Effects of methylene blue on ovarian antioxidant capacity, histopathology, and antioxidant-related gene expression in a letrozole-induced PCOS-like ovarian phenotype in mice

Abstract Background/aim Polycystic ovary syndrome (PCOS) is a common reproductive endocrine condition affecting women of reproductive age. In PCOS, ovarian granulosa cells undergo excessive oxidative stress, which results in aberrant follicular development and irregular ovulation. Recent work suggested that methylene blue (MB) has antioxidant effects in a variety of disorders. Therefore, in this study, we investigated the effect of MB on antioxidant capacity, ovarian histopathological alterations, and the expression of antioxidant-related genes in the ovaries of mice with a letrozole-induced PCOS-like ovarian phenotype. Materials and methods In this study, we established a letrozole-induced PCOS-like ovarian phenotype in mice using letrozole. Thirty mice were randomly assigned to three groups of 10: a control group, a letrozole-induced PCOS-like ovarian phenotype group, and an MB-treated group. Mice in the MB group received an intraperitoneal injection of a 0.5 mg/kg dose of MB every day for 2 weeks. Total antioxidant capacity was measured in the ovaries. Histopathologic morphology of the ovaries was also observed. Finally, we used RT-qPCR to detect the expression of Nrf2 and FOXO3a. Results Ovarian TAC levels were significantly lower in the letrozole-treated group than in the control group ( P < 0.0001), whereas MB treatment significantly increased TAC compared with the letrozole-induced PCOS-like ovarian phenotype group ( P < 0.001). MB treatment was associated with a partial normalization of estrous-stage distribution and ovarian histopathological features. In addition, MB significantly increased the mRNA expression of Nrf2 and FOXO3a compared with the letrozole-induced PCOS-like ovarian phenotype group ( P < 0.0001 and P < 0.01, respectively). Conclusion These findings suggest that MB increases ovarian total antioxidant capacity and is associated with partial improvement in ovarian morphology and estrous cyclicity in mice with a letrozole-induced PCOS-like ovarian phenotype. However, because specific oxidative damage markers were not assessed, the findings do not establish a direct reduction in oxidative damage. The increased mRNA expression of Nrf2 and FOXO3a may reflect modulation of redox-responsive pathways; however, pathway activation could not be confirmed at the protein level.