Fig 2: Comparison of clinical manifestations at different doses of 3-NPA. (A, B) 3-NPA had no significant effects on body weight. (C) 3-NPA could prolong the mean length of the diestrus phase and decrease the mean length of the estrous phase after two weeks of treatment, especially when used at a dose of 50 mg/kg. (D) 3-NPA could prolong the mean length of the diestrus phase and decrease the mean length of the estrous phase after three weeks of treatment, especially when used at a dose of 40 mg/kg and 50 mg/kg. (E, F) 3-NPA reduced the ovarian index, especially when used at a dose of 40 mg/kg and 50 mg/kg. (G, H) Serum E2 concentration in different groups; mice treated with 40 mg/kg 3-NPA for 3 weeks and those treated with 50 mg/kg 3-NPA for 2 or 3 weeks showed a significant decrease. (I, J) Serum FSH concentration in the different groups; mice treated with 40 mg/kg or 50 mg/kg 3-NPA for 3 weeks showed a significant increase. (K, L) Serum AMH concentration in different groups; mice treated with 40 mg/kg 3-NPA for 3 weeks and those treated with 50 mg/kg 3-NPA for 2 or 3 weeks showed a significant decrease. (M) In the mice treated with 50 mg/kg 3-NPA for 2 weeks, the number of primordial follicles decreased. (N) In the mice treated with 40 mg/kg or 50 mg/kg 3-NPA for 3 weeks, the number of primordial follicles decreased, the number of atretic follicles increased, and the number of growing follicles did not change significantly. (N=8 in all assays; *P<0.05, **P<0.01, and ***P<0.001 compared with the control group, NS: none significant).

Fig 3: 40 mg/kg 3-NPA significantly induced manifestations similar to those of POI. (A) 40 mg/kg 3-NPA could prolong the mean length of the diestrus phase and decrease the mean length of the estrous phase after three weeks of treatment. (B) 40 mg/kg 3-NPA could decrease the number of primordial follicles and increase that of atretic follicles. (C–F) 40mg/kg 3-NPA decreased the ovarian index (C), increased serum FSH levels (D), decreased serum E2 levels (E), decreased serum AMH levels (F), and decreased mouse fertility (G). (H) Although the 40 mg/kg 3-NPA group had fewer pups, the difference was not significant. (I, J) The offspring of the mice treated with 40 mg/kg 3-NPA showed lower weight than the control group (N=8 in all assays; *P<0.05, **P<0.01).