Circulating cytokeratin 19 fragments in patients with benign nodules and carcinomas of the thyroid gland

Cytokeratin 19 (CK19) is an acidic protein of 40 kDa that is part of the cytoskeleton of epithelial cells and is highly expressed by differentiated thyroid carcinomas, mainly of the papillary subtype. The soluble fragments of CK19 (Cyfra 21.1) can be measured by immunometric assays employing specific monoclonal antibodies. The present study was planned to assess the serum expression of Cyfra 21.1 in patients with benign thyroid nodules and thyroid malignancies. We enrolled 135 patients with histologically proven benign thyroid nodules (n=79) and thyroid carcinomas (n=56). No differences were found in serum Cyfra 21.1 levels between patients with benign nodules and patients with carcinomas. When thyroid malignancies were subdivided according to tumor histology, serum Cyfra 21.1 increased significantly from classical differentiated thyroid carcinomas (papillary or follicular) to less differentiated or undifferentiated carcinomas (poorly differentiated or anaplastic). CK19 release into the bloodstream is strongly related to the apoptotic pathway, and particularly to hyperproliferation-related apoptosis. These pathways characterized anaplastic and poorly differentiated thyroid carcinoma but not classical forms of differentiated thyroid carcinoma. Consequently, Cyfra 21.1 may be regarded as a circulating marker of poorly differentiated and anaplastic thyroid carcinoma. Additionally, a role of Cyfra 21.1 as a dedifferentiation marker in patients with classical differentiated thyroid carcinomas may be postulated and should be explored by further focused studies.     

Significance of AgNOR measurement in thyroid lesions

OBJECTIVE: To evaluate the discriminating potential of AgNOR area measurement and count in thyroid tumors using static cytometry equipment. STUDY DESIGN: Slides were analyzed by a computerized system for image analysis, CAS 200 (Becton & Dickinson, U.S.A.), using the Cell Measurement computer program (CAS 200, Becton & Dickinson). The argyrophilic reaction (NORs) was evaluated with a 400-fold amplification directly from the computer monitor. RESULTS: Thirty-three cases were analyzed for AgNOR staining. The cases studied included 3 goiters, 10 follicular adenomas, 6 Hürthle adenomas, 4 follicular carcinomas, 7 papillary carcinomas, and 3 Hürthle carcinomas. A total of 6,600 nuclei were evaluated. For statistical purposes, lesions were classified as benign and malignant, and both the number and the area of counted NORs showed very similar values. The NORs median among 19 benign tumors was 1.484 (SD +/- 0.265) and of 14 malignant tumors was 1.436 (SD +/- 0.414); the NORs areas were 2.6584 (SD +/- 1.0653) and 2.3643 (SD +/- 0.6320), respectively. CONCLUSION: Our results showed that AgNOR evaluation was not a significant parameter to discriminate between malignant and benign thyroid lesions.     

Expression of obestatin and ghrelin in papillary thyroid carcinoma

Ghrelin and obestatin are two peptide hormones with opposing roles in the control of appetite: orexigenic and anorexigenic, respectively. Loss of appetite is a common, serious complication of many forms of malignancy. The goals of this study were to investigate: (i) whether there are differences in ghrelin and obestatin peptide expression in thyroid tissues from a series of papillary carcinoma cases and normal controls, and (ii) whether there are correlations between tissue ghrelin and obestatin levels in series of papillary carcinoma cases and normal controls. Immunohistochemical analysis showed that in sections of benign human thyroid tissue, anti-ghrelin antibody reacted with intense staining in colloid-filled follicles. In benign thyroid tissues, colloids displayed plentiful dispersion in comparison with papillary microcarcinomas, whereas colloids in malignant thyroid tissues were uncommon. We found markedly lower tissue ghrelin levels in thyroid tissue of patients with papillary carcinomas, compared with normal thyroid tissues (P = 0.001). Immunohistochemical analysis also showed that obestatin in papillary carcinoma stained positively to various degrees. Obestatin tissue levels in papillary carcinomas tended to be slightly higher than those in normal thyroid tissue, but this was not statistically significant (P = 0.29). We also report that thyroid tissue of patients with Hashimoto’s thyroiditis produced ghrelin and obestatin at similar levels as in normal thyroid tissue, even though colloid in Hashimoto’s disease is scarce. We conclude that depressed expression of ghrelin, but not obestatin, is specific to papillary carcinoma, and this difference might constitute a diagnostic tool to differentiate papillary carcinoma from normal thyroid tissue. We currently do not know how these peptides are regulated and what factors are involved in papillary carcinoma, which inhibit the expression of ghrelin but not obestatin. This issue warrants further studies.     

Comparison of morphonuclear features in normal, benign and neoplastic thyroid tissue by digital cell image analysis.

We analyzed the morphonuclear features of thyroid cell nuclei from 10 normal tissues (10 multinodular goiters), 10 benign tissues (10 adenomas) and 10 neoplastic tissues (10 carcinomas--2 follicular, 2 medullary and 6 papillary). These morphonuclear features quantitatively described the nuclear size, DNA content and chromatin texture of 200-400 Feulgen-stained cell nuclei that were digitized by means of a cell image processor. Nuclear DNA estimations revealed that the benign thyroid tumors showed an aneuploidy level that was not different from that of the neoplastic tissues. Morphometric measurements showed a significant and positive correlation between an increase in nuclear area and the development of thyroid pathology--i.e., from normal to a neoplastic stage. We also observed a significant decrease in chromatin condensation and heterogeneity from normal to neoplastic thyroid tissue. In the specific case of thyroid tumors, such criteria were not found sufficient per se for a diagnosis of malignancy. More detailed data banks will be necessary for this purpose.     

Cell population kinetics and DNA content during thyroid carcinogenesis

The proliferation kinetics and DNA content of thyroid follicular cells in rats were studied by autoradiography and cytophotometry. Continuous treatment of animals with methylthiouracil (MTU) results in hyperplasia followed by tumour growth in the thyroid gland. The mitotic index (MI) increases from 0.006 +/- 0.002% in controls to 0.13 +/- 0.06% in hyperplasia and to 0.09 +/- 0.03% in malignant cells. The same is true for the labelling index (LI) which rises from 0.08 +/- 0.003% in controls to 1.4 +/- 1.1% in hyperplasia and to 1.0 +/- 0.6% in follicular adenomas. The S-phase duration (TS) is shortened from 8.0 +/- 1.2 hr in controls to 6.0 +/- 1.4 hr in animals treated for 9 months with MTU and prolonged to 15.4 +/- 2.1 hr in papillary carcinomas. In all MTU-treated animals a decrease in the value of the potential population doubling time (TPD) and thyroid weight doubling time (TD) was observed. The cell loss factor (phi) decreases in animals treated for 3 months with MTU and increases during the stage of tumour growth in the gland (animals treated 12-15 months with MTU). DNA measurements in the nuclei of hyperplastic and neoplastic thyroid tissues reveal cells with values exceeding that in control animals. However, no difference was found in the DNA content between thyroid adenomas and carcinomas, nor between thyroid hyperplasia and neoplasia.     

Production and properties of novel human thyroid cancer specific monoclonal antibodies.

Monoclonal antibodies (TCM-7, -9 and -12) against human thyroid differentiated cancers were established by screening with human thyroid cancers, normal and benign thyroid tissue, and normal human serum protein. A monoclonal antibody (TCM-9) with strong specificity for human thyroid cancer but not for Graves' disease, adenoma or normal thyroid, was shown to recognize a 300 K protein but not to bind to native or mature human thyroglobulin. When TCM-9 was used in immunohistochemical staining tests on more than 30 types of non-thyroid lesions, no reactivity of TCM-9 was observed except with skin immature teratoma, lip squamous carcinoma and stomach adenocarcinoma, which revealed weak reactivities. TCM-9 also showed strong reactivity with two undifferentiated thyroid cancer cell lines and one tissue specimen. Thus TCM-9 is a novel monoclonal antibody against the thyroid cancer.     

Simultaneous expression of keratan sulphate epitope (a sulphated poly-N-acetyllactosamine) and blood group ABH antigens in papillary carcinomas of the human thyroid gland

Summary The monoclonal antibody 5-D-4 recognizes heavily sulphated forms of keratan sulphate epitope. It reacted strongly with the cell surfaces of most thyroid papillary carcinomas from all the individuals examined, independently of the blood group of the patients. Cells of follicular variants of papillary carcinomas were also labelled by 5-D-4. In contrast, no labelling with this antibody was observed in other types of thyroid neoplasms, or in normal tissues. The reactivity of 5-D-4 with papillary carcinomas was markedly reduced or abolished by prior digestion with endo-β-galactosidase keratanase II, or N-glycosidase F. Although keratanase digestion had no effect on 5-D-4 labelling, it revealed the binding sites ofGriffonia simplicifolia agglutinin II (GSA-II), which recognizes terminalN-acetylglucosamine in a limited number of carcinoma cells from some individuals. Blood group ABH antigens, which are simultaneously expressed together with keratan sulphate epitope in cancer cells, were eliminated by digestion with endo-β-galactosidase and N-glycosidase F, but were resistant to keratanase and keratanase II treatment. These results indicate that keratan sulphate oligosaccharides are cancer-associated and are probably oncofoetal antigens, as are the blood group antigens in human thyroid glands. The results suggests that poly-N-acetyllactosamine, which is ubiquitously and consistently produced in papillary carcinomas, is modified in two different ways: sulphation on the 6-position of at least some units of either galactose, orN-acetylglucosamine or both, and decoration of non-reducing termini with the blood group antigens. Along with the endo-β-galactosidase-GSA-II labelling procedure, labelling with 5-D-4 may be a useful diagnostic means for distinguishing papillary carcinoma from other types of thyroid neoplasms.     

Cytologic characteristics of pulmonary papillary adenoma. A case report

BACKGROUND: Pulmonary papillary adenoma is a benign pulmonary neoplasm. Previously pulmonary papillary adenoma was described in terms of immunohistochemistry and ultrastructure. However, there are no previous reports describing the cytologic characteristics of pulmonary papillary adenoma. CASE: A 50-year-old male was admitted for evaluation of a coin lesion in the left upper lung field. Radiologic images showed a solid, round tumor approximately 25 mm in diameter in the left upper lung. Transbronchial needle aspiration biopsy (TBNA) was performed, and small numbers of atypical cells were collected. Adenocarcinoma was suggested clinically, and left upper segmentectomy was performed. The histologic diagnosis was pulmonary papillary adenoma. Imprint cytology of the cut surface of the tumor showed tumor cells arranged in sheets that contained scant or vesicular cytoplasm. The nuclei were oval or round, without obvious anisokaryosis, and their chromatin was fine, without hyperchromasia. Cytologically, the nuclei of the tumor cells in the imprint specimen (38.70 +/- 8.69 microns 2) were uniform in size and similar to the atypical cells in the TBNA specimen (38.29 +/- 11.56 microns 2) but significantly larger than the nuclei of the bronchial cells (23.61 +/- 5.98 microns 2) (P < .0001). CONCLUSION: The cytologic appearance of pulmonary papillary adenoma was characterized morphologically and morphometrically. The possibility of cytodiagnosis by TBNA was suggested.     

Lectin histochemistry in rat thyroid tumours

The thyroid tumours and background goiterous and adenomatous lesions induced in rats with diisopropanolnitrosamine (DIPN) plus methylthiouracil (MTU), and regenerative thyroid tissues after wounding were studied by lectin histochemistry. Ten weeks after cessation of the carcinogen treatments, carcinomas invading the surrounding tissues and blood vessels (13/20) and papillary micronodules (11/20) were formed in the thyroid tissues. In general, the carcinoma lesion was solitary, and the papillary micronodules were multiple in a single thyroid gland. Among the lectins tested, Maclura pomifera (MPA) and Solanum tuberosum (STA) showed specific binding with both carcinoma and papillary micronodule lesions, but not with the background goiterous and adenomatous lesions and regenerative thyroid tissues. The former both lesions showed higher labelling indices with BUdR or 3H-thymidine and poorer thyroglobulin accumulation than the latters, thereby indicating their enhanced proliferative capability and depressed potency of cyto-differentiation. The common cytological and histochemical properties of carcinoma lesions and papillary micronodules allow us to regard the latter as pre-invading carcinoma lesions. The lectins MPA and STA may be, therefore, used as the specific markers of malignancy in rat thyroid carcinogenesis.     

Mechanisms and pathogenesis of thyroid cancer in animals and man

The transformation of the normal fully differentiated thyroid follicular cell to the rapidly growing undifferentiated anaplastic thyroid carcinoma cell involves a number of stages which have been defined morphologically and are now being related to various growth pathways and to molecular biological defects. The two main factors involved in this transformation are growth stimulation and mutagenesis. Growth stimulation alone, through elevated TSH, can lead to the development of thyroid tumours, usually benign, and retaining TSH dependency in some cases. Mutagens alone, if growth is suppressed, do not produce tumours, the combination of mutagens and increased growth is a potent carcinogenic regime. Non-genotoxic carcinogenesis in the thyroid involves growth, without mutagenesis the agent often causes this through affecting one component of thyroid hormone synthesis or metabolism, leading to a fall in thyroid hormone levels and a rise in TSH. Growth stimulation increases the rate of cell division, and therefore increases the chance of a mutation. Continued growth increases the change of subsequent events, in particular loss of heterozygosity in a tumour suppressor gene. The main oncogenes involved in human thyroid carcinogens are ras in the follicular tumour pathway, and ret in the papillary carcinoma pathway. p53 is involved in the progression of either papillary or follicular adenoma to an undifferentiated carcinoma. In experimental thyroid carcinogenesis, ras is again involved, with a link between the mutagenic agent used and the type of ras gene showing mutation. Analysis of the involvement of different growth factors and oncogenes in thyroid carcinogenesis suggests that genes related to the two receptors concerned with normal TSH stimulated growth, TSH receptor and the IGF1 recpptor may be involved in the progression of thyroid tumours of follicular pathology. Several tyrosine kinase receptors with unknown ligands or of uncertain physiological function are linked to papillary carcinoma. The recent large increase in papillary carcinoma of the thyroid in children exposed to fallout from the Chernobyl nuclear accident underlines the importance of understanding the pathobiology of thyroid neoplasia.     

HSP70和P53蛋白在甲状腺乳头状癌中的表达及其与临床病理特征的关系

目的：探讨热休克蛋白70（HSP70）和P53蛋白在甲状腺乳头状癌组织中的表达及其与临床病理特征的关系。方法：应用免疫组织化学Envision法检测35例甲状腺乳头状癌组织、15例甲状腺良性病变组织及15例正常甲状腺组织中HSP70和P53蛋白的表达,分析HSP70和P53蛋白的表达与甲状腺乳头状癌临床病理学特征的关系。结果：甲状腺乳头状癌组织中HSP70和P53阳性表达率明显高于甲状腺良性病变组织及正常甲状腺组织（P〈0．01）。HSP70在甲状腺乳头状癌组织中的表达与是否有颈部淋巴结转移、浸润深度以及AJCC分期密切相关（P〈0．01）,而与年龄、性别、肿瘤的大小以及分化程度无关（P〉0．05）。P53在PTC组织中的表达与组织分化程度、是否有淋巴结转移、浸润深度以及AJCC分期密切相关（P〈0．01）,而与年龄、性别、肿瘤的大小无关（P〉0．05）。HSP70和P53蛋白在PTC组织中的表达呈正相关性（r=0．679,P〈0．01）。结论：HSP70和p53蛋白在PTC中均呈高表达,并有协同作用,两者可作为预测PTC的生物学行为和预后的参考指标：     

Nuclear grooves. How specific are they?

OBJECTIVE: To study the significance of nuclear grooves in thyroid and extrathyroid benign and malignant lesions. STUDY DESIGN: In this retrospective study, 26 cases were selected randomly, consisting of papillary carcinoma of the thyroid (7), papillary adenocarcinoma (9), normal cervical smear (5) and benign bronchioalveolar lavage (BAL) (5). In all cases hematoxylin and eosin- and Papanicolaou-stained smears were studied for nuclear grooves. RESULTS: Except for one case of papillary adenocarcinoma, nuclear grooves were discernible in all cases. The highest number of nuclear grooves was seen in normal cervical smears, followed by papillary carcinoma of the thyroid, papillary adenocarcinoma and BAL smears. CONCLUSION: The number of grooves did not correlate with either benign or malignant conditions. These findings cast serious doubt about the diagnostic significance of nuclear grooves. Probably nuclear grooves are nonspecific findings and should be considered only in association with other diagnostic features.     

Proliferative activity as detected by immunostaining with Ki-67 and proliferating cell nuclear antigen in benign and malignant epithelial lesions of the human parotid gland

OBJECTIVE: A retrospective immunohistochemical study of parotid gland lesions was designed to evaluate the diagnostic and prognostic value of the proliferating cell nuclear antigen (PCNA) and Ki-67 with monoclonal antibodies PC 10 and MIB-1, respectively. STUDY DESIGN: Tissue samples comprised normal parotid gland (N, n = 10), chronic sialadenitis (CS, n = 8), Warthin's tumor (W, n = 10), benign pleomorphic adenoma (BPA, n = 8), mucoepidermoid carcinoma (MEC, n = 13), carcinoma in pleomorphic adenoma (CPA, n = 8) and adenoid cystic carcinoma (ACC, n = 12). The morphometric parameters for PCNA and MIB-1 comprised the PI and MI labelling indices (the numerical percentage of positive nuclei), NAP and NAM (the numerical density of positive nuclei), and NPI and NMI (volume corrected index). RESULTS: The values of MIB-1 parameters increased progressively in benign lesions in comparison with the N group and in malignant neoplasms in comparison with nonneoplastic groups and benign lesions. Values for all parameters in BPA were significantly lower than those in malignant groups. Spearman rank correlation analysis showed a highly positive correlation between the morphometric parameters and severity of the lesions. The mean values of MI and NMI were significantly higher in patients who died of the malignant tumors than in those who survived. The same quantitative parameters for PCNA did not differ significantly from those obtained for MIB-1 and showed similar trends. CONCLUSION: PCNA and MIB-1 indices are reliable markers for discriminating between benign and malignant tumors of the parotid gland, and the parameters PI, MI, NPI and NMI may have prognostic applications.     

DNA flow cytometry of breast carcinoma after acetic-acid fixation

Aqueous acetic acid was used to fix and store specimens of tissue prior to dissociation into nuclear suspensions for flow cytometric quantitation of DNA. The optimum concentration was 20 volumes of glacial acetic acid in 80 volumes of distilled water. Both neoplastic and benign nuclei were easily released from the fixed tissue blocks by slicing and shaking. Residual undissociated tissue was suitable for microscopic examination to confirm its identity. The nuclei fluoresced brightly after staining with propidium iodide, and yielded histograms similar to those from unfixed samples. Acetic-acid fixation resulted in slightly broader G1 and G0 peaks in the DNA histograms in comparison to unfixed cells, but fluorescent debris was less and correlation between the flow cytometric S-phase fraction (SPF) and in vitro thymidine labelling index (TLI) was better than with unfixed cells. Twenty-one of thirty-nine acetic-acid-fixed breast carcinomas had DNA indices in excess of 1.0 (increased nuclear DNA content in comparison to benign cells), and eighteen had DNA indices of 1.0 (normal or near-normal). The SPF was usually in excess of the TLI, but the two were significantly correlated (r = 0.72, P less than 0.0001). However, a significant correlation of SPF with TLI held only for the group with DNA index greater than 1.0. DNA indices greater than 1.0 were associated with high SPF and TLI, and high SPF and TLI each associated with low content of estrogen receptor.     

Psammoma bodies in fine needle aspirate of the thyroid in lymphocytic thyroiditis

Psammoma bodies are concentric, laminated microcalcifications that are regarded as nearly specific markers in the thyroid gland for the presence of papillary carcinoma. While psammoma bodies have been seen rarely in some benign thyroid diseases, there appear to be no reports of psammoma body formation in lymphocytic or Hashimoto's thyroiditis. We report a case of Hashimoto's thyroiditis in which psammoma bodies were identified in a fine needle aspiration specimen of the thyroid and in histologic sections of the right thyroid lobectomy; papillary carcinoma was not found in either specimen. We conclude that psammoma bodies may be seen in any benign process, such as nodular goiter or lymphocytic thyroiditis, that produces reactive papillary hyperplasia of thyroid epithelium, as well as in papillary carcinoma. However, the finding of psammoma bodies in a fine needle aspirate without corroborating cytologic evidence of papillary cancer is still an indication for surgical removal of the thyroid nodule since these structures are reliable markers for occult papillary carcinoma of the thyroid, despite the rarity of their formation in benign diseases.     

The number of nucleoli in benign and malignant thyroid lesions: a useful diagnostic sign in cytological preparations

The slides of fine needle aspiration cytology specimens from 99 cases of cold thyroid nodules with known histology were reviewed and the number of nucleoli per nucleus counted and correlated with the different histopathological groups. Significant differences were observed between benign and malignant thyroid lesions in the number of nucleoli in the cytological material. Lower values were present in nodular goitres and follicular adenomas compared to carcinomas. In benign lesions the majority of nuclei contained one nucleolus and nuclei with two, three or more nucleoli were less frequent than in follicular, papillary, medullary and anaplastic carcinomas. Only one case of follicular adenoma had cells containing three or more nucleoli compared to more than half the cases of follicular carcinoma.