Fig. 2: Mettl3^Gdf9 cKO mice display follicle developmental defects and POF.

A Representative ovarian histology of 6-week-old, 8-week-old WT, and Mettl3^Gdf9 cKO mice. The primary, secondary, and antral (indicated with PF, SF, and AF, respectively) follicle stages are indicated by the black arrow. Scale bar, 50 μm. B Quantification of the numbers of different types of follicles in ovaries from 6-week-old, 8-week-old WT, and Mettl3^Gdf9 cKO mice. Primordial, primary, secondary, and antral follicles (indicated with PrF, PF, SF, and AF, respectively) were counted. n.s., p > 0.05; *p < 0.05; **p < 0.01 by two-tailed Student’s t-test. Data represent the mean ± SEM (n = 3). C Confocal immunofluorescence staining with γH2AX antibody (red) and DAPI (blue) in ovary sections from 6-week-old WT and Mettl3^Gdf9 cKO mouse ovaries. Scale bars, 20 μm. D Graph showing the quantification of γ-H2AX staining. ***p < 0.001 by two-tailed Student’s t-test. Data represent the mean ± SEM (n = 4). E Detection of apoptosis in granulosa cells by TUNEL kit performed in paraffin sections at different stages of follicle development of 6-week-old WT and Mettl3^Gdf9 cKO mouse ovaries. Secondary and antral follicles (indicated with SF and AF, respectively) were counted. Scale bars, 20 μm. F Graph showing the quantification of the number of TUNEL positive granulosa cells at different follicle stages. *p < 0.05; **p < 0.01 by two-tailed Student’s t-test. Data represent the mean ± SEM (n = 3). G, H FSH and LH levels of WT and Mettl3^Gdf9 cKO mice. Serum samples were collected from female mice of both genotypes (12 to 20 weeks old) and were killed for measurement of FSH and LH levels. FSH: WT, n = 5; Mettl3^Gdf9 cKO, n = 9. LH: WT, n = 8; Mettl3^Gdf9 cKO, n = 9. **p < 0.01; ***p < 0.001 by two-tailed Student’s t-test. Data represent the mean ± SEM. I Immunohistochemistry with MVH antibody in ovary sections from 24-week-old WT and Mettl3^Gdf9 cKO mice. Scale bar, 50 μm.