Evaluation Of The Efficacy Of A Gossypol-loaded Intrauterine Device On Adenomyosis In An ICR Mouse Model

Objective:To evaluate the efficacy of a gossypol acetate -loaded intrauterine device on adenomyosis,and explore the possible and the mechanism of local gossypol acetate in an ICR modelMethods:(1) Preparation of the gossypol-loaded intrauterine device(IUD),gossypol acetate and silicone gel were mixed at the ratio of 1:9(w:w).To test the release rate of gossypol acetate from the IUD in vitro,the gossypol acetate concentration was detected using the high-performance liquid chromatography(HPLC) method.(2) To test the release rate of gossypol acetate from the IUD in vivo,medication with the ICR mice with gossypol acetate-loaded IUD(1mg/20g)(n=24),taking the blood samples after the medication respectively.The blood was collected in tubes with heparin sodium,the plasma separated by centrifugation.The gossypol acetate concentration in the plasma was assayed using the HPLC method.(3) An adenomyosis animal model was established using Institute for Cancer Research,Swiss-derived(ICR) mice,grafted with a single pituitary gland.The mice were sacrificed by cervical dislocation 4 months after implantation and the uterine tissue investigated by naked eye observation,histology using hematoxylin-eosin(HE) staining and immunohistochemical method,the others of implantation group,operation but not be implanted group and control group were samly deal with 6 months after operation.(4) Four months after grafting,IUDs with three different quantities of gossypol acetate were prepared and used to treat the ICR mice with adenomyosis.After 2 months of treatment with a gossypol acetate-loaded IUD,the number of adenomyosis nodules, the hematoxylin-eosin staining scores and ER expression were measured and compared with mice given daily oral gossypol acetate and controls(no adenomyosis).Results:(1) The releasing characteristics of gossypol aceate-loaded IUD in vitro can be described by two-phase dynamic model.The accumulative releasing percentage of 24h was 25%,the release then followed zero-order kinetics,It released about 0.3%everyday after 15th day.The gossypol acetate was continually and steadily released over 50 days when the ratio of gossypol acetate/carrier was 1:9.(2) The in vivo release test showed that the concentration of plasma gossypol acetate gradually increased to a peak level of 0.30μg/l on the 2th day,which was then maintained for 2 months.(3) The histology confirmed that adenomyosis had developed in 17 of 19 ICR mice (89%) 4 months after grafting with a pituitary gland.,adenomyosis had developed in 8 of 8 ICR mice(100%) 6 months after implantation.(4) As the gossypol acetate dose increased,the nodule number decreased reaching significance at a dose of 0.2mg per 20 g body weight(P＜0.05) Maximal effect was achieved with an IUD loaded with 0.4mg gossypol acetate per 20g body weight,the nodules,the HE scores and ER expression were significantly decreased in mice treated with gossypol acetate—loaded IUD or intragastric gossypol acetate. Conclusion:The ICR mice are a suitable animal model for adenomyosis.A gossypol acetate-loaded IUD is a reliable method of administration for gossypol acetate and that a gossypol acetate-loaded IUD is an effective way to treat adenomyosis in ICR mice.