Ultrastructural localization of thyroid peroxidase, hydrogen peroxide-generating sites, and monoamine oxidase in benign and malignant thyroid diseases

Despite thyroid tissue heterogeneity, biochemical and morphological features have been associated with certain thyroid diseases. We analyzed the ultracytochemical localization of thyroperoxidase (TPO), TPO-associated hydrogen peroxide-generating sites (H(2)O(2) sites), and monoamine oxidase (MAO) in terms of morphology and biochemical TPO activity in abnormal thyroids. We examined 11 cases of nontoxic multinodular goiter, 5 cases of Hashimoto's thyroiditis, 1 case of oncocytic (Hürthle or oxyphilic cell) adenoma, 5 cases of Graves' disease, 4 cases of papillary carcinoma, and 4 cases of perinodular normal tissue. In the perinodular tissue, TPO was detected mainly in the nuclear envelope, rough endoplasmic reticulum (RER), and subapical vesicles, but not in the apical surface. In multinodular goiter, heterogeneous TPO reactivity ranging from almost null to strongly positive was detected in similar locations as in the perinodular tissue, and was absent in the microvilli. Follicular cells from Hashimoto's thyroiditis displayed TPO in the nuclear envelope and the scarce RER. Remarkably, oncocytic cells from both Hashimoto's thyroiditis and oncocytic adenoma, typically packed with mitochondria, displayed evident TPO reaction exclusively in mitochondrial cristae. In Graves' disease, the nuclear envelope, enlarged RER, and apical vesicles were strongly TPO positive, and microvilli also exhibited TPO activity. Papillary carcinoma cells were negative for TPO. The localization and characteristics of TPO activity in the H(2)O(2) sites were similar to that of TPO in all tissues. MAO was positive in mitochondria of perinodular tissues, multinodular goiter, and oncocytes and negative in Hashimoto's thyroiditis and Graves' disease. Interestingly, MAO was intensely positive in the nuclear envelope of papillary carcinoma but unreactive in mitochondria. Biochemical TPO activity was increased in multinodular goiter and Graves' disease. In conclusion, several changes in ultracytochemical characteristics of TPO, H(2)O(2) sites, and MAO were associated with thyroid disease. Nonmalignant oncocytic cells exhibited an unusual mitochondrial location of TPO and H(2)O(2) sites. The distribution of MAO in nuclear envelope of papillary carcinoma cells could be a further feature of malignancy.     

促甲状腺激素、甲状腺过氧化物酶抗体和促甲状腺激素受体抗体检测在甲状腺疾病中的应用价值

探讨促甲状腺激素（TSH）、甲状腺过氧化物酶抗体（TPOAb）、促甲状腺激素受体抗体（TRAb）在甲状腺疾病中的诊断价值。对44例Graves病（GD）、29例桥本甲状腺炎（HT）、13例亚甲炎患者和26例正常对照,采用电化学发光免疫分析法测定TSH、TPOAb、TRAb的含量,分析其在几组病例中的意义。三病例组的TSH、TPOAb、TRAb含量与正常对照组比均有显著差异（P〈0.01）;HT组TPOAb的测值和阳性率又显著高于GD组,而GD组TRAb的测值和阳性率都显著高于HT组,经统计学检验两者均有显著意义（P〈0.05）。结果说明,TSH是甲状腺功能的非常敏感的特异性参数,TPOAb、TRAb的检测对鉴别诊断GD和HT有着重要意义。     

Marked increase of CD5 + B cells in hyperthyroid Graves' disease

We examined the proportions of B lymphocytes bearing CD5 cell surface antigen (CD5+ B cells), which are capable of making autoantibodies, in peripheral blood from patients with various thyroid diseases. The level of CD5+ B cells was markedly increased (>9.0%) above the normal range (0.5-7.7%) in untreated, hyperthyroid patients with Graves' disease, although about 10% of the patients had no detectable serum thyroid-stimulating hormone (TSH) receptor antibody (TRAb). However, the levels of CD5+ B cells were normal in untreated patients with destructive thyrotoxicosis due to aggravation of Hashimoto's thyroiditis or subacute thyroiditis. In patients with stimulated hyperthyroid Graves' disease the levels of CD5+ B cells were correlated with those of thyroid hormones and TRAb, all significantly increased. However, once hyperthyroidism was controlled by anti-thyroid drugs, CD5+ B cells were decreased, followed in turn by reduction of TRAb. We conclude that the proportion of CD5+ B cells is useful as a therapeutic index and for diagnosis of Graves' disease and its differentiation from destruction-induced thyrotoxicosis.     

Thyroid follicular cell function after non-lethal complement membrane attack

Terminal complement complexes have been identified around thyroid follicles in Graves' disease and Hashimoto's thyroiditis, and the concentrations of such complexes are increased in the sera of these patients, suggesting a role for complement activation and membrane attack complexes (MAC) in autoimmune thyroiditis. This has been investigated further using cultured human and rat thyroid cells. Thyrocytes were resistant to lysis by homologous complement, in contrast to the effects of heterologous (rabbit) complement. The formation of non-lethal amounts of MAC, using reactive lysis or classical pathway activation, significantly reduced cAMP production by these cells in response to thyroid-stimulating hormone (TSH) (P less than 0.01); similar effects were seen with thyroid-stimulating antibodies. Thyroid cells were able to recover rapidly from complement attack after washing and incubation for 30 min. Non-lethal MAC formation also resulted in reactive oxygen metabolite production, detected by luminol-dependent chemiluminescence in three out of five thyroid cell preparations tested. Ionomycin, but not TSH, also stimulated reactive oxygen metabolite production. These results suggest that repeated or continuous sub-lethal complement attack on thyroid cells may exacerbate hypothyroidism in Hashimoto's thyroiditis, or partially counter the effects of thyroid-stimulating antibodies in Graves' disease. Furthermore, the production of reactive oxygen metabolites in these circumstances could increase the intra-thyroidal inflammatory response; oxygen radical scavenging by anti-thyroid drugs (which are concentrated by thyrocytes) may account in part for the amelioration of thyroiditis observed with such treatment.     

CD4~+CD25~+调节性T细胞在自身免疫性甲状腺疾病中数量和功能变化

目的:探讨自身免疫性甲状腺疾病患者外周血中CD4+CD25+调节性T细胞(Tregs)的数量和功能变化。方法:采用化学发光法测定20例初发Graves’病人、20例初发桥本甲状腺炎(HT)患者及20例健康体检者血清中促甲状腺素(TSH)、总三碘甲状腺原氨酸(TT3)、总甲状腺素(TT4)、甲状腺球蛋白抗体(TgAb)和甲状腺过氧化酶抗体(TPOAb)的水平;用流式细胞仪分析外周血单个核细胞(PBMC)中CD4+T细胞及CD4+CD25+Tregs的数量;采用磁珠分选技术分选5例HT病人和5例健康体检者PBMC中CD4+CD25+Tregs和CD4+CD25-T细胞,采用MTT法检测CD4+CD25+Tregs对自身CD4+CD25-T细胞增殖的抑制作用;提取各组PBMC的总RNA,经Real time-PCR检测TGFβ-1、Foxp3 mRNA的表达水平。结果:流式细胞检测结果显示,初发Graves’病人、初发HT患者外周血PBMC中CD4+T细胞数量与正常人比较无差异(P<0.05);初发HT患者外周血PBMC中CD4+CD25+Tregs占CD4+T细胞的比率为(1.55%±0.49%),明显低于正常对照组(2.86%±1.04%)(P<0.05);初发Graves’病人外周血PBMC中CD4+CD25+Tregs占CD4+T细胞的比率为(3.25%±0.97%),与正常对照组(2.86%±1.04%)相比无显著性差异(P<0.05)。MTT结果显示,初发HT患者CD4+CD25+Tregs对自身CD4+CD25-T细胞增殖的抑制百分率为15.7%±5.36%,与正常组(41.7%±9.87%)相比显著降低(P<0.05)。Real time-PCR结果显示,初发Graves’病人、初发HT患者PBMC的TGFβ-1 mRNA表达水平分别为(0.37±0.10)和(0.43±0.09),均明显低于正常对照组(1.02±0.04)(P<0.05);初发Graves’病人、初发HT患者PBMC的Foxp3 mRNA表达水平分别为0.62±0.09和0.42±0.29,均明显低于正常对照组(0.99±0.17)(P<0.05)。结论:本研究结果提示,HT患者外周血中CD4+CD25+Tregs的数量和功能明显降低。Graves’病和HT患者外周血PBMC中TGFβ-1、Foxp3 mRNA表达水平明显降低。     

Characterization of different types of experimental autoimmune thyroiditis in the Buffalo strain rat

We have compared features of experimental thyroiditis in the Buffalo strain rat induced by neonatal thymectomy, immunization with rat thyroglobulin (Tg) and complete Freund's adjuvant, and subcutaneous administration of trypan blue or 3-methylcholanthrene. The disease produced by neonatal thymectomy resulted in significantly worse thyroiditis and higher antibody levels than the other models and shared other features in common with Hashimoto's thyroiditis. In particular TSH levels were elevated, Tg antibodies had a restricted subclass distribution, the thyroid was infiltrated by both B cells and T cells (comprising equal numbers of W3/25 and Ox8 positive cells) and thyroid follicular cells were Ia antigen-positive in some of the thymectomy group animals. In the other forms of thyroiditis, the thyroid infiltrate was mainly composed of macrophages or dendritic cells and B cells. Different pathogenic mechanisms are probably involved in these models; the disease produced by neonatal thymectomy shows the closest similarity to Hashimoto's thyroiditis.     

Evidence of HTLV-I in thyroid tissue in an HTLV-I carrier with Hashimoto's thyroiditis

Human T-lymphotropic virus type I (HTLV-I) protein and messenger RNA (mRNA) for HTLV-I were examined in thyroid tissues from two patients with Hashimoto's thyroiditis and serum anti-thyroid antibody. The virus envelope protein and signals for the mRNA were detected in many of the follicular epithelial cells of the thyroid tissue from one of the patients, respectively, by immunohistochemistry and in situ hybridization. PCR-Southern blotting revealed the presence of HTLV-I DNA in the thyroid tissue, in which the viral protein and mRNA were detected, although no virus particles were found in the epithelial cells by electron microscopy. HTLV-I virus was not present in the thyroid tissue from the second patient. The present findings suggest that infection of thyroid tissue with HTLV-I is associated with the pathogenesis of Hashimoto's thyroiditis in some patients.Abbreviations HTLV-I Human T-lymphotropic virus type 1     

Latent autoimmune thyroiditis in untreated patients with HCV chronic hepatitis: a case-control study

In order to establish a relationship between Hepatitis C virus (HCV) chronic infection and autoimmune thyroiditis, 97 untreated patients with biopsy-proven HCV chronic hepatitis and 97 controls were studied. An ultrasound examination of the thyroid and an assay of serum thyroid-stimulating hormone (TSH), thyroid hormones and anti-thyroid antibodies were performed in all cases. The overall prevalence of thyroid abnormalities was higher in patients than in controls (17 vs. 4%, P<0.01) and the prevalence of anti-thyroid antibodies was significantly different between the two groups (P<0. 02). HCV patients with (n=13) compared to HCV patients without anti-thyroid antibodies (n=84) were older, predominantly female, and more frequently had increased serum TSH levels or a hypoechogenic pattern of the thyroid gland, while Knodell's score and prevalence of cirrhosis were similar. Latent autoimmune thyroiditis is more frequent in untreated HCV patients than in controls. This finding raises questions about the mechanism of autoimmunity induced by HCV and provides an explanation for the high rate of overt autoimmune thyroiditis during interferon treatment in these patients.     

运用ROC曲线方法评价TRAb、TPO-Ab和TGA在Graves'病和HT鉴别诊断中的意义

目的:应用临床诊断性能(ROC)曲线方法评价TSH受体抗体(TRAb)、甲状腺过氧化物酶抗体(TPO-Ab)和甲状腺球蛋白抗体(TGA)在Graves'病(格雷夫斯病)和桥本甲状腺炎鉴别诊断中的意义.方法:以甲状腺细针穿刺细胞学检查结果作为诊断金标准,以采用化学发光法测定63例自身免疫性甲状腺病患者血清的TRAb、TP...     

Thyroid function tests

About 80% of thyroid disease consists of thyroid-specific autoimmune diseases, Hashimoto's disease and Grave's disease. To diagnose thyroid diseases, testings for (1) thyroid function and (2) pathogenetic autoantibodies are indispensable. To assess thyroid function, serum hormone concentrations, such as TSH, FT4 and FT3 are measured. Among these hormones, serum TSH concentrations are the most reliable and informative regarding thyroid function, correcting indicating a hyperthyroid, euthyroid or hypothyroid state. Therefore, TSH measurement appears to be the first choice in selecting the hormone determination. Reference intervals for normal healthy subjects of TSH are around 0.4-5.0 microU/ml. The second choice for thyroid function assessment are FT4 which supersedes total T4(TT4). TT4 is affected by changes in serum thyroid hormone binding proteins(TBG, TTR, Albumin). For example, euthyroid pregnant women whose serum TBG are physiologically higher than those of non-pregnant women show augmentation of TT4. However, FT4 depicts within reference intervals, although measurement of FT4 alone is unable to detect any abnormality of thyroid hormone binding proteins. According to its plasma concentration and binding affinity, FT3 measurement deserves no more significance than T3. Another important test for thyroid diseases is to detect serum autoantibodies against thyroid tissues, such as TgAb, TPOAb. Much more important is TSH receptor antibody which differentiates Graves' disease from Hashimoto's thyroiditis. In patients who show hyperthyroidism and some very uncommon hypothyroidism, TSH receptor antibodies should be measured. Three indicators are available as routine tests; TRAb measured by radioreceptor assay; TSAb determined by bioassay using cultured porcine thyroid cells. Usually, TRAb activity clinically correlates well with TSAb. TSBAb was initially discovered in patients with severe hypothyroidism with atrophic thyroid gland. TSBAb blocks thyroid stimulating activity of TSH and consequently causes severe hypothyroidism. TRAb and TSAb are very useful to diagnose and follow patients with Grave's disease.     

Dipeptidyl aminopeptidase IV staining of cytologic preparations to distinguish benign from malignant thyroid diseases.

Staining for dipeptidyl aminopeptidase IV (DAP IV [EC: 3.4.14.5]) activity was applied to aspiration biopsy specimens or imprint preparations of surgical biopsy specimens from thyroid tumors. Material was obtained from 55 patients with histologically proven thyroid diseases: 9 with papillary carcinoma, 5 with follicular carcinoma, 11 with follicular adenoma, 13 with adenomatous goiter, 8 with Hashimoto's thyroiditis, and 9 with other benign conditions. Most tumor cells, follicular lumina in cell clusters, and intranuclear inclusions were strongly positive for DAP IV in all examples of papillary or follicular carcinoma. In contrast, only a few epithelial cells were labeled for DAP IV in follicular adenoma and adenomatous goiter. Some Hürthle cells in Hashimoto's thyroiditis also were positive for DAP IV. When a DAP IV scoring system based on the percentage of positive cells and staining intensity was used, all benign tissues except one (from a follicular adenoma) were found to have extremely low scores. These results indicate that staining for DAP IV activity is a simple but useful tool to aid in distinguishing benign from malignant thyroid neoplasms.     

Chronic lymphocytic thyroiditis in a cynomolgus macaque (Macaca fascicularis)

Chronic lymphocytic thyroiditis characterized by multifocal follicular lymphoid cell infiltrates with germinal centers, thyroid acinar atrophy and pituitary cell hyperplasia/hypertrophy of the adenohypophysis was detected in a vehicle control, 4-year-old female Cynomolgus macaque in a routine toxicology study. Lymphoid cells of germinal centers were positive for the B-cell marker CD20 by immunohistochemistry (IHC), while remaining lymphocytes were positive for the T-cell marker CD3. Hypertrophied/hyperplastic pituitary cells were positive for thyroid stimulating hormone (TSH) by IHC, consistent with an adaptive response due to removal of hormonal negative feedback from the diseased thyroid gland. Features of this case are similar to chronic lymphocytic thyroiditis in humans, an autoimmune disorder also known as Hashimoto's disease. Chronic lymphocytic thyroiditis with compensatory pituitary changes may occur spontaneously in young, clinically normal cynomolgus macaques and its presence in drug treated animals should be interpreted with caution.     

Ultrastructural localization of endogenous peroxidase activity in benign thyroid diseases

Ultrastructural localization of endogenous thyroid peroxidase under benign pathological conditions such as toxic diffuse goiter, non-toxic multinodular goiter, and adenoma, and in normal tissue was studied. Peroxidase activity was visualized by a cytochemical reaction for electron microscopy. In toxic diffuse goiters and most non-toxic multinodular goiters, reaction product for peroxidase was observed not only in the cytoplasm but also at the external surface of microvilli of follicular cells. In normal thyroid tissues and adenomas, peroxidase was visualized only in the cytoplasm. Peroxidase activity at the external surface of microvilli of the follicular cells was found in the tissues obtained from the goiters which showed "hot" radioiodine scintigram. These findings suggest that follicles with peroxidase activity at the external surface of microvilli in non-toxic multinodular goiter are "autonomous follicles" and that peroxidase at the external surface of microvilli plays some role in active iodine uptake.     

ULTRASTRUCTURAL LOCALIZATION OF ENDOGENOUS PEROXIDASE ACTIVITY IN BENIGN THYROID DISEASES

Ultrastructural localization of endogenous thyroid peroxidase under benign pathological conditions such as toxic diffuse goiter, non-toxic multinodular goiter, and adenoma, and in normal tissue was studied. Peroxidase activity was visualized by a cytochemical reaction for electron microscopy. In toxic diffuse goiters and most non-toxic multinodular goiters, reaction product for peroxidase was observed not only in the cytoplasm but also at the external surface of microvilli of follicular cells. In normal thyroid tissues and adenomas, peroxidase was visualized only in the cytoplasm. Peroxidase activity at the external surface of microvilli of the follicular cells was found in the tissues obtained from the goiters which showed "hot" radioidine scintigram. These findings suggest that follicles with peroxidase activity at the external surface of microvilli in non-toxic multinodular goiter are "autonomous follicles" and that peroxidase at the external surface of microvilli plays some role in active iodine uptake.     

过氧化物酶缺陷型激素合成障碍性甲状腺肿的核医学特征

用放射免疫法测定过氧化物酶缺陷型激素合成障碍性甲状腺肿病人血清TT3,TT4,FT3,FT4,TGAb,TMAb,用高敏感免疫放射法测定TSH,并进行碘过氨酸钾释放实验和~(99m)锝甲状腺静态显像,实验显示:该类病人血清甲状腺素水平降低,TSH水平升高,TGAb呈阴性反应;碘过氯酸钾释放试验呈阳性反应;核素显像显示甲状腺均匀性肿大,摄取~(99m)锝功能增加,结果表明:该类病人甲状腺细胞合成甲状腺素的关键酶—过氧化物酶活性低下,使进入甲状腺细胞的碘离子氧化为碘分子的过程受到抑制,该酶活性降低使甲状腺素水平降低,反馈性刺激垂体TSH分泌增加,在TSH刺激下,甲状腺增生肿大,摄取~(131)碘和~(99m)锝功能增加,SPECT静态显像甲状腺放射性分布浓聚,与桥本甲状腺炎鉴别诊断,测定血清甲状腺自身抗体TGAb,TMAb具有重要意义     

Thyroid autoantigens and human T cell responses.

We investigated the ability of T cells from patients with Hashimoto's thyroiditis and with Graves' disease as well as control donors to proliferate in response to thyroid peroxidase (TPO) and thyroglobulin using (i) lymphoid cells from different lymphoid organs; (ii) unfractionated or CD8- depleted lymphoid suspensions or T cells + autologous low density cells (LDC); (iii) 200-microliters well cultures and 20-microliters hanging-drop microcultures; and (iv) intact TPO and thyroglobulin, denatured thyroglobulin and 12 synthetic peptides predicted on the basis of the amino acid sequence of TPO to be T cell epitopes. In 200-microliters well cultures, proliferative responses (assessed in terms of 3H-thymidine uptake) to intact TPO or thyroglobulin, digested thyroglobulin or synthetic TPO peptides were not significantly different in unfractionated or CD8-depleted lymphoid suspensions from blood, thyroid or lymph nodes of TPO/thyroglobulin autoantibody-positive patients, autoantibody-negative patients or control donors. In contrast, blood T cells from some high titre patients with Hashimoto's thyroiditis (but not from healthy individuals) proliferated in response to intact thyroglobulin or TPO presented by autologous LDC in hanging-drop microcultures. Heat denatured thyroglobulin (with which thyroglobulin autoantibodies do not interact) did not stimulate proliferation and this observation, together with the ability of T cells from some patients to respond to intact thyroglobulin in the absence of LDC, indicated that thyroglobulin-specific B cells may be involved in antigen presentation. As we were unable to demonstrate proliferation by blood T cells + LDC from all thyroglobulin antibody-positive patients with Hashimoto's thyroiditis, our studies suggest that the presence of sufficient precursor T cells, as well as the number and type of antigen-presenting cells, are critical for T cell proliferative responses to human TPO and thyroglobulin.     

Thyroid blocking antibodies in thyroiditis

Serum from a woman with a history of Hashimoto's thyroiditis, who had given birth to two children with congenital hypothyroidism, contained potent TSH blocking activity. Immunoglobulin preparation from this serum abolished completely TSH-stimulated cAMP production in human thyroid membranes. The blocking activity was associated with the IgG fraction absorbed to and eluted from a Protein A column. The stimulation of adenylate cyclase by a preparation of thyroid-stimulating antibodies from a patient with Graves' disease was also inhibited by the antibodies. In contrast, no effect was observed upon fluoride-stimulated cAMP production. The data indicate that the antibody activity was directed against the TSH receptor. Immunoglobulin preparations from 22 other patients with Hashimoto's thyroiditis and 16 patients with subacute thyroiditis were examined for the existence of TSH receptor blocking antibodies. A blocking activity was found in two of the 22 Hashimoto patients. No such activity was found in the patients with subacute thyroiditis. It appears that thyroid blocking antibodies sometimes contribute to hypothyroidism associated with Hashimoto's thyroiditis.     

Cytokines, thyroid autoantibody synthesis and thyroid cell survival in culture

In autoimmune thyroid disease lymphoid cells infiltrating the thyroid gland occur in conspicuous aggregates or as a diffusely distributed population invading the thyroid follicles. Consequently cytokines secreted by activated T cells or macrophages could influence neighbouring thyroid cells as well as other lymphocytes. We have investigated this possibility using recombinant cytokines. Thyroid cell survival was assessed in terms of mitochondrial dehydrogenase activity in monolayers exposed to tumour necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-1 (IL-1 alpha and beta) and interleukin-2 (IL-2) in the presence or absence of thyroid-stimulating hormone (TSH). Neither TNF-alpha nor IL-2 affected thyroid cell survival, IFN-gamma was usually inhibitory and IL-1 alpha slightly enhanced cell survival in some experiments. However, the effects were small and variable and were not enhanced by potentially synergistic combinations of cytokines, longer periods of exposure, or different culture conditions. In contrast, IFN-gamma, IL-2 and TNF-alpha inhibited the ability of thyroid lymphocytes from patients with Graves' disease and Hashimoto's thyroiditis to synthesize autoantibodies to thyroid peroxidase (TPO) and thyroglobulin (Tg). Comparison of lymphoid populations isolated by digestion and/or mechanical disaggregation indicated that a population of activated B cells, plasma cells and T cells, intimately associated with thyroid cells since they could only be extracted by digestion, was influenced by cytokines. Our studies suggest that in addition to its well-recognized ability to induce MHC class II antigens on thyroid cells, IFN-gamma may inhibit thyroid cell proliferation and TNF-alpha, IFN-gamma and IL-2 may down-regulate thyroid autoantibody synthesis.     

Expression of intercellular adhesion molecule-1 (ICAM-1) on thyroid epithelial cells in Hashimoto's thyroiditis but not in Graves' disease or papillary thyroid cancer.

In order to study the possible role of antigen-independent adhesion systems in thyroid autoimmunity, we evaluated by indirect immunofluorescence the expression of lymphocyte functional antigen-1 (LFA-1) and its ligand ICAM-1 on mononuclear cell infiltrates (when present) and thyroid follicular cells of four patients with Hashimoto's thyroiditis, 30 with Graves' disease, five with papillary cancer, two with follicular adenoma, and two normal thyroid specimens. The expression of MHC class I and class II antigens was also evaluated. Most mononuclear infiltrates were LFA-1 positive, as expected. A positivity for ICAM-1 on follicular cells was observed in three out of four Hashimoto's thyroiditis specimens; such a phenomenon was totally absent in Graves' disease or any other pathological condition, or in normal tissue. MHC class II expression on thyrocytes was observed in all the patients with Hashimoto's thyroiditis, in 27 out of 30 with Graves' disease and in three out of five papillary cancer specimens.