Research Paper Volume 11, Issue 22 pp 10532—10556

Figure 5. Analysis of CS-induced DNA damage in ovarian somatic cells in vitro. (A) Representative IF of cells stained with γH₂AX (red) after 16, 36 and 72 hrs of treatment with 10 μM CS. (B) The graph reports the quantification of γH2AX positive cells percentage. Data are shown as mean ± SEM of three experiments. Statistical differences vs control **p<0.01 ***p<0.001 ****p<0.0001; CS 48 hrs vs CS 72 hrs d = p<0.0001. Note that cells showed a progressive increase in percent of positive cells that peaked after 16 hrs followed by a marked decrease at 72 hrs. (C–F) Representative double IF for γH2AX (green) and FOXL2 (red) in (C) scattered fibroblastoid and epithelioid cells, (D) a large epithelioid colony (D’–D’’’ higher magnifications from D), (E) a little colony of small epithelioid cells and (F) GCs from isolated secondary follicles after CS-treatment for 24 hrs. Scale bar = 100μm. (G) Representative IF of cells stained with MLH1 (green) after 16, 24, 36, 48 and 72 hrs of treatment with 10 μM CS. Scale bar = 100 μm (H) The graph reports the Mean Correlated Total Cell Fluorescence (CTCF) in ovarian cells treated with/out CS as indicated. The fluorescence intensity was determined in each cell by ImageJ software. Data are expressed as means ± SEM of three experiments. Statistical differences vs control ***p<0.001 ****p<0.0001; CS 16 hrs vs CS 24 hrs a = p<0.05; CS 36 hrs vs CS 48 hrs d = p<0.0001; CS 48 hrs vs CS 72 hrs d = p<0.0001. Note that cells showed a progressive increase in the nuclear expression of MLH1.