| Objective:The pathological mechanism of chemotherapy-induced premature ovarian failure is still unclear and there is still no effective treatment for premature ovarian failure.In premature ovarian failure,the pathological manifestation of the ovary is characterised by fibrosis,and blockade of HMGB1 has been shown to relieve the symptoms of tissue fibrosis.Therefore,this study hopes to investigate the mechanism and role of high mobility group protein-1 in chemotherapy-induced premature ovarian failure in mice as a new target,and provide an advanced concept for therapy of POF.Method:Part I: The animal models of chemotherapy-induced premature ovarian failure in mice were established,and the model divided into modeling group(POF group),treated with chemotherapeutic drugs and control group(CON group),treated with drug solvent control.Mice were executed 14 days after injection of chemotherapeutic drugs,and their serum and ovary tissues were taken.HE stain,Masson stain,immunohistochemistry and Western Blot were used to detect the phenotype of mouse ovarian tissue and the changes of HMGB1 content.Part II: The animal models of chemotherapy-induced premature ovarian failure in mice with blocked HMGB1 were established using glycyrrhizin,and grouped as follows:glycyrrhizin(GL group)treatment group and normal saline(NS group)control group.The injection concentration was 25 mg/kg.As in the first part,mice were injected with chemotherapeutic drugs to establish the model,and the mice were injected with glycyrrhizin on the sixth,eighth,tenth and twelfth days after the injection of chemotherapeutic drugs for a total of four treatments of 200 μl/each.Mice were executed 14 days after injection of chemotherapeutic drugs,and their serum and ovary tissues were taken.As in the first part,the same technologies were used to detect the phenotype of mouse ovarian tissue and the changes of HMGB1 content and analyze its mechanism to protect POF.Result:(1)An animal model of chemotherapy-induced premature ovarian failure in mice was successfully established.The histomorphological results of HE staining showed that the follicular structure of the mouse ovaries was destroyed after the use of chemotherapeutic drugs,the number of primordial follicles was decreased,the number of atretic follicles was increased,the arrangement of granulosa cells at all levels of follicles was disturbed,nuclear consolidation was observed,and the shape of sinus follicles was irregular.Masson staining showed that the interstitial fibrosis was more severe.(2)The findings of immunohistochemistry indicated that HMGB1 in the CON group was mainly found in the nucleus,while in the POF group,HMGB1 was not only in the nucleus,but also outside the nucleus.Meanwhile the expression of HMGB1 was upregulated.It is suggested that HMGB1 positive cells increase in chemotherapy-induced premature ovarian failure and can be transferred from the nucleus to the cytoplasm or even outside the cell to play a active role.The results of WB indicated that the level of HMGB1 in the ovarian tissues after modeling was higher than that in the CON group(P<0.05).ELISA results can be seen that the expression of HMGB1 was increased in serum of mice in the modeled group.(3)After treatment with glycyrrhizin to block HMGB1,WB results indicated that the amount of HMGB1 protein expression in the ovarian tissue was reduced compared to the untreated group.The results of immunohistochemistry indicated that HMGB1 was mainly expressed in the nucleus of mouse ovaries.ELISA results suggested that the expression of HMGB1 in mouse serum was reduced.Immunofluorescence results showed that the expression of TGF-β and F4/80 in the ovaries was reduced,and the number of macrophages was decreased.It was suggested that blocking HMGB1 could protect against premature ovarian failure by reducing the release of TGF-β from macrophages.Conclusion:(1)HMGB1 is involved in the development of premature ovarian failure in chemotherapy-induced mice,and HMGB1 expression is upregulated in ovarian tissues of mice with premature ovarian failure.(2)GL as an HMGB1 blocker may protect against premature ovarian fibrosis in premature ovarian failure by reducing the release of TGF-β from macrophages.(3)HMGB1 molecule may be a new target for the treatment of premature ovarian failure and provide new ideas for treatment. |
