 0.05).