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Background: Rutin (quercetin-3-rhamnosyl-glucoside), a flavonoid, is derived from plants and has antioxidant properties. Objectives: This study aimed to evaluate the effect of different concentrations of rutin on mouse ovary heterotopic allotransplantation. Methods: The present animal experimental study was conducted on 40 female adult Balb/c mice weighing 30±5 g at the Jundishapur University of Medical Sciences, Ahvaz, Iran, during 2016 - 2018. The mice were divided by permuted block randomization into 8 groups (n = 5): OVX (ovariectomy), as the negative control; normal (positive control); OVX + OVA (ovariectomy and transplantation) (control), treated with 0.5 mL of normal saline; OVX + OVA + 10 mg/kg of rutin; OVX + OVA + 30 mg/kg of rutin; OVX + OVA + 60 mg/kg of rutin; OVX + OVA + 100 mg/kg of rutin; and the autograft. Groups were treated daily. Fourteen days after transplantation, ovarian grafts were collected and processed histologically for follicle number counting. Serum estrogen (E2) and progesterone (P4) levels were evaluated. Furthermore, the expression of Estrogen Receptor alpha (ERα), Estrogen Receptor beta (ERβ), and Progesterone Receptor (PR) in the uterine endometrial tissue was tested using qRT-PCR and western blotting. Results: A decrease in the number of mature follicles and increase in the number of atretic follicles (mean ± SD: OVX + OVA + 30 = 19.00± 1.000, OVX + OVA + 60 = 25.00± 5.000, and OVX + OVA + 100 = 23.00 ± 2.646) were observed in all groups treated with rutin in comparison with the control group (mean ± SD: 12.33 ± 2.517) (P value < 0.05). The level of E2 and P4 (mean ± SD: OVX + OVA + 100 = 6.133 ± 1.026) increased in comparison with the OVX + OVA group (mean ± SD: 0.4667 ± 0.2517) (P value < 0.05). The protein expression of ERα (mean ± SD: OVX + OVA + 10 = 1.615 ± 0.1701 and OVX + OVA + 30 = 1.744 ± 0.1779) in comparison with the control group (mean ± SD: 0.7089 ± 0.1131), and ERβ (mean ± SD: OVX + OVA + 10 = 0.7747 ± 0.4365, OVX + OVA + 30 = 0.9220 ± 0.1245, OVX + OVA + 60 = 0.7701 ± 0.2150, and OVX + OVA + 100 = 0.6676 ± 0.1547) increased in a dose-dependent manner in all groups treated with rutin in comparison with the OVX + OVA group (mean ± SD: 0.1534 ± 0.06109) (P value < 0.05). No significant changes in PR were found in groups treated with rutin in comparison with the control group. Conclusions: The results of the present study indicated that rutin increases E2 and P4 levels in ovarian hetero allograft mice. Rutin also upregulated the expression of ERα and ERβ but had no significant effect on PR.