The Immunomodulatory Effect Of ADSCs Combined With Estrogen On Regulatory T Cells In Patients With Premature Ovarian Insufficiency

RESEARCH OBJECTIVE:In recent years,studies have shown that POI is associated with autoimmune disorders,showing a decrease in peripheral blood regulatory T cells(Treg).The application of mesenchymal stem(MSCs)from different sources in POI animal models shows that MSCs can effectively improve the POI induced by chemotherapy.However,little is known about the use of MSCs for immune-induced POI.Estrogen(17P-E2)can promote the proliferation of Treg cells and enhance their immunosuppressive activity.Therefore,we investigated the immunomodulatory effect of human adipose-derived mesenchymal stem cells(ADSCs)combined with 17?-E2 on Treg cells in POI patients.METHODS:The first chapter:primary culture of ADSCs and flow identification of ADSCs and adipogenic induction,osteogenic induction.Isolation of PBMCs from healthy and POI patients by density gradient centrifugation.Under the stimulation of phytohemagglutinin(PHA),PBMCs was co cultured with 72h with different concentrations of ADSCs and 17?-E2 to detect the proliferation rate of lymphocytes,the mRNA levels of Foxp3,TGF-?1 and IFN-? and the proportion of Treg.The second chapter:In the first chapter,1×105 ADSCs and 10-9mol/L 17?-E2 with the best effect on promoting Treg cell proliferation were selected as the combination group.Under the stimulation of PHA,PBMCs were co-cultured with control group,10-9mol/L 17?-E2 group and 1×105 ADSCs group for 72h.Then we detected the mRNA levels of Foxp3,TGF-?1 and IFN-? and the proportion of Treg.RESULTS:The first chapter:The original cultured ADSCs had high purity,good activity and stable phenotype.ADSCs were positive for CD29,CD44 and CD 105 and negative for CD34,CD45 and human leukocyte antigen DR(HLA-DR).ADSCs have adipogenic and osteogenic differentiation ability.Compared with the control group,different concentrations of ADSCs and 17?-E2 could promote the proliferation of Treg cells,up-regulate the mRNA expressions of Foxp3 and TGF-?1 and down-regulate the mRNA expression of IFN-?.The second chapter:Compared with the control group,each experimental group could up-regulate Foxp3 and TGF-?1 mRNA levels and down-regulate the mRNA level of IFN-??and promote the expression of Treg cells?the combination group was the best.CONCLUSION:The first chapter:both ADSCs and 17?-E2 can play an immunomodulatory role and induce regulatory T cell production,in which 1 × 105 ADSCs and 10-9 mol/L 17?-E2 are the best concentrations.The second chapter:ADSCs and estrogen can promote the proliferation of Treg by promoting the expression of Foxp3,the secretion of TGF-?1 and the inhibition of the secretion of IFN-??ADSCs combined with 17?-E2 can play a synergistic role in inducing the immune regulation of Treg.