Bisphenol a downregulates GLUT4 expression by activating aryl hydrocarbon receptor to exacerbate polycystic ovary syndrome

Abstract

Background: Bisphenol A (BPA) levels are high in women with polycystic ovary syndrome (PCOS). The mechanism by which BPA induces abnormal glucose metabolism in PCOS patients is largely unknown.

Methods: Serum and urine samples were collected from women with and without PCOS (control) at the reproductive medicine center with informed consent. Non-PCOS patients who received in vitro fertilization were recruited for collection of ovarian follicular fluid and granular cells. Wild-type C57BL/6 and AhR ^-/- mice were used to verify the effects of BPA on PCOS. Real-time PCR, western blotting, and ELISA were conducted to analyze the function of BPA. Chip-qPCR verified the role of AhR in GLUT4 transcription. Flow cytometry was performed to determine glucose uptake.

Results: A positive correlation was observed between BPA concentration and serum BPA levels in PCOS patients. BPA aggravated the changes in PCOS with abnormal glucose metabolism, impaired fertility, and increased body fat. Mechanistically, we showed that BPA activated AhR and led to decreased glucose transport via GLUT4 downregulation in ovarian granular cells. Therefore, the use of inhibitors or knockout of AhR could effectively rescue BPA-induced metabolic disorders in PCOS mice.

Conclusions: Our results revealed that BPA suppressed GLUT4 expression and induced abnormal glucose metabolism by activating AhR, causing insulin resistance, and is thus a potential contributor to the development of PCOS. Therefore, AhR could be a potential new therapeutic target for PCOS. Video Abstract.

Keywords: Aryl hydrocarbon receptor; Bisphenol a; Glucose transporter 4; Ovarian granulosa cells; Polycystic ovary syndrome.

Fig. 1

A Urine and B serum BPA levels between two groups

Fig. 2

After administration with various concentrations of BPA for 21 days, A-C the hormones were detected. D The number of pups was monitored. Data are presented as mean ± SEM. * P < 0.05 and *** P < 0.001

Fig. 3

Female mice were randomly divided into four groups: CON (oil), BPA, DHEA, BPA + DHEA (n = 10 of each group). During the treatment for 21 days, body weight of mice was measured every day (A). B Body weight gain, C body fat, D body fat ratio, E body lean and F body lean ratio. G Blood glucose and H insulin levels for indicated treatment. I, J Oral glucose tolerance tests (OGTTs) (K, L) and insulin tolerance tests (ITTs) were performed on mice of the four groups. M The homeostasis model assessment of insulin resistance (HOMA-IR) index was calculated of four groups. Data are represented as the mean ± SEM. One-way ANOVA with a Tukey post hoc test. * P < 0.05; ** P < 0.01, and *** P < 0.001

Fig. 4

A Representative estrous cycles were determined after another 10 days. E, estrus; P, proestrus; D, diestrus; M, metestrus. B Representative ovary sections were stained with H&E (scale bar = 500 μm). The serum levels of C estrogen, D testosterone, and E LH/FSH ratio. Data are presented as mean ± SEM. P values were calculated using one-way ANOVA, * P < 0.05; *** P < 0.001

Fig. 5

A Cell vitality was observed with CCK8. B Elevated AhR, Cyp1a1 and Cyp1b1 expression were confirmed by qPCR. KGN cells were treated with PBS, BPA (10 μmol/L), CH223191(10 μmol/L) or BPA/CH223191 for 24 h, and the resulting effects on AhR location were determined by (C) western blots and (D) immunofluorescence. E Cell vitality and (F) glucose uptake was analysed. Values are presented as mean ± SEM; *p < 0.05; **p < 0.01; ***p < 0.001

Fig. 6

A 2D and 3D interactions between BPA and AhR. After administration with various concentrations of BPA for 21 days, B Oral glucose tolerance tests (OGTTs) (C) and insulin tolerance tests (ITTs) were performed on mice of the four groups. D Glucose uptake was evaluated by measuring 2-NBDG uptake using flow cytometry (FACS). E Western blots showing the expression of GLUT4 after the indicated treatment. F ChIP-qPCR analysis was performed with an anti-AhR antibody and GLUT4 promotor-specific primers. Data are presented as values relative to the control group value. G mRNA expression was determined by qPCR. Protein expression was determined by western blotting. The data shown are representative of three independent experiments, and the error bars represent the mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant

Fig. 7

Schematic diagram. Schematic model indicating at which the BPA will disturb ovarian granular cells glycometabolism exacerbating PCOS. BPA suppressed GLUT4 expression and induced abnormal glucose metabolism by activating AhR, causing insulin resistance, and is thus a potential contributor to the development of PCOS