TSH改变亚临床甲减小鼠胸腺中T细胞发育及TSH在免疫系统中的作用

发布时间：2018-06-24 01:43

本文选题：T细胞发育 + 亚临床甲状腺功能减退　；参考：《山东大学》2017年硕士论文

【摘要】：研究背景:促甲状腺素(thyrotropin,TSH)被认为仅仅甲状腺调节。然而,新的证据表明,高血清TSH水平可能存在潜在的有害的影响。高血清TSH水平,是非酒精性脂肪肝病和心血管疾病的独立危险因素,而且往往伴随着慢性炎症、胰岛素抵抗、氧化应激。亚临床甲状腺功能减退症(subclinical hypothyroidism,SCH),是一种临床上较为常见的隐匿性疾病,其特点是甲状腺功能血清学检测表现现为促甲状腺激素水平增高,但游离甲状腺素水平尚正常,是甲状腺机能减退障碍的早期阶段。亚甲减症的发病率呈现逐渐升高的趋势,并对健康有潜在的害影响。SCH在成年人中的患病率约为4-100%。胸腺是重要的中枢免疫器官,是T淋巴细胞离开骨髓后发育的重要场所。胸腺为T淋巴细胞的发育,提供了一个特定的微环境,既胸腺微环境。对胸腺内T细胞发育、分化以及成熟的研究,是当今免疫学研究的重要热门领域。研究表明,T细胞表达促甲状腺激素受体(TSHreceptor,TSH-R),而SCH中胸腺T细胞发育的变化尚未完全阐明。研究目的:本实验旨在阐明亚甲减小鼠模型中,TSH对胸腺T细胞发育的影响,SCH小鼠模型中胸腺的细胞组成情况以及不同发育阶段的T细胞凋亡变化。研究方法:1.SCH小鼠模型的构建:应用甲巯基咪唑喂养小鼠,甲巯基咪唑剂量为0.08mg/kg/d,甲巯基咪唑喂养16周时,采用I125标记放射免疫的方法,检测小鼠的血清中游离甲状腺素(FT4)水平,采取Elisa的方法测验小鼠血清TSH水平。通过评估小鼠的血清甲状腺功能,来构建SCH小鼠模型,使其血清检测符合SCH诊断标准,既血清TSH水平升高,甲状腺激素水平无显著变化的,研究TSH对T细胞发育的影响。2.SCH小鼠胸腺中不同发育阶段T的免疫学组成分析:应用多色彩流式分析的方法检测小鼠胸腺中T细胞不同发育阶段各T细胞亚群的比例。3.胸腺中不同发育阶段T细胞凋亡比例的测定:应用凋亡试剂盒检测各T细胞亚群细胞凋亡的比例。4.小鼠胸腺内凋亡相关蛋白检测:采取Western Blot法测验胸腺p-ERK/ERK的蛋白水平变化。5.胸腺近期输出功能检测:提取小鼠脾脏DNA,实时定量PCR检测小鼠脾脏T细胞DNA中TREC水平。结果:1.SCH小鼠模型的构建:甲巯基咪唑喂养小鼠16周,SCH组小鼠血清FT4测值与对照组小鼠相比,无明显差异性的变化(P0.05);SCH模型组血清TSH水平明显高于对照组小鼠(P0.05),两组血清TSH水平差异具有统计学意义。2.SCH小鼠胸腺中不同发育阶段T的免疫学组成分析:SCH小鼠胸腺重量比对照组增加18%。胸腺中CD4 +和CD8 +单阳性(single positive,SP)胸腺细胞分别增加了38%和44%。3.胸腺中不同发育阶段T细胞凋亡比例的测定:通过检测凋亡,我们发现在SCH小鼠胸腺中,Annexin-V+细胞比例减少(p0.05),证明TSH减少SCH小鼠胸腺T细胞凋亡。4.小鼠胸腺内凋亡相关蛋白检测:Western blot结果显示,SCH小鼠胸腺中p-ERK 1/2增加,表明SCH小鼠胸腺组织中ERK通路被激活。5.胸腺近期输出功能检测:PCR检测小鼠脾脏组织T细胞受体切除环(TREC),结果显示,SCH组小鼠脾脏T细胞中TREC增加(p0.05)。结论及意义:这些结果表明,TSH促进SCH小鼠胸腺T细胞发育,增强胸腺近期输出。其机制可能是通过抑制胸腺细胞的凋亡,抗凋亡作用可能是通过p-ERK/ERK信号通路。
[Abstract]:Background: thyrotropin (TSH) is thought to be only thyroid regulation. However, new evidence suggests that high serum TSH levels may have potentially harmful effects. High serum TSH levels are independent risk factors for nonalcoholic fatty liver disease and cardiovascular disease, and are often accompanied by chronic inflammation, insulin resistance, and oxidation. Stress. Subclinical hypothyroidism (subclinical hypothyroidism, SCH) is a clinically common occult disease characterized by thyroid function serological detection showing elevated levels of thyroid stimulating hormone, but free thyroxine level Shang Zhengchang is an early stage of hypothyroidism. Subhypothyroidism is a subgrade of hypothyroidism. The incidence of the disease is increasing gradually, and the potential harm to health of the.SCH in adults is about 4-100%. thymus, which is an important central immune organ and an important place for the development of T lymphocytes to leave the bone marrow. The thymus provides a specific microenvironment for the development of T lymphocytes, and the thymus microenvironment. Research on the development, differentiation and maturation of T cells in the thymus is an important area of immunology research today. Studies have shown that T cells express TSHreceptor (TSH-R), and the changes in the development of T cells in the thymus gland have not been fully elucidated. The purpose of this study was to elucidate the thymus T thin in the subsubsubtract mice model. The effect of cell development, the cell composition of the thymus in the SCH mouse model and the changes of T cell apoptosis in different developmental stages. Research methods: Construction of 1.SCH mice model: using methimidazole to feed mice, a dose of methimidazole at a dose of 0.08mg/kg/d, and 16 weeks after a mercapto methimidazole feeding, using I125 labelled radioimmunoassay to detect small amounts. The serum level of free thyroxine (FT4) in the rat serum was tested by Elisa method to test the level of TSH in the serum of mice. By evaluating the thyroid function of the mice, the SCH mouse model was constructed to make its serum test conform to the SCH diagnostic standard. The serum TSH level was elevated and the thyroid hormone level was not significantly changed. The effect of TSH on the development of T cells was studied. The analysis of the immunological composition of T in the thymus of.2.SCH mice: detection of the proportion of T cell subgroups in the different developmental stages of T cells in the thymus gland by multi color flow analysis. The ratio of apoptosis in the different developmental stages of the.3. thymus of the.3. thymus was measured: the apoptosis ratio of each T cell subgroup was detected by the apoptotic test kit. 4. the detection of apoptosis related proteins in the thymus of mice: the Western Blot method was used to test the protein level of p-ERK/ERK in the thymus, and the short-term output function of.5. thymus was detected: the spleen DNA in mice was extracted and the TREC level in the T cell DNA of the spleen was detected by PCR in real time. The result: the construction of 1.SCH mouse model: methimidazole feeding mice for 16 weeks, SCH group mice. There was no significant difference in serum FT4 values compared with the control group (P0.05); the serum level of TSH in the SCH model group was significantly higher than that of the control group (P0.05). The difference in serum TSH level between the two groups was statistically significant in the immunological analysis of T in the different developmental stages of the thymus of.2.SCH mice: the thymus weight of SCH mice was more than the 18%. thymus in the control group. CD4 + and CD8 + single positive (single positive, SP) thymic cells increased the percentage of apoptotic T cells in 38% and 44%.3. thymus, respectively. By detecting apoptosis, we found that the proportion of Annexin-V+ cells in the thymus of SCH mice decreased (P0.05), which showed that TSH decreased the apoptosis phase in the thymus gland of SCH mice. The Western blot results showed that the p-ERK 1/2 in the thymus of SCH mice increased, indicating that the ERK pathway in the thymus of the SCH mice was activated to detect the short-term output function of the.5. thymus: PCR detection of the T cell receptor excision ring in the spleen tissues of mice (TREC). The results showed that TSH promoted the development of thymus T cells in SCH mice and enhanced the recent output of thymus. The mechanism may be through inhibiting the apoptosis of thymus cells, and the anti apoptosis effect may be through the p-ERK/ERK signaling pathway.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R581.2

【相似文献】