Effect Of Aikangkangtai On The Balance Of Th1/Th2 Cell Balance And Apoptosis In AIT Model Rats

ObjectThis study was aimed to explore the mechanism of Jia-Yan-Kang-Tai(JYKT)on Th1/Th2 immune cell imbalance and Apoptosis in Rats with Autoimmune Thyroiditis.MethodsFifty female Lewis rats were randomly divided into the normal group,model group,JYKT high-dose,middle-dose and low-dose group by their weight,with 10 rats in each group.Autoimmune thyroiditis(AIT)rat model was established by the method of subcutaneous injection of Thyroglobulin from porcine thyroid gland and Freund's adjuvant in the other groups except the normal group for six weeks.Those groups were treated with deionized water,and JYKT formula granules gavage administration respectively for 8 weeks,which included normal group(deionized water lml/100(g.d)),model group(deionized water lml/100(g.d)),JYKT low-dose group(JYKT granules 0.708g/(kg.d)),JYKT middle-dose group(JYKT granules 1.417g/(kg·d)),and JYKT high-dose group(JYKT granules 2.834g/(kg.d)).After gavage administration,blood was taken from abdominal aorta in anesthetized rats.Serum thyroglobulin antibody(TgAb)and thyroid peroxidase antibody(TPOAb)were measured by Elisa.The ratio of Thl cell,Th2 cell and Thl/Th2 was measured by flow cytometry.IL-4,IL-5,IFN-?,TNF-? and IL-lain plasma were measured by AimPlex Multiple Immunoassays for Flow.Thyroid tissue of all rats were sampled and given HE dyeing.Apoptosis measured by TUNEL,expression distribution of IFN-?,IL-4,Fas,FasL,Bcl-2 and Bax in Thyroid tissue were observed under immunohistochemical staining.RT-PCR was used to measure mRNA expression of Fas?FasL?Bax?Bcl-2?FLIp?Caspase-3 in spleen.The experimental data were analyzed with IBM SPSS Statistics 20 statistical software.Results(1)Serum detection:the significant improvement of TPOAb and TGAb of rats in model group(P<0.01),comparing with normal rats;comparing with model rats,AIT rats in JYKT high-dose group showed that TPOAb(P<0.05)and TGAb(P<0.01)decreased significantly.(2)Morphological detection:HE dyeing showed that lymphocytic infiltrates in rat thyroid tissue decreased significantly and follicular epithelial cells were more complete in the three JYKT groups,compared with the model group,A slight decreased in the content of the Thyrocolloid in the three JYKT groups,but it was better than the model group's.In addition,thyroid follicular structure was complete in the three JYKT groups.The high dose group was the best of the three groups.(3)The detection of AimPlex Multiple Immunoassays for Flow:comparing with the normal group,IFN-y.TNF-??IL-1??IL-5(P<0.01)and IL-4(P<0.05)increased in the model group;comparing with the model group,IFN-y(P<0.05)decreased in the high-dose group;comparing with the model group,TNF-?(P<0.05)decreased in middle-dose and high-dose group;comparing with the model group,IL-ladecreased in the low-dose group(P<0.05)and high-dose group(P<0.01);comparing with the model group,IL-4(P<0.05)decreased in low-dose,middle-dose and high-dose group;comparing with the model group,IL-5 decreased in the low-dose group(P<0.05)and high-dose group(P<0.01).(4)Flow cytometry detection:comparing with the model group,the ratio of Thl cell(P<0.01),Th2 cell(P<0.05)and Thl/Th2(P<0.05)decreased in JYKT high-dose group.(5)Immunohistochemical staining detection:?serum cytokines:compared with the model group,the protein level in Thyroid Tissue of IFN-??IL-4 was decreased significantly in the three JYKT groups(P<0.01);?Apoptosis factor:compared with the model group,the protein level in Thyroid Tissue of Fas?Fas-L?Bax was decreased and Bcl-2 was increased significantly in the three JYKT groups(P<0.05,P<0.01).(6)Roche TUNEL detection:The number of apoptotic cells in the model group is more than that in the normal group,and is mainly located in the infiltration area of the inflammatory cells;compared with the model group,the number of apoptotic cells decreased in low-dose and middle-dose group.In addition,the high dose group was more obvious.(7)Real time-PCR detection:comparing with model rats,AIT rats in JYKT high-dose group showed that the mRNA level in spleen of Fas?FasL?Bax?FLIp?Caspase-3 was decreased and Bcl-2 was increased significantly in the high-dose group(P<0.01).Conclusions(1)Jia-Yan-Kang-Tai was able to suppress thyroid autoantibodies and lymphocyte infiltration of AIT rats.(2)Jia-Yan-Kang-Tai could decrease the levels of cytokines and adjusting Thl/Th2 immune cell imbalance.(3)Jia-Yan-Kang-Tai can relieve AIT by adjusting apoptosis protein and gene to reduce apoptosis in Thyroid Tissue.