The purpose of this study was to investigate whether the local use of estradiol after anterior cruciate ligament (ACL) reconstruction with a polyethylene terephthalate (PET) artificial ligament graft could promote graft-bone healing. A total of 45 New Zealand white rabbits underwent ACL reconstruction with a PET ligament graft. The experimental groups were administered a local estradiol injection at either a low dose after surgery or a high dose after surgery, and the control group did not receive an injection. Computed tomography (CT) scans and blood sample collection were routinely performed in all three groups. Over time, the serum estradiol levels increased in both experimental groups, and the CT images revealed a trend of a shrinking bone tunnel area in all three groups. The rabbits were randomly sacrificed at 2, 4, and 8 weeks after surgery. The load to failure and stiffness of the experimental groups were significantly higher than those of the control group at 4 and 8 weeks. The histological study identified more bone mineralization in the experimental groups at 4 weeks after surgery compared to the control group. This study showed that the use of estradiol is a promising approach in promoting graft-bone healing in rabbits undergoing ACL reconstruction with a PET ligament graft.