Effect Of Blimp-1 On Abortion Induced By Autoimmune Thyroiditis Through PD-1/PD-L1 Pathway

Background: Previous studies have investigated that autoimmunity thyroiditis with normal thyroid function is associated with recurrent miscarriage,however,the exact mechanism remains unclear.Thyroid peroxidase antibodya(TPOAb)and thyroglubiolin antibody(Tg Ab)are important marker antibodies.A large thyroid disease survey " show that the positive rate of TPOAb in women of childbearing age was 12.9%,the positive incidence of pathological gastosis was 8.6%.The most common and serious adverse pregnancy outcome resulting from maternal thyroid dysfunction is autoimmune thyroiditis,which is miscarriage,especially recurrent miscarriage.Regulatory T cell(Regulatory T cells,Treg),it plays a crucial role in the abortion process leading to autoimmune diseases.It has been demonstrated that an increase or block of the programmed cell death molecule 1(PD-1)/ programmed cell death ligands(PD-L1)costimulatory pathway is directly linked to abortion caused by autoimmune disease.The effect of PD-1/PD-L1 on Treg is mainly through inhibition of protein kinase B/mammalian target of rapamycin signaling(AKT /m TOR)signal conduction is realized.PD-1/PD-L1 negative costimulation signal can reduce the downstream m TOR expression and further regulate the differentiation,proliferation,metabolism and survival of T cells.It plays an important role in regulating cell growth,proliferation,energy metabolism and cell cycle.B lymphocyte induced maturation protein 1,(Blimp-1),it also plays an important role in abortion caused by autoimmune diseases.Blimp-1 is an inhibitory transcription factor that regulates the terminal differentiation of T cells and B cells by direct binding to PD-1 promoter and inhibition of nuclear factor of activated T-cells.(Nuclear factor of activated T-cells,cytplasmic 1,NFATc1)thereby inhibiting PD-1.Pregnancy actually reflects the mother's immune tolerance to embryonic antigens,which includes the immune tolerance of mother-fetus interface and peripheral immune tolerance.Once the balance of immune tolerance is broken,the moyher will show immune rejection to the fetus leading to miscarriage occure.However,the interaction and influence of Blimp-1,PD-1/PD-L1,m TOR and Treg in pregnancy loss induced by autoimmune thyroiditis are rarely reported in the literature.Objective:The purpose of this study was to investigate the effect of Blimp-1 on the PD-1/PD-L1 pathway and the downstream role of m TOR and Treg in AIT induced abortion.Methods:(1)Animals model building: CBA/J female mice were immunized with mouse thyroglobulin to induce the abortion model of pregnant mice with autoimmune thyroiditis were induced by mating with Balb/c male mice.(2)Keep the serum,take the complete thyroid,placenta,spleen.(3)Detection of Blimp-1 in placenta and spleen by immunohistochemistry and immunofluorescence as well as PD-1/PD-L1.Expression of m TOR and Foxp3 protein,a representative molecule of Treg.(4)Application of Western blot(Immunoblotting)the placenta and spleen Blimp-1 were detected.PD-1 /PD-L1;The expression level of m TOR and Foxp3.(5)Analysis of Blimp-1,the level of PD-1/PD-L1,m TOR and Treg in placenta and spleen.Results:(1)Compared with the control group,the lavel of Tg Ab in the serum of the experimental group was higher than that in the control group.(P<0.05);It showed a large number of lymphocytes infiltration,follicular cells were destroyed,the model was successfully constructed.(2)Immunohistochemical results showed that compared with con group,Blimp-1,the expression of m TOR in placenta was increased.PD-1 and the expression of PD-L1 was decreased.Immunofluorescence results showed that the expression of Foxp3 in TG group was significantly lower in the con group.(3)The results of Western blot Showed that the expression of Blimp-1 and the expression of m TOR in placenta was significantly higher than that in con group(P<0.05);Compared with con group,TG group PD-1.Both PD-L1 and Foxp3 decreased(P<0.05).(4)Spleen IHC-P(immunohistochemistry)similarly,the expression of Blimp-1 and m TOR in the Tg group increased while the expression of PD-1 and PDL-1 and Foxp3 decreased significantly.(5)The results of splenic Western blot were slightly different.Compared with con group,the Blimp-1 of TG group was significantly higher than that of con group(P<0.05);PD-1,PD-L1 and Foxp3 decreased significantly(P<0.05);There was no significant difference in m TOR between TG group and con group(P > 0.05).Conclusion:(1)200 micro g mouse Tg protein(m Tg)immunized with CBA/J female mice could successfully construct the Tg Ab positive autoimmune abortion model:Lymphocyte infiltration and thyroid follicular cell destruction were observed on the 13 th day of gestation and the embryo absorption rate of pragnent mice was significantly increased and the embryo development was abnormal after immunization.And mouse TG protein(m Tg)after immunization,the mice in the experimental group showed higher TG antibody titers.(2)The abnormal increase of Blimp-1 in Tg Ab positive pregnant rats may lead to the change of PD-1/PD-L1 inhibition pathway resulting in the imbalance of fetal tolerance and peripheral tolerance in Tg Ab positive pregnant rats by regulating Treg cells through m TOR signaling pathway,which may participate in the occurrence of abortion.