自身免疫性甲状腺炎自身抗原的实验研究进展

目前认为甲状腺球蛋白 (Tg) ,甲状腺过氧化物酶 (TPO)和促甲状腺激素受体 (TSHR)可能是参与自身免疫性甲状腺炎发生的主要自身抗原 ,就近年来对Tg、TPO和TSHR等甲状腺自身抗原表位及其抗原性等研究进展做一介绍。     

TPO mRNA和HGF/c-Met在甲状腺良恶性结节中的表达及意义

目的 探讨甲状腺过氧化物酶(TPO)mRNA和肝细胞生长因子及其受体蛋白(HGF/c-Met)在良恶性甲状腺结节中的表达及意义.方法 应用逆转录聚合酶链反应技术检测55例甲状腺结节新鲜标本的TPO mRNA丰度,采用免疫组织化学法检测600例甲状腺结节石蜡标本HGF/c-Met的表达.结果 甲状腺癌HGF/c-Met表达率显著高于甲状腺良性结节,乳头状癌高于滤泡型癌(P<0.01).甲状腺恶性结节TPOmRNA的阳性表达率显著低于甲状腺良性病变.结论 HGF/c-Met和TPO的联合检测在良恶性甲状腺病变的鉴别中有一定应用价值.     

乳头状甲状腺癌抑癌基因TSHR、P16和RAS启动子甲基化研究

目的 研究促甲状腺激素受体(TSHR)、P16和RAS 3个抑癌基因在乳头状甲状腺癌的表达,分析其启动子甲基化与肿瘤发生的关系.方法 对50例乳头状甲状腺癌组织和32例对照组织(20例结节性甲状腺肿,12例甲状腺腺瘤)提取RNA后,以RT-PCR检测3个抑癌基因mRNA的表达情况,采用甲基化PCR(MSP)检测上述组织中3个抑癌基因启动子区甲基化的情况,并对3个抑癌基因甲基化和未甲基化的组织随机进行测序.结果 50例乳头状甲状腺癌组织中,有34例(68.0%)TSHR基因、27例(54.0%)P16基因、30例(60.0%)RAS基因启动子发生了甲基化;32例对照组织中,有7例(21.9%)TSHR基因、5例(15.6%)P16基因、10例(31.3%)RAS基因启动子发生了甲基化;乳头状甲状腺癌组TSHR、P16、RAS基因启动子甲基化率均显著高于对照组(均P<0.05).TSHR、P16和RAS mRNA在乳头状甲状腺癌中的表达量明显低于对照组织(0.41±0.11对0.63±0.08,0.51±0.17对0.72±0.22,0.56±0.10对0.67±0.16,均P<0.05).经DNA测序证实,3个抑癌基因启动子发生甲基化的其CpG岛碱基未发生改变,仍为CG;未发生甲基化的碱基由CG变为TG.结论 3个抑癌基因启动子甲基化与乳头状甲状腺癌的发生和发展均相关.     

糖尿病大鼠甲状腺组织TG,TPO,TSHR与RAGE mRNA表达的实验研究

目的 探讨糖尿病大鼠甲状腺组织甲状腺球蛋白(TG)、甲状腺过氧化物酶(TPO)、促甲状腺激素受体(TSHR)与晚期糖基化终末产物受体(RAGF)mRNA表达的变化.方法 Wistar大鼠112只,随机分为糖尿病组(DM 组)27只、糖尿病氨基胍治疗组(AG组)28只、糖尿病胰岛素治疗组(INS组)32只和对照组(N组)25只,采用逆转录-聚合酶链反应检测造模后12和20周各组大鼠甲状腺组织TG、TPO、TSHR和RAGE mRNA的表达.结果 TG mRNA表达:12周各组之间无统计学差异(P>0.05).20周DM组低于AG组(P<0.01)、INS组(P<0.01)和N组(P<0.01).后3组间无统计学差异(P>0.05).TPO mRNA表达:12周DM组低于AG组(P<0.05)、INS组(P<0.001)和N组(P<0.001),AG组低于INS组(P<0.001)和N组(P<0.001),INS组低于N组(P<0.01).20周DM组低于INS组(P<0.01)和N组(P<0.001),与AG组无统计学差异,AG组低于INS组(P<0.05)和N组(P=0.001),INS组与N组无统计学差异.TSHR mRNA表达:12周DM组高于AG组(P<0.05)、INS组(P<0.05)和N组(P<0.01),后3组间无统计学差异(P>0.05).20周各组之间差别无统计学意义(P>0.05).RAGE mRNA表达:12周和20周均表现为DM组高于AG组(P<0.001)、INS组(P<0.01)和N组(P<0.001),INS组高于AG组(P<0.05)和N组(P<0.01),AG组与N组无统计学差异.结论 糖尿病大鼠甲状腺组织TG mRNA、TPO mRNA表达降低,TSHR mRNA表达增高,可能与高糖毒性所致甲状腺组织RAGE mRNA表达增高有关.     

酪氨酸对体外培养大鼠甲状腺细胞功能相关基因表达的影响

目的 研究酪氨酸对体外培养大鼠甲状腺细胞甲状腺球蛋白(TG)、甲状腺过氧化物酶(TPO)、钠碘转运体(NIS)及促甲状腺素受体(TSHR)mRNA表达的影响.方法 将体外培养大鼠甲状腺细胞分为对照组、酪氨酸Ⅰ组、酪氨酸Ⅱ组,每组5例,分别用含0、5％、10％酪氨酸的培养液培养96 h,收集甲状腺细胞,应用实时荧光定量PCR检测TG、TPO、NIS、TSHR mRNA表达.结果 对照组、酪氨酸Ⅰ组、酪氨酸Ⅱ组TG、NIS mRNA表达分别为1.11±0.36、2.08±0.45、2.52±0.65和1.21±0.36、1.94±0.57、2.10±0.56,组间比较差异有统计学意义(F值分别为10.48、4.42,P均＜0.05),其中酪氨酸Ⅰ组、酪氨酸Ⅱ组TG、NIS mRNA表达明显高于对照组(P均＜ 0.05).上述3组TPO、TSHR mRNA表达分别为1.02±0.54、1.14±0.29、1.17±0.38和0.81±0.44、0.91±0.30、0.90±0.19,组间比较差异无统计学意义(F值分别为0.18、0.26,P均＞0.05).结论 酪氨酸可能影响甲状腺细胞功能相关基因表达,增加碘在甲状腺的储存,通过作用于碘泵,调节甲状腺细胞的碘摄取功能.     

甲状腺乳头状癌组织中TSHR基因启动子甲基化研究

目的探讨甲状腺乳头状癌组织中促甲状腺激素受体基因（Thyroid Stimulating Hormone Receptor,TSHR）基因启动子区5端CpG岛甲基化改变的特点与临床特征的关系。方法采用甲基化特异性PCR（MSP,methylation-specific PCR）方法检测TSHR基因启动子甲基化情况。结果（1）22／34例甲状腺乳头状癌组织中检测到TSHR基因启动子甲基化,9／34例甲状腺乳头状癌癌旁组织中检测到TSHR基因启动子甲基化,癌组织中TSHR基因启动子甲基化率显著增高（χ^2=10．019,P=0．002〈0．05）;（2）有淋巴结转移的甲状腺乳头状癌组织（15／18例）TSHR基因启动子的甲基化显著高于无淋巴结转移组（7／16例）（χ^2=5．812,P=0．016〈0．05）。结论TSHR基因启动子异常甲基化是甲状腺乳头状癌发展过程中的分子事件之一,可能影响了甲状腺乳头状癌细胞的摄碘的功能。     

本刊2009年第4期部分文题介绍

1.维生素D与自身免疫性甲状腺疾病2.表达TSHR的腺病毒诱导的Graves病小鼠模型的研究进展3.自身免疫性甲状腺疾病与乳腺癌的关系4.选择性垂体甲状腺激素抵抗的新认识5.药物治疗Graves病停药后复发的相关因素研究进展6.糖尿病大鼠甲状腺组织TG、TPO、TSHR与RAGE mRNA表达的试验研究7.1型糖尿病免疫诊断标志物的研究进展     

半乳糖凝集素3表达在良恶性甲状腺肿瘤鉴别中的意义

目的　探讨半乳糖凝集素3表达对鉴别良恶性甲状腺肿瘤的临床价值。方法　采用免疫化学染色法(SP法)观察72例手术切除的甲状腺肿瘤标本半乳糖凝集素3的表达。结果　半乳糖凝集素3在恶性肿瘤的细针抽吸(FNA)细胞涂片与组织切片中均高水平表达,在良性肿瘤中不表达或低表达。良性与恶性甲状腺肿瘤在FNA细胞涂片或组织切片中半乳糖凝集素3的表达差异有统计学意义(χ2 =43. 73,χ2 =48. 16,均P<0. 01)。相同性质的甲状腺肿瘤FNA细胞涂片与组织切片的半乳糖凝集素3表达一致,两者之间差异无统计学意义(χ2 =0. 04,χ2 =0. 19,均P>0. 05)。结论　半乳糖凝集素3在良恶性甲状腺肿瘤中表达不同,恶性肿瘤中表达显著增高,是鉴别良恶性甲状腺肿瘤可靠的分子标记物,可用于FNA涂片标本及组织标本检测。     

碘与慢性淋巴细胞性甲状腺炎

慢性淋巴细胞性甲状腺炎(Chronic LymphoidThyroiditis),首先由Hashimoto于1912年报道,亦称桥本氏甲状腺炎(Hashimoto Thyroiditis)。近几十年的研究发现,其发病机理与自身免疫有关,故又称自身免 疫甲状腺炎。慢性淋巴细胞性甲状腺炎是一种典型的器官特异性自身免疫疾病,其自身抗体针对自体的甲状腺组织成分。如甲状腺球蛋白(Tg)、甲状腺过氧化物酶(TPO)等自身抗原。慢性淋巴细胞甲状     

甲状腺自身抗体的基础和临床进展

迄今已知的人类甲状腺自身抗原有促甲状腺激素受体(TSHR)、甲状腺球蛋白(TG)、甲状腺过氧化物酶(TPO)和钠-碘转运体(NIS)4种,与其相对应的抗体分别为TSHR抗体(TRAbs)、TGAb、     

Detection of thyrotropin-receptor messenger ribonucleic acid (mRNA) and thyroglobulin mRNA transcripts in peripheral blood of patients with thyroid disease: sensitive and specific markers for thyroid cancer

Because thyroid cancer cells express functional TSH receptors (TSHR), TSHR-mRNA in peripheral blood might serve as a tissue-/cancer-specific marker. We measured circulating TSHR-mRNA by RT-PCR in 51 normal controls, 27 patients with benign thyroid disease, 67 patients with treated differentiated thyroid cancer (DTC), and eight patients with newly diagnosed DTC, preoperatively. Results were compared with thyroglobulin (Tg) mRNA and serum Tg levels. TSHR-mRNA signals were not detected in normal controls and in 24 of 27 (89%) patients with benign thyroid disease. All 19 patients with treated DTC with evidence of distant or local disease tested positive for TSHR-mRNA (sensitivity 100%). Among patients with no evidence of disease, TSHR-mRNA was detected in 1 in 48 (specificity 98%). Six of the eight newly diagnosed DTC patients tested preoperatively were positive for TSHR-mRNA. The concordance between TSHR-mRNA and Tg-mRNA and between TSHR-mRNA and serum Tg was 95%. Fourteen patients with DTC (21%) had Tg antibodies, three with local disease (all positive for TSHR-mRNA), and 11 with no evidence of disease (all negative for TSHR-mRNA).Our results indicate that TSHR-mRNA and/or Tg-mRNA in peripheral blood are both equally sensitive and specific markers for monitoring thyroid cancer patients. Their principal value resides in the Tg antibody-positive patients in whom a positive or a negative mRNA value might have indicated or obviated the need for a whole-body scan. Furthermore, the high specificity combined with their ability to predict thyroid cancer preoperatively suggests a potential role in detecting thyroid cancer in patients with thyroid nodules.     

Semi-quantitative comparison of the differentiation markers and sodium iodide symporter messenger ribonucleic acids in papillary thyroid carcinomas using RT-PCR

OBJECTIVE: To investigate the levels of expression of the sodium iodide symporter (NIS) and three differentiation markers (thyroglobulin (Tg), thyroid peroxidase (TPO) and thyrotrophin receptor (TSH-R)) in 35 patients with primary (n=31) or recurrent (n=4) papillary thyroid carcinoma, and to compare the findings with clinical data. METHODS: We performed a multiplex semi-quantitative RT-PCR to analyse the relative levels of expression of Tg, TPO and TSH-R mRNAs, and a separate semi-quantitative RT-PCR for NIS mRNA. RESULTS: Tg, TPO and TSH-R mRNAs were expressed in all the patients, whereas NIS mRNA was expressed in all but eight. Analysis of the expression of the differentiation markers in all patients showed a significant correlation among Tg, TPO and NIS. With regard to the relationship between the expression of each gene and the MACIS score, there was significant correlation only for the Tg gene (P<0.05). CONCLUSIONS: The levels of expression of NIS mRNA correlated significantly with those of Tg and TPO mRNAs, but not with those of TSH-R mRNA. The relationship with clinical stage and prognostic score, however, varied among these differentiation markers.     

Thyroglobulin and thyroid peroxidase share common epitopes recognized by autoantibodies in patients with chronic autoimmune thyroiditis

Monoclonal antibodies specific for human thyroid peroxidase (TPO) were prepared by the hybridoma technique using hyperimmune spleen cells from mice immunized with TPO purified from thyroid glands from patients with Graves' disease. Use of the microenzyme-linked immunosorbent assay method revealed that some of the monoclonal antibodies cross-reacted strongly with human thyroglobulin (Tg). Conversely, monoclonal anti-Tg antibodies cross-reacted with TPO, albeit to a lesser degree. Some anti-Tg autoantibodies in serum from patients with chronic autoimmune thyroiditis purified by Tg affinity chromatography bound TPO, and such binding was completely inhibited by Tg. Western blotting experiments revealed that thyroid microsomal 103K proteins recognized by mouse monoclonal and polyclonal anti-TPO antibodies were recognized by some monoclonal anti-Tg antibodies and anti-Tg autoantibodies, and conversely, that 19S Tg was recognized by some monoclonal anti-TPO antibodies. TPO was immunoprecipitated by anti-Tg autoantibodies isolated by Tg affinity chromatography. On the other hand, the specificity for TPO of the anti-Tg autoantibodies was not identical with that of anti-TPO autoantibodies. These cross-reactivities were not due to contamination of TPO with Tg or vice versa, or to contamination of the anti-Tg autoantibody preparations with anti-TPO autoantibodies. Taken together, these data indicate that Tg and TPO share common antigenic determinants and that some of those determinants are recognized by autoantibodies in the serum of patients with chronic autoimmune thyroiditis.     

Low specificity of blood thyroglobulin messenger ribonucleic acid assay prevents its use in the follow-up of differentiated thyroid cancer patients

Thyroglobulin (Tg) is a glycoprotein specifically synthesized by follicular thyroid epithelium. After thyroidectomy and remnant (131)I ablation, serum Tg is a specific and sensitive marker for the presence of thyroid cancer tissue, and its measurement is fundamental in the follow-up of patients affected by differentiated thyroid carcinomas (DTCs), being even more sensitive than diagnostic whole-body scan. Unfortunately, serum Tg measurement becomes useless in approximately 15-25% of DTC cases who are positive for anti-Tg antibodies that interfere with the Tg measurement. In these cases, Tg mRNA measurement has been proposed as an alternative to serum Tg determination. The aim of this study was to verify the sensitivity and specificity of Tg mRNA measurement, performed by quantitative real-time RT-PCR, in a series of 100 subjects (80 DTC patients and 20 controls). From our data, the sensitivity and the specificity of the blood Tg mRNA measurement are 82.3 and 24.2%, respectively, with a positive predictive value and a negative predictive value of 65.6 and 43.7%, respectively. The comparison of the Tg mRNA with the serum Tg, measured by both chemiluminescent and ultrasensitive ELISA methods, confirmed the low specificity of the Tg mRNA assay. The hypothesis that Tg mRNA detectable levels could be predictive of future recurrences is not supported by the long follow-up (median, 7 yr; range, 3-29 yr) of our disease-free patients, who did not develop any recurrences in their clinical history. Moreover, nine disease-free patients, who showed positive levels of Tg mRNA (11.8-336 pg equivalents/ micro g RNA), were confirmed to be serum Tg free, both in basal conditions and after recombinant human TSH stimulation, 4 yr after the Tg mRNA detection. In conclusion, we demonstrated that the Tg mRNA assay is of poor utility in the follow-up of DTC patients. On the contrary, serum Tg measurement is a very sensitive and specific thyroid tumor marker, and we recommend that the follow-up of patients affected by DTC must be performed using serum Tg rather than blood Tg mRNA measurement.