The Role Of Treg Cells In Auto-immune Ovarian Failure Induced By Thymectomy

Background and objectivePremature ovarian failure (POF) refers to a syndrome consisting of amenorrhea,infertility and elevated levels of gonadotropins and reduced levels of oestrogen in a womanbefore the age of40. It affects approximately1%of women under the age of40years,0.1%of women under30years and0.01%of women younger than20years. The causes of thiscondition include: genetic aberrations, autoimmune ovarian damage, infection,chemotherapy and environmental factors. In some cases the underlying cause is unknown,in which case autoimmunity may play an important role. In mice, thymectomy in day3afterbirth results in changes in the ovary characterized by follicle atresia, oophoritis andlymphocyte infiltration of the follicles, leading to premature ovarian failure.Thymus-produced CD4+CD25+T-regulatory (Treg) cells suppress the activation andexpansion of self-reactive T cells that can potentially cause autoimmune disease, and play akey role in the maintenance of immunologic self-tolerance, thereby preventing autoimmunedisease including POF. It is hoped that a better understanding of the molecular mechanismof POF will lead to effective treatment for the condition.Methods1. Female BALB/c mice were isolated when visibly pregnant. Thymectomy wasperformed by the suction techniques on3-day-old anesthetized mice (D3tx). Alternatelittermates were used as sham-thymectomized controls. Serum was collected at the time ofsacrifice for measurement of follicle-stimulating hormone (FSH), estradiol （E2）,luteinizing hormone （LH） by ELISA. The ovaries were examined by the use ofimmunohistochemistry.2. The changing levels of the CD4+CD25+Foxp3+cells and T lymphocyte subsets inthe para-aortic lymph node, inguinal lymph node, spleen and peripheral blood of the D3txmice and the control mice were examined by flow cytometry.3. The CD4+CD25+Treg cells were separated using magnetically activated cell sorting,and injected intraperitoneally into d3tx mice following3days. The circulating sex hormone levels of serum sex hormone and ovarian histology were then examined. The CD4+CD25+Foxp3+cells and T lymphocyte subsets in different lymph nodes were compared.Results1. The circulating serum hormone concentration in the D3tx mice and control mice,the FSH concentration in the D3tx group was higher than the control mice,and the E2,LHconcentration was significantly lower than the control mice.2. In the D3tx mice, there were chronic inflammatory changes in the ovary, significantreduction of follicle count and infiltration of the stroma by lymphocytes.3. In the D3tx mice, the percentage of CD4+CD25+Foxp3+among CD4+T cells in theperipheral blood of rapidly increased from1stto2ndweek, and then decreased until9thweek.4. In the D3tx mice,the T lymphocyte cells was decreased and the Treg cells/CD4cellratios expanded simultaneously compared the control mice.5.Transfer of Treg cells derived from adult mice into D3tx mice successfully preventedthe development of premature ovarian failure in thymectomized mice.6. Following treatment of D3tx with Treg cells derived from adult mice, there was asignificant increase in Treg cells/CD4cell ratios in spleen and peripheral blood, but therewas no difference in the ratio in the para-aortic lymph node and inguinal lymph nodebetween the D3tx mice with and without adoptively transfered with Treg cells.Conclusion:Thymectomy in day3mice results in ovarian changes characterized by follicle atresia,oophoritis and lymphocyte infiltration of the follicles, leading to premature ovarian failure.It is suggested that autoimmune ovarian disease is a chronic inflammatory disordermediated by T-cells. The use of Treg cell was adoptively transferred into D3tx mice thatsuccessfully prevented the development of AOD in thymectomized mice, especially Tregfrom ovarian lymph nodes (LN) preferentially suppress AOD. In D3tx mice, increased Tregcan modify AOD at the early phase, but they cannot fully prevent autoimmune diseasedevelopment.