fig6

Figure 6. RCC1 knockdown enhances DNA damage and impairs DDR in drug-resistant CRC cells. (A) Western blot analysis of γ-H2AX expression in drug-resistant CRC cells treated with 38.43 μM 5-FU or 500 nM Doxo for 48 h, with or without RCC1 knockdown for 48 h; (B) Representative comet assay images of drug-resistant CRC cells treated with the indicated drugs for 24 h. Scale bar: 100 μm; (C and D) Quantification of DNA damage by comet assay, including percentage of DNA in tail (C) and tail moment (D) across treatment groups (one-way ANOVA); (E) Schematic of the experimental design for monitoring DNA damage repair dynamics via γ-H2AX immunofluorescence at 0, 6, 12, and 24 h post-drug withdrawal; (F and G) Representative immunofluorescence images showing time-dependent changes in γH2AX-positive cells after treatment with 7.68 μM 5-FU (F) or 500 nM Doxo (G) in drug-resistant CRC cells with or without RCC1 knockdown. Scale bar: 50 μm; (H) Quantification of γH2AX-positive cells at 24 h after drug removal in the indicated groups (two-way ANOVA). ns, not significant; ^***P < 0.001; ^**P < 0.01; ^*P < 0.05. RCC1: Regulator of chromosome condensation 1; DDR: DNA damage response; CRC: colorectal cancer; 5-FU: 5-fluorouracil; Doxo: doxorubicin; ANOVA: analysis of variance.