The Study Of GARP Expression Level And Promoter Methylation Of Peripheral Blood Mononuclear Cells In Hashimoto’s Thyroiditis Patients

ObjectiveTo research the expression and significance of GARP m RNA and protein,Foxp3 m RNA and TGF-β m RNA in peripheral blood mononuclear cell（PBMC）of patients with Hashimoto’s thyroiditis（HT）.And research methylation changes in the promoter region of GARP gene to explore the effect on the expression of GARP and HT disease.Methods32 HT patients meeting the inclusion criteria in outpatients were selected,and 32 healthy people were picked out as the control group.Fasting venous blood was collected from all subjects,and serum TSH,TT3,TT4 and thyroid antibody Tm Ab and Tg Ab were determined by chemiluminescence immunoassay.In addition,5 ml peripheral venous blood was collected,and PBMC was isolated to detect GARP m RNA,Foxp3 m RNA,TGF-β m RNA,and analyze the correlation between GARP,Foxp3,TGF-β and indicators of thyroid inflammation.Total protein was extracted from PBMC of 15 HT patients and 15 healthy controls,and the expression of GARP protein was detected by Western Blotting（WB）.One part was used to extract DNA,and Methylation Specific PCR（MSP）was used to detect the methylation status of GARP promoter in the samples.Results1.The age of HT group and control group were（42.28±12.345）and（41.13±10.388）,respectively;and the gender composition（male/female）were（2/30）and（3/29）,respectively.There were no significant differences in age,sex,TT3,TT4 and TSH between the two groups（P>0.05）,the levels of Tg Ab and Tm Ab were significantly different（P<0.001）.2.The relative expression levels of GARP m RNA,Foxp3 m RNA and TGF-β m RNA in PBMC of HT patients were 0.27（0.16-0.62）,0.21（0.11-0.32）and 0.45（0.29-0.62）,respectively;Significantly lower than the control group 0.74（0.58-1.20）,0.66（0.48-1.16）,1.01（0.88-1.18）,the difference was statistically significant（P<0.001）,and the relative expression of GARP m RNA was negatively correlated with Tg Ab level（r=-0.421,P=0.016）,and was positively correlated with TT3（r=0.453,P=0.009）.The decrease of GARP protein expression in HT group was lower than that in control group,and the difference was statistically significant（P<0.001）.3.The methylation level of GARP promoter in HT group was higher than that in control group,and the difference was statistically significant（P=0.008）.Conclusions1.Decreased expression of GARP,Foxp3 and TGF-β in HT.Abnormal expression of GARP and TGF-β may be an important mechanism for the occurrence of HT.Ultimately,the number of Treg cells decreased and the immunosuppressive activity decreased,thus participating in the occurrence and development of HT.2.The downregulation of GARP of HT is related to the highly methylated promoter of HT,and inhibition of the methylation of GARP promoter may increase its expression,thus changing the occurrence of HT.This may be a new target for clinical intervention of HT.