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. 2022 Mar 18;28(10):2147–2159. doi: 10.1158/1078-0432.CCR-22-0100

Figure 4.

Figure 4. Effects of CHEK1 silencing and prexasertib. A, The relative expression of CHEK1 and CHEK2 in paired ULMS and myometrium. ACTB was used as a reference gene to normalize expression, and the relative expression was compared using the paired t test. B, Validation of CHEK1 or CHEK2 suppression following transfection with 3 nmol/L of siRNA for CHEK1 (siCHEK1) or CHEK2 (siCHEK2). C, The proliferation of siCHEK1 transfected cells. Cell viability was measured at 24, 48, and 72 hours, and luminescence was compared using the Dunnett's test. Experiments were performed in triplicate and repeated three times. D, The expression of CHEK1 and pCHEK1(Ser296) protein in prexasertib-treated cells. Cells were treated with each concentration of prexasertib for 16 hours. E, Immunofluorescent of γH2AX in prexasertib-treated cells. Cells were treated with 0 or 100 nmol/L prexasertib for 24 hours. The experiments were performed in three independent replicates and the percentage of γH2AX-positive cells was compared using the Welch's t test. The green and blue colors indicate γH2AX and Hoechst, respectively, and the scale bars, 10 µm. F, The combination effect of prexasertib and cisplatin. Cells were treated with each drug concentration for 72 hours, and the percentage of growth inhibition is shown relative to untreated controls. Experiments were performed in triplicate and repeated three times. Drug synergy was analyzed using CompuSyn software. Error bars represent standard errors of the mean, *, P < 0.05; **, P < 0.01; and ***, P < 0.001.

Effects of CHEK1 silencing and prexasertib. A, The relative expression of CHEK1 and CHEK2 in paired ULMS and myometrium. ACTB was used as a reference gene to normalize expression, and the relative expression was compared using the paired t test. B, Validation of CHEK1 or CHEK2 suppression following transfection with 3 nmol/L of siRNA for CHEK1 (siCHEK1) or CHEK2 (siCHEK2). C, The proliferation of siCHEK1-transfected cells. Cell viability was measured at 24, 48, and 72 hours, and luminescence was compared using the Dunnett's test. Experiments were performed in triplicate and repeated three times. D, The expression of CHEK1 and pCHEK1(Ser296) protein in prexasertib-treated cells. Cells were treated with each concentration of prexasertib for 16 hours. E, Immunofluorescent of γH2AX in prexasertib-treated cells. Cells were treated with 0 or 100 nmol/L prexasertib for 24 hours. The experiments were performed in three independent replicates and the percentage of γH2AX-positive cells was compared using the Welch's t test. The green and blue colors indicate γH2AX and Hoechst, respectively; scale bars, 10 µm. F, The combination effect of prexasertib and cisplatin. Cells were treated with each drug concentration for 72 hours, and the percentage of growth inhibition is shown relative to untreated controls. Experiments were performed in triplicate and repeated three times. Drug synergy was analyzed using CompuSyn software. Error bars represent standard errors of the mean, *, P < 0.05; **, P < 0.01; ***, P < 0.001.