| Background:The pituitary hormone thyrotropin(thyroid stimulating hormone,TSH)has been thought to regulate solely the thyroid gland.However,new evidences have emerged showing that high serum TSH level,may have potential deleterious effects.Subclinical hypothyroidism(SCH),is a common clinically occult disease,which is characterized by elevatedthyroid stimulating hormone levels,with normal free thyroxine level in serological detection of thyroid function.SCH is the early stage of hypothyroid disorder.The incidence of subclinical hypothyroidism is gradually increasing,and it has potential harm to health.The prevalence of subclinical hypothyroidism in adults is about 4-10%.Thymus is an important central immune organ,which is an important site for the development of T lymphocytes.The thymus provides a specific microenvironment for the development of T lymphocytes,as well as the thymic microenvironment.The study on the development,differentiation and maturation of T cells in thymus is an important field of immunology.High serum TSH level is an independent risk factor for nonalcoholic fatty liver disease and cardiovascular diseases,and often accompanied by low grade chronic inflammation,insulin resistance and oxidative stress.Although developing T cells express thyroid stimulating hormone receptor(TSH-R),the changes of T cell development in thymus in SCH have not been fully clarified.Objective:This experiment is intended to clarify the effect of TSH on thymus T cell development in SCH mice and to compare the cell composition of thymus and the apoptosis of T cells of different developmental stages.Methods:1.Construction of subclinical hypothyroidism mice model:Usingmethimazole(dose of methimazole,0.08 mg/kg/d)feeding mice for 16 weeks.The serum thyroid function was assessed in mice to construct subclinical hypothyroidism serum mice meet diagnostic criteria for subclinical hypothyroidism.Serum free thyroid hormone in mice(FT4)level was detected using the 1125 labeling method of radioimmunoassay.TSH level in serum was detected by Elisa.SCH mouse model,which is characterized by elevated serum TSH but similar thyroid hormone levels,were used to studythe role of TSH in T cell development.2.T cell developmental stages and thymus composition analysis in subclinical hypothyroidism mice:The frequency of developmental stages of thymocytes subpopulations was distinguished usingflow cytometry.Surface molecules expression was detected to distinguish the developmental stages of thymocytes.3.The percentage of apoptosis of T cells ofdifferent developmental stages in thymus:The percentage of apoptosis in T cell subsets was detected by apoptosis kit.4.Detection of apoptosis related proteins in thymus:The changes of extracellular-regulated kinases(ERK)1/2 protein level was determined by Western Blot analysis.5.Detection of recent thymic output function:Spleen DNA extraction and RT-PCR analysis to detectTREC.Results:1.Based on the characteristic elevated serum TSH levels only in SCH,we established the SCH mouse model.The SCH mice exhibited equal serum fT4 levels and higher TSH levels compared with their littermate controls.2.Thymusweight of SCH mice increased 18%compared with controls.Importantly,the frequencies of CD4+ and CD8'single positive(SP)thymocytes increased 380%and 440,%respectively.3.We demonstrated that TSH protected thymocytes from apoptosis as evidenced by a significant decrease of Annexin-V positive thymocytes in SCH mice(p<0.05).4.Western Blotanalysis showed thatp-ERK 1/2 in thymus was activated in SCH mice.5.With analysis of T-cell receptor excision circles(TREC),we found that TSH increased recent thymic emigrants(RTEs)in spleen tissue in SCH mice.Summary and conclusions:Thus,these results suggest that TSH promoted T cell development and enhanced the thymic recent output in SCH mice,possibly by suppression of apoptosis of thymocytes,indicating that modification of the ERK signaling pathways. |
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