Fig 2: Caspase-induced apoptosis leads to POF degradation. (A-B) WB and grey analysis of caspase3 expression in POFs (POF1 to POF4). Values are means ± SEM of three experiments. Asterisks indicate significant differences (* P<0.05 and ** P<0.01). (B-E) Few TUNEL (green) marker labeled occurred in cells from POF1 to POF3. (D) Negative Control. Scale bar: 50 µm.

Fig 3: Steatosis-like morphology occurring in the granulosa cells from POFs. (A) HE staining was used to observe steatosis-like granulosa cells in POF6. Scale bars: 50 µm and 10 µm. (B) Oil Red O staining was used to verify lipid droplets existing in the coalescence of POF6. Scale bars: 100 µm and 50 µm. (C-E) TEM was used to observe the large quantity of lipid droplets existing in POF3, POF4 and POF6.

Fig 4: Visualization of MAPK3 expressed in ovarian granulosa cells of the PCOS, BSTJF and control groups by double-labeling immunofluorescence with FoxL2 (winged helix for head transcription factor gene2, specific expression in adult ovarian granulosa cells) (green) and MAPK3 (red). Nuclei were counterstained with 40,6-diamidino-2-phenylindole (DAPI) (blue fluorescence). All scale bars are 100 µm. The expression ratio of the target protein is shown by bar plots (n = 4 in each group). Data were presented as the mean ± SEM. One-way ANOVA for multiple comparisons and LSD test for pairwise comparisons between groups

Fig 5: GRP78 and BCL2 co-expressed in the granulosa layer from POFs. (A-C) Histological sections of POFs (POF1 to POF3) were given immunofluorescent labels with ER stress marker GRP78 (Red) and mitochondria anti-apoptosis marker BCL2 (Green), showing the main distribution in the granulosa layer. Scale bar: 20 µm. (D) Negative Control. (E-F) TEM was used to observe ER lesions, concentric round, dilatation and vesiculation and hyperplasia, respectively (arrows). (H-J) TEM was used to observe normal mitochondria and vacuolated mitochondria, respectively (arrowheads). GL represents the granulosa layer.