B7-1、CD40和CD54分子在Graves病患者甲状腺组织中的表达

目的探讨T细胞共刺激分子B7-1、CD40和CD54在Graves病(GD)患者甲状腺组织中的表达及其发病机制。方法采用免疫组织化学方法(HRP)观察51例GD患者和50例非毒性结节性甲状腺肿(NTG) 患者甲状腺组织T细胞共刺激分子B7-1、CD40和CD54的表达,应用Mias 99图像分析系统进行定量分析。结果 B7-1、CD40和CD54分子在所有GD患者的甲状腺组织中呈阳性表达,强阳性染色区域主要分布在甲状腺滤泡的表面和细胞质,在浸润的淋巴细胞区部分呈强阳性表达;而在NTG患者的甲状腺组织几乎不着色。B7-1、 CD40和CD54分子阳性表达例数、阳性百分率、阳性颗粒面积及平均吸光度值,在GD组明显高于NTG组(P< 0．01)。结论 T细胞共刺激分子B7-1、CD40和CD54的异常表达在GD的发病中起一定的作用。     

自身免疫性甲状腺疾病甲状腺组织中凋亡调控蛋白Fas/FasL、Bcl-2/Bax的表达及意义

目的探讨凋亡调控蛋白Fas/FasL、Bcl-2/Bax在Graves病(GD)和桥本氏病(HT)中的表达及意义。方法取外科手术切取的GD及HT病人甲状腺组织标本,颈部手术时取正常甲状腺组织作为对照;采用免疫组织化学方法检测甲状腺标本Fas/FasL、Bcl-2/Bax的表达和分布状况。结果(1)Fas/FasL在GD和HT甲状腺滤泡上皮细胞表达均较正常对照组增高(P<0.01)。(2)在GD甲状腺滤泡上皮细胞Bcl-2及Bax表达明显高于HT组及正常对照组,以Bcl-2表达强度显著高于同组Bax(P<0.01),而在HT,Bcl-2表达强度与正常对照组比较差异无统计学意义。(3)GD与HT相比较,GD甲状腺滤泡细胞及浸润淋巴细胞Fas,Bcl-2抗原表达均强于HT,但其浸润淋巴细胞表达强度明显低于同组甲状腺滤泡细胞(均P<0.05)。结论Fas/FasL高表达和Bcl-2的低表达可能引起HT甲状腺滤泡上皮细胞的凋亡。在GD,Fas、Bax诱导细胞凋亡的作用可能被高表达Bcl-2对抗,从而致甲状腺体积增大、功能亢进。此外,Bcl-2与Bax表达强度的比值对于凋亡的调控有重要影响作用。     

CD28-T细胞亚群在类风湿关节炎患者外周血和关节液中的表达

目的 探讨CD28-T细胞亚群在类风湿关节炎(RA)患者外周血和关节液中的变化和意义。方法 随机选择RA患者45例,取新鲜抗凝外周血单个核细胞(PBMC),其中15例同时提取关节液单个核细胞( SFMC),以流式细胞技术检测CD28-T细胞数量及其表面可诱导共刺激分子(ICOS)的表达。2 组间比较用独立样本t检验。结果 ①与PBMC相比,RA患者SFMC中CD4+CD28+ ICOS+、CD4+CD28-ICOS+、CD8+ CD28+、CD8+ CD28+ ICOS+T细胞明显升高[(36±19)％与(15±8)％,t=-4.234,P＜0.01;(2.1±2.2)％与(0.6±1.4)％,t=-3.143,P＜0.01;(62±15)％与(47±18)％,t=-2.885,P＜0.01;(9±9)％与(3±3)％,t=-2.131,P＜0.05];CD8+CD28-T细胞明显降低[(38±15)％与(54±18)％,t=2.975,P＜0.01];CD8+ CD28- ICOS+、C1D4+CD28+和CD4+CD28-T细胞无明显变化（P＞0.05）。②同一RA患者SFMC与PBMC相比,CD4+CD28+ICOS+、CD8+ CD28+T细胞明显升高[(38±18)％与(16±10)％,t=-4.065,P＜0.01;(61±16)％与(41±21)％,t=-2.883,P＜0.01];CD8+ CD28-T细胞明显降低[(39±16)％与(59±21)％,t=2.949,P＜0.01]。③缓解期与活动期RA患者相比,PBMC中CD4+CD28-、CD8+ CD28-、CD28-ICOS+T细胞无明显变化（P＞0.05）。结论 RA患者关节液中CD28-T细胞亚群失衡和ICOS分子表达异常,可能是导致RA关节损伤的重要机制。     

Graves病患者甲状腺组织内IL-6及gp130表达增强

对8例Graves病(GD)甲状腺组织内白细胞介素6(IL-6)及其受体(gp80、gp130)表达进行研究,发现GD组甲状腺组织内IL-6mRNA、gp130mRNA表达增高;GD中甲状腺滤泡细胞和淋巴细胞胞浆IL-6表达水平显著升高.提示GD患者甲状腺内IL-6/gp130信号通路的激活可能参与GD的发病.     

Graves病患者甲状腺细胞凋亡的初步研究

目的　研究Graves病 (GD)甲状腺细胞凋亡和凋亡相关蛋白Fas、可溶性Fas(sFas)及Fas配基 (Fas L)的表达特征 ,探讨Fas介导的细胞凋亡与GD发病机制之间的内在联系。方法甲状腺组织取自 7例GD手术患者和 3例意外死亡健康个体 ,采用原代甲状腺细胞 (TEC)培养技术、流式细胞术、ELISA及半定量RT PCR法检测TEC凋亡率及Fas、sFas和Fas L的表达情况。结果(1)GDTEC凋亡率为 (3.42± 0 .81) %,明显高于正常对照 (P <0 .0 1)。 (2 )GDTECFas表达率为 (8.3± 1.7) %,与正常对照相比差异无显著性。 (3)GDTEC培养上清液中sFas含量明显高于正常对照(P <0 .0 1)。 (4 )正常及GDTEC均表达Fas及sFasmRNA ,而Fas LmRNA仅在GDTEC表达。与正常对照相比 ,GDTECFasmRNA差异无显著性 ,sFasmRNA含量则明显升高 (P <0 .0 1)。结论GD患者甲状腺细胞存在细胞凋亡和Fas系统的异常表达 ,提示Fas介导的细胞凋亡参与GD的发病过程 ,sFas产生增加可能与甲状腺细胞增殖有关。     

自身免疫性甲状腺疾病甲状腺组织中bcl-2家族蛋白的表达

目的研究凋亡相关基因bcl鄄2家族蛋白bcl鄄2、mcl鄄1、bcl鄄XL和bax在自身免疫性甲状腺疾病(AITD)甲状腺组织中的表达特征及与AITD发病机制之间的内在联系。方法以甲状腺腺瘤旁正常甲状腺组织为对照(C组,20例),采用免疫组织化学ElivisionTM二步染色法检测凋亡相关蛋白bcl鄄2、mcl鄄1、bcl鄄XL和bax在桥本甲状腺炎(HT组,33例)和Graves病(GD组,28例)患者甲状腺组织中的表达与分布。结果bcl鄄2蛋白表达强度GD组>C组>HT组(P<0.01);mcl鄄1蛋白表达强度GD组>C组>HT组(P<0.01);bcl鄄XL蛋白表达强度HT组和GD组强于C组(P<0.01),但HT组和GD组间差异无统计学意义(P>0.05);bax蛋白的表达强度HT组>GD组和C组(P<0.01),但GD组和C组间差异无统计学意义(P>0.05);HT组中,在淋巴细胞浸润区域附近的甲状腺滤泡上皮细胞(TEC)bcl鄄2表达弱,bax和mcl鄄1表达强;远离淋巴细胞浸润区域的TECbcl鄄2表达强,bax和mcl鄄1表达弱(P<0.05)。结论(1)抗凋亡bcl鄄2和mcl鄄1蛋白在HT中表达的减弱以及在GD中表达的增强对于HT甲状腺滤泡细胞凋亡的增加和GD甲状腺滤泡细胞的增殖可能起一定作用;(2)bax蛋白在HT中表达增强所起的促凋亡的作用对疾病的发生发展起一定作用;(3)bcl鄄2与bax表达强度的比值对于凋亡的调控起重要作用;(4)bcl鄄2家族蛋白bc     

Graves病白细胞减少易感性与HLA-DRB1基因多态性的关联

目的探讨天津地区汉族Graves病(GD)白细胞减少易感性与HLA-DRB1基因多态性的关联.方法采用聚合酶链反应-序列特异性引物(PCR-SSP)方法检测45例GD白细胞减少患者、50例GD白细胞正常患者和90名正常对照的HLA-DRB1等位基因频率.结果(1)在不考虑白细胞变化的情况下,GD患者DRB1*08基因频率明显高于对照组(P＜0.01,RR=2.62),DRB1*07基因频率明显低于对照组(P＜0.01,RR=0.24).(2)GD白细胞减少组DRB1*08(P＜0.01,RR=4.17)和DRB1*15(P＜0.05,RR=1.69)基因频率较对照组显著增高,DRB1*07基因频率明显低于对照组(P＜0.01,RR=0.13).(3)GD白细胞减少组DRB1*08基因频率(P＜0.01)和DRB1*15基因频率(P＜0.05)均明显高于白细胞正常组,DRB1*09(P＜0.05)基因频率明显低于白细胞正常组.结论天津地区汉族GD白细胞减少易感性与HLA-DRB1*08,HLA-DRB1*15基因频率增加有关;GD的保护性与HLA-DRB1*07基因频率减少有关.     

甲状腺特异转录因子1、主要组织相容性复合物1在自身免疫性甲状腺疾病时甲状腺组织中的表达

应用免疫组化检测主要组织相容性复合物1(MHC-1)、甲状腺特异转录因子1(TTF-1)在Graves病(GD)、桥本甲状腺炎(HT)甲状腺滤泡上皮的表达.结果显示MHC-1、TTF-1在GD、HT的表达显著强于对照组,这种表达差异在GD、HT的发病中可能起重要作用.     

CD54、CD80、HLA-DR在Graves病和桥本甲状腺炎甲状腺滤泡上皮中的表达

目的观察正常人、Graves病（GD）、桥本甲状腺炎（HT）、甲状腺组织CD54、CD80、HLA—DR的表达。方法免疫组化SP法测定上述各组甲状腺组织CD54、CD80、HLA—DR的表达水平。结果正常甲状腺组织甲状腺滤泡上皮细胞（TEC）几乎不表达CD54和HLA—DR，而GD组和HT组有较高水平的CD54和HLA-DR表达，以HT表达水平最高，3组间比较差异有统计学意义（均P〈0．01）；HT患者甲状腺CD80表达高于GD组和正常人，差异有统计学意义（均P〈0．01），GD组甲状腺CD80表达很低，与正常人相比差异没有统计学意义。结论CD54和HLA-DR在GD和HT的发病机制中起重要作用，CD80在HT的发病中起重要作用。     

细胞角蛋白19、波形蛋白、血管内皮生长因子C和环氧化物酶2在Graves病和桥本甲状腺炎的表达及意义

目的 探讨细胞角蛋白19(CK-19)、波形蛋白(vimentin)、血管内皮生长因子C(VEGF-C)和环氧化物酶2(COX-2)表达在弥漫性毒性甲状腺肿(diffuse toxic goiter,Graves病,GD)和桥本甲状腺炎(HT)发生和发展中的作用.方法 采用免疫组织化学SP法检测57例Graves病和58例桥本甲状腺炎中CK-19、波形蛋白、VEGF-C、COX-2的表达.结果 CK-19、VEGF-C、COX-2表达于甲状腺滤泡上皮细胞的胞浆,波形蛋白在间质和滤泡上皮细胞的胞浆均有表达.HT中CK-19和VEGF-C表达阳性率(86.2％、96.6％)及阳性强度均高于GD(43.9％、56.1％,均P＜0.05),波形蛋白和COX-2表达的阳性率(100％、93.1％)近似于GD( 100％、91.2％),但COX-2的阳性强度明显高于GD(均P＜0.05).CK-19阳性率在GDⅢ型(81.3％)明显高于Ⅰ型(15.8％)和Ⅱ型(40.9％),HT的P型(100％)明显高于L型(66.7％).VEGF-C阳性率在GDⅢ型(87.5％)明显高于Ⅰ型(36.8％)和Ⅱ型(50.0％,均P＜0.05).结论 综合检测CK-19、波形蛋白、VEGF-C和COX-2四种免疫学指标,对探讨GD和HT发生、发展过程及预后评估具有一定的参考价值.     

Immunohistochemical analysis of bcl-2, Bax and Bak expression in thyroid glands from patients with Graves' disease

In order to clarify the role of apoptosis and the expression of Bcl-2 family proteins in the pathology of Graves' disease (GD), we evaluated the apoptosis by in situ end-labeling of fragmented DNA and the expression of Bcl-2, Bax and Bak by immunohistochemistry in thyroid tissues from 20 patients with GD and in normal thyroid tissues from 6 patients with follicular adenoma (N). Apoptotic nuclei were found in thyrocytes and in germinal center of lymphoid follicles. Bcl-2 was strongly expressed in both GD and N thyrocytes. Bax was not expressed in either GD or N thyrocytes. Bak was expressed in thyrocytes from 5 of 20 patients with GD, while it was detected in all N thyrocytes. In lymphoid follicles Bcl-2 was expressed in the mantle zone, while Bax and Bak were both expressed in the germinal center. The percentage of apoptotic nuclei in GD thyrocytes was low (0~3.6%), and negatively correlated with the weight of the thyroid glands resected (rs = -0.43, P<0.05). It was greater in Bak-positive GD thyrocytes than in Bak-negative ones (mean +/- SD; 1.7 +/- 0.7% vs. 0.7 +/- 0.9%, P<0.05). These findings suggest that the differential expression of Bcl-2 family proteins in both thyrocytes and lymphoid follicles may be involved in the pathology of GD.     

Expression of intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 and homing factor CD44 after engraftment of Graves' lymphocytes in xenotransplanted human thyroid tissue in athymic nude mice.

The expression of adhesion molecules on thyrocytes and endothelium cells plays an important role in the pathogenesis of Graves' disease (GD). The intercellular adhesion molecule-1 (ICAM-1), the vascular cell adhesion molecule-1 (VCAM-1), and the homing receptor CD44 are responsible for the specific migration of lymphocytes in autoimmune thyroid diseases (AITD) (homing). Eight weeks after transplantation of thyroid tissue from 26 patients with nonautoimmune thyroid disease (nontoxic nodular goiter [NTG]) into nude mice, peripheral (PBL) and intrathyroidal lymphocytes (ITL) from 14 patients with NTG and 12 patients with GD were grafted into the animals. Two days after lymphocyte engraftment, the thyroid transplants were examined histologically (HE) and immunohistologically stained with monoclonal antibodies directed against ICAM-1, VCAM-1, and CD44. After injection of GD lymphocytes, thyroid transplants expressed significantly more ICAM-1, VCAM-1, and CD44 than after injection of NTG lymphocytes. This expression was even more pronounced after grafting of GD intrathyroidal lymphocytes. Our data demonstrate that only GD lymphocytes induce the expression of adhesion molecules and homing factor CD44, both of which play an important role in the migration of lymphocytes and induction of the autoimmune process.     

Evaluation of insulin-like growth factor II, cyclooxygenase-2, ets-1 and thyroid-specific thyroglobulin mRNA expression in benign and malignant thyroid tumours

OBJECTIVE: We evaluated three markers (insulin-like growth factor II (IGF-II), cyclooxygenase-2 (COX-2) and ets-1) of thyroid growth stimulation and cell transformation together with a thyroid-specific marker (thyroglobulin (Tg)) for their potential to differentiate benign and malignant follicular thyroid neoplasia (FN). DESIGN AND METHODS: mRNA expression levels were determined by real-time PCR in 100 snap-frozen thyroid samples: 36 benign thyroid nodules with different histology and function (19 cold (CTN) and 17 toxic thyroid nodules (TTN)), 36 corresponding normal thyroid tissues of the same patients, eight Graves' disease (GD) thyroids, 10 follicular thyroid carcinomas (FTC) and 10 papillary thyroid carcinomas (PTC). RESULTS: Mean IGF-II and COX-2 levels were not significantly altered between benign and malignant thyroid nodules (IGF-II) or nodular (FTC, TTN, CTN) and normal thyroid tissues (COX-2). In contrast, eight- to tenfold upregulation of ets-1 was observed in PTC and three- to fourfold upregulation of ets-1 was observed in FTC (and GD) compared with benign thyroid nodules and normal thyroid tissues. In addition, thyroglobulin mRNA expression was markedly downregulated (50- to 100-fold) in FTC, PTC and GD samples compared with benign nodular and normal thyroid tissues. Hence an ets-1/Tg ratio >20 distinguished differentiated thyroid cancer from benign nodular or normal thyroid tissue. We then studied ets1- and Tg mRNA expression levels in fine needle aspiration cytology (FNAC) samples. However, in a consecutive series of 40 FNAC samples only equivocal results were obtained on 38 benign and two malignant (FTC) thyroid tumour samples. CONCLUSIONS: Upregulation of ets-1 and downregulation of Tg mRNA expression occur in differentiated thyroid cancer and may facilitate pre-operative identification of thyroid malignancy depending on further evaluation of these potentially promising markers in a larger series of benign and malignant thyroid tumours and their FNAC samples.     

甲状腺肿瘤组织中雌激素受体β1和β2的表达及其意义

采用RT-PCR检测10例甲状腺乳头状腺癌(PTC)和10例癌旁正常甲状腺组织雌激素受体(ER)β1、ERβ2以及ERα的表达,同时用免疫组织化学方法检测107例甲状腺癌、56例甲状腺腺瘤和10例正常甲状腺组织中ERβ1和ERβ2表达,并结合甲状腺癌ERα表达及其它主要临床病理参数进行分析.结果显示,ERα mRNA在PTC组织的表达明显高于癌旁正常组织,ERβ2 mRNA在PTC组织的表达明显低于癌旁正常组织(P＜0.05);正常甲状腺组织、甲状腺腺瘤和甲状腺癌中3种ER的表达差异均有统计学意义( P＜0.05);ERβ1和ERβ2在甲状腺癌中的表达与其组织类型有关;甲状腺组织中ERβ1和ERβ2的表达与ERα呈低、中度负相关(ERβ1:r=-0.206,P=0.042;ERβ2:r=-0.545,P＜0.01);ERβ2的表达还与淋巴结转移以及TNM分期有关.提示ERβ2的表达缺失可能导致甲状腺滤泡上皮细胞的增生和癌变以及肿瘤的演进,并且有一定的预后评估价值.     

Cathepsin B activity and protein levels in thyroid carcinoma, Graves' disease, and multinodular goiters.

Cathepsin B (CB) is involved in the hydrolysis of thyroglobulin (Tg) and thought to be regulated by thyroid stimulating hormone (TSH) in the normal thyroid. Our analyses of 91 thyroid tissues from 71 patients with Graves' disease (GD), multinodular goiter (MNG), papillary carcinoma (PC), or follicular carcinoma (FC), demonstrated a 2-fold increase in expression of CB in GD and an average increase of 1.5-fold in MNG (varying from 10-fold below normal to 6-fold above normal in MNG nodules), as might be predicted by the altered functional status of thyroid follicular cells in those diseases. However, CB activity was not downregulated in conjunction with the known "blocking effect" of malignancy on many thyroid functions, but rather increased on average 9-fold in papillary carcinomas (n = 33), and also showed a marked increase in 2 follicular carcinomas. Activity measurements were confirmed by CB protein detection on Western blot with moderately increased CB protein levels demonstrated in GD, variable expression in nodules of MNG, and markedly increased protein expression in carcinomas. In all diseased states, increased protein was detected primarily as overexpression of the 27 kd heavy chain of 2-chain mature CB and less frequently as overexpression of 31 kd single-chain mature CB. However, an additional 35 kd protein form was noted in 3 of 9 PCs, 1 of 2 FCs, and 1 of 4 GD cases but in none of 10 MNG cases. In conjunction with elevated CB activity plus additional protein bands on Western blots, altered patterns of CB immunohistochemical staining were observed, irrespective of the type of thyroid disease, suggesting certain common changes in CB expression, posttranslational processing, and vesicular trafficking. In summary, GD and MNG thyroid tissues demonstrated altered CB expression in keeping with predicted functional changes in thyroid follicular cells, while increased CB expression in carcinomas indicated a more pathological role for CB in thyroid cancers, possibly related to the processes of invasion or metastasis.     

Characterisation of DEHAL1 expression in thyroid pathologies

Iodotyrosine dehalogenase 1 (DEHAL1) is a transmembrane protein involved in the recycling of iodide in the human thyroid. The aim of the present study was (I) to investigate whether DEHAL1 expression is different in differentially functioning thyroid pathologies and (II) to evaluate DEHAL1 as a possible marker of thyroid cell differentiation. DESIGN AND METHODS: Real-time PCR for DEHAL1 and its isoform DEHAL1B was performed in a series of 105 thyroid specimens, including toxic thyroid nodules (TTN), Graves' disease (GD) thyroids, benign cold thyroid nodules (CTN), normal thyroid tissues and thyroid cancers (follicular thyroid carcinomas (FTC), papillary thyroid carcinomas (PTC), partially differentiated thyroid cancers (PDTC) and anaplastic thyroid carcinomas (ATC)). In addition, DEHAL1 protein expression was studied by immunohistochemistry in 163 benign and malignant thyroid pathologies and normal thyroids. RESULTS: (I) The highest DEHAL1 mRNA levels were found in GD thyroids, while downregulation of DEHAL1 and DEHAL1B mRNA occurred in PTC and ATC (P<0.001 and <0.05 respectively); (II) DEHAL1 protein was overexpressed in TTNs and GD thyroids with predominant apical staining in all samples; (III) a weaker and patchy staining pattern was found in CTNs and normal thyroids; (IV) in differentiated thyroid cancers (FTC and PTC), a diffuse cytoplasmic DEHAL1 expression was found; and (V) in PDTC and ATC, DEHAL1 expression was faint or absent. CONCLUSION: Upregulation of DEHAL1 protein expression and sublocalisation of DEHAL1 at the apical cell pole in TTNs and GD thyroids is consistent with increased thyroid hormone turnover during thyrotoxicosis. Diffuse cytoplasmatic localisation or downregulation of DEHAL1 expression in thyroid cancers suggests alteration or loss of DEHAL1 function during thyroid cell dedifferentiation.