Fig. 4

The impact of (R)-NAF and (S)-NAF on the reproductive and metabolic characteristics of PCOS mice caused by letrozole and high-fat diet (HFD) (n = 4–8). A Schematic diagram of the experimental protocols for (R)/(S)-NAF in treating letrozole with HFD-induced PCOS mice. B The serum levels of T. C The mRNA expression of UGT2Bs in the ovarian tissue of mice. D Immunohistochemical analysis of the protein expression level of UGT2Bs in ovarian tissues of each group, scale bar = 100 µm. E The histogram shows the H-score of UGT2Bs IHC in each group. D The mRNA expression of UGT2Bs in the ovarian tissue of mice. F Ovarian morphology was represented using hematoxylin and eosin staining, with a scale bar of 300 μm, showing primary follicles (triangle), secondary follicles (square), mature follicles (asterisk), cystic follicles or atretic follicles (asterisk), and corpus leteum (CL). G Representative of estrous cycles. H Percentages of each follicle type per ovary. Different stages of follicles include primordial follicles (PrF), primary follicles (PF), secondary follicles (SF), mature follicles (MF), atretic follicles (AF), and corpus leteum (CL). I Differential HOMA-IR of each group. J-M The serum levels of progesterone, androstenedione, DHT, and E2. N Representative images of ovarian stained by TUNEL assay, cell nuclei were stained by DAPI (blue), and apoptotic cells were stained with TUNEL (green). Scale bar = 100 μm. O Quantification of the proportion of ovarian apoptotic cells as detected by TUNEL assay in each group. The data is displayed as mean ± SD. Statistical analysis was performed using one-way ANOVA with Tukey’s post-hoc test for normally distributed data, or the Kruskal–Wallis test for non-normally distributed or unequal variance data. Significance levels: ^*P < 0.05, ^**P < 0.01, ^***P < 0.001 (vs. vehicle group); ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 (vs. model group); ns, no significant difference (P > 0.05)