Backgroud Beta-cypermethrin and emamectin benzoate are widely used for the prevention and control of pests in the greenhouse planting industry, and their combined exposure may increase the accumulation of beta-cypermethrin and emamectin benzoate in organisms and affect human health.

Objective Based on the changes in reproductive hormone levels in the hypothalamic-pituitary-ovarian (HPO) axis, to investigate the effect of combined exposure to beta-cypermethrin and emamectin benzoate on the estrous cycle of female mice.

Methods Twenty-four healthy adult SD rats were randomly divided into a blank control group, a beta-cypermethrin group (Beta-CYP, 53 mg·m⁻³), an emamectin benzoate group (EMB, 8 mg·m⁻³), and a beta-cypermethrin and emamectin benzoate combined exposure group (Beta-CYP+EMB, Beta-CYP 53 mg·m⁻³ + EMB 8 mg·m⁻³). Six rats in each group were exposed to the designed treatment protocol by aerosol inhalation 6 d a week for 13 weeks. The general condition of the rats was observed in real time during the treatment. From the 12th week of exposure, a 10-day reproductive tract smear was performed on the rats to observe the estrous cycle. The rats were neutralized on the second day after the end of the treatment protocol, and the ovarian tissues were stained with HE to observe histopathological changes. Serum levels of gonadotropin-releasing hormone (GnRH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) were measured by ELISA. Experimental results were expressed as mean ± standard deviation ( \overlinex\pm s ). One-way ANOVA was used for comparison among groups, and LSD test for pairwise comparison between groups. The significance level was α=0.05.

Results After four weeks of the treatment protocol, the rats in the Beta-CYP group and the Beta-CYP+EMB group continued to be hyperactive and irritable, while the EMB group showed symptoms of mental disorder, decreased activity, and slow response. On the 90th day of the treatment protocol, the body weight of rats in the control group increased to (314.51±2.44) g, and that in the Beta-CYP+EMB group only increased to (253.47±1.50) g. There was no abnormal cellular morphology in the control group; however, small deeply stained nuclei appeared in the Beta-CYP group, the EMB group, and the Beta-CYP+EMB group, and abnormal morphological development of keratinized epithelial cells in the Beta-CYP+EMB group was found. The estrous cycle of rats in the control group was (97.83±4.17) h, and compared with the control group, the estrous cycles of rats in the Beta-CYP group, the EMB group, and the Beta-CYP+EMB group were prolonged to (134.33±7.53) h, (126.50±5.28) h, and (156.00±6.66) h, respectively. The results of ANOVA showed that the numbers of leukocytes (527.17±15.83), keratinized epithelial cells (35.67±4.32), and non-keratinized epithelial cells (70.50±4.51) in the vaginal smears during diestrus in the Beta-CYP+EMB group were significantly lower than those in the control group (752.50±28.89, 50.50±2.74, 101.33±7.92) (P<0.001). The hormone levels of GnRH and FSH in the control group were (5.13±0.59) and (0.76±0.09) IU·L⁻¹ respectively, while the levels in the Beta-CYP+EMB group were increased to (16.86±0.59) and (3.80±0.19) IU·L⁻¹ respectively (P<0.05). The levels of LH and E2 in the control group were (12.93±0.81) IU·L⁻¹ and (22.23±1.44) pmol·L⁻¹ respectively, and the levels in the Beta-CYP+EMB group were decreased to (5.63±0.41) IU·L⁻¹ and (10.45±0.78) pmol·L⁻¹ respectively (P<0.05).

Conclusion The combined exposure to beta-cypermethrin and emamectin benzoate may ultimately affect the estrous cycle of female rats by interfering with the secretion of reproductive hormones involved in the HPO axis.