Comparison of nuclear DNA content in primary and metastatic papillary thyroid carcinoma.

Nuclear DNA values were determined in 40 primary papillary thyroid carcinomas, as well as in 52 corresponding local recurrences and metastases were observed either at the time of diagnosis or up to 20 years later. The patient population consisted of 34 survivors and 6 nonsurvivors. In survivors, both the primary tumors and their recurrences and metastases exhibited a majority of cells with DNA values within the normal diploid region, whereas nonsurvivors showed increased and scattered DNA values. In all cases, the primary tumors and the corresponding recurrences and metastases showed similar DNA distribution patterns even if many years had passed between the detection of the primary tumor and the metastases. The results indicate that in papillary thyroid carcinomas, the DNA distribution patterns in the primary tumor and the corresponding recurrences or metastases are generally similar throughout the entire period of disease.

Mouse papillary lung tumors transplacentally induced by N-nitrosoethylurea: evidence for alveolar type II cell origin by comparative light microscopic, ultrastructural, and immunohistochemical studies.

A histogenetic study was designed to evaluate controversial findings on the cell of origin of tubular/papillary lung tumors in mice, i.e., bronchiolar Clara cell versus alveolar type II cell. N-Nitrosoethylurea (0.5 mmol or 0.74 mmol/kg) was given to pregnant C3H (C3H/HeNCr MTV-) and Swiss Webster [Tac:(SW)fBR] mice as a single i.p. injection on Day 14, 15, 16, or 18 of gestation. The offspring were studied at various ages ranging from 7 days to 52 wk. Serial sections of the whole lung (100 to 200 sections per mouse) showed that solid/alveolar and papillary tumors arose from the pulmonary acinus, invading the bronchioles only as the tumors grew. Furthermore, a mixture of solid and papillary patterns within a single module did not represent a merging of two tumors but a progression from the solid to the papillary form. By use of two rabbit antisera against mouse lung surfactant apoproteins found in normal alveolar type II cells, it was shown by the avidin-biotin peroxidase complex procedure, by the peroxidase-antiperoxidase technique, and by indirect immunofluorescence that both solid and papillary tumors contained these proteins that are specific markers for alveolar type II cells. With a rabbit anti-rat Clara cell antiserum, none of the tumors studied was immunoreactive while normal Clara cells were reactive. The nitroblue tetrazolium formazan stain for dehydrogenase enzymes, found particularly in Clara cells, did not reveal these enzymes in any lung tumors from either strain. Ultrastructurally, no typical features of the mature Clara cell were detected in papillary or other pulmonary neoplasms. However, all tumors showed characteristic alveolar type II cell structures such as various stages of lamellar body formation, although these features were less well differentiated in the papillary tumors. Argentaffin dense bodies, representing lysosomes and immature forms of lamellar bodies, were commonly observed in papillary tumors. Some features of the papillary tumors such as cell shape, high glycogen content, and primary cilia were equivalent to those seen in pulmonary epithelial precursor cells during fetal development. With age, the papillary tumors became invasive, accumulated neutral lipids, and developed bizarre cleaved nuclei and lamellated nuclear pseudoinclusions. In conclusion, the papillary lung tumors of the mouse, at least those induced transplacentally by N-nitrosoethylurea, constitute less well-differentiated or poorly differentiated alveolar type II cell adenomas or carcinomas with fetal morphological and biochemical properties.

Gastrin-releasing peptide gene expression in developing, hyperplastic, and neoplastic human thyroid C-cells.

Gastrin-releasing peptide (GRP), the mammalian homolog of bombesin, is often studied as a prototypic neuroregulatory hormone and growth factor, but its own regulation and physiological roles remain to be fully defined. We now demonstrate that the GRP gene is expressed in human thyroidal calcitonin (CT)-containing neuroendocrine cells (C-cells) in an ontogenic pattern similar to its expression in pulmonary neuroendocrine cells and is also expressed at high levels in C-cell hyperplasias and neoplasias (medullary carcinomas of the thyroid). Mean GRP-like immunoreactivity is 20 times higher in 3-week-old to 5-month-old infants than in normal adults, with six of seven infants having GRP levels 6- to 67-fold higher than those in normal adults, the highest levels occurring at 2-2.5 months. CT levels are about 100 times greater than GRP levels at all time intervals, with levels of GRP and CT being linearly correlated (r = 0.98). By RNA blot analysis, GRP mRNAs are increased in neonatal thyroids compared to adult thyroids. In situ hybridization and immunoperoxidase analyses localize GRP mRNAs and peptide to a majority of C-cells in fetuses and neonates, but to only 5-18% of C-cells in normal adults. The majority of developing C-cells have a dendritic morphology, suggesting a paracrine role, although this morphology is not observed in adult C-cells. In addition, for unknown reasons, an increased percentage of C-cells positive for GRP occurs in normal thyroid adjacent to GRP-negative follicular adenomas and papillary carcinomas, an association that we term perineoplastic. We hypothesize that GRP gene expression may play a role in both normal and neoplastic growth processes.

Medullary thyroid carcinomas express chromogranin A and a novel neuroendocrine protein recognized by monoclonal antibody HISL-19.

PURPOSE AND METHODS: A number of endocrine peptides and proteins are expressed by medullary thyroid carcinoma (MTC). The expression of two newly appreciated neuroendocrine tumor markers, chromogranin A (CgA) and the endocrine antigen defined by monoclonal antibody HISL-19, was determined in 14 MTCs by immunohistology to evaluate the clinical utility of these markers in the diagnosis of MTC. Papillary, follicular, and undifferentiated thyroid tumors were also evaluated along with an MTC cell line. The same tissues were evaluated with antibodies to human calcitonin. RESULTS: All human calcitonin antibodies were found to react with the MTCs. In addition, all MTCs were reactive for CgA and the antigen detected by antibody HISL-19. CgA was generally present in the human calcitonin-containing cells, whereas the HISL-19 antigen had a more distinctive distribution. The other thyroid tumors failed to show reactivity with any of the three antibodies. CONCLUSION: Our results demonstrate that, in addition to human calcitonin, MTCs commonly express CgA and the antigen defined by antibody HISL-19. Our observations thus add to the repertoire of endocrine substances produced by MTC. These studies also demonstrate the clinical value of immunohistologic procedures for two novel antigens in distinguishing MTCs from other thyroid tumors.

Immunohistochemical analysis of benign and malignant papillary lesions of the breast.

The histocytological diagnostic criteria and recently developed immunohistochemical procedures selective for either the epithelial or the myoepithelial mammary cells have been tested in a series of 60 cases of papillary lesions of the breast. These included 15 benign solitary intraductal papillomas, 41 papillary carcinomas (29 pure and 12 associated with other types of in situ or invasive ductal carcinoma), and four cases of "suspected" papillary carcinomas. Markers for epithelial cells (EMA) and for apocrine metaplasia (GCDFP-15) did not permit a distinction between benign and malignant papillary lesions; however, immunocytochemical staining for CEA using monoclonal antibodies, and for actin (a marker of the myoepithelial cells) was discriminative in this respect. Benign papillomas have a basal layer of actin-rich myoepithelial cells; the cytoplasm of the epithelial cells is CEA negative. Papillary carcinomas lack the myoepithelial layer, except in areas where multiple papillomas are present, associated with ductal or papillary cancer. CEA was detected in 85% of carcinomas. Two of the cases of "suspected carcinoma" lacked myoepithelial cells and were interpreted as carcinomas. It is concluded that the immunocytochemical methods for cell markers can offer valuable data in the study and diagnosis of papillary lesions of the breast; it is difficult, however, to be categorical in borderline cases since in our experience, the behavior of the malignant papillary lesions of the breast is usually favorable. Residual foci of multiple intraductal papillomas were found in seven cases of papillary carcinoma, supporting the pre-neoplastic potential of this condition.

Serum and tissue total sialic acid as a marker for human thyroid cancer.

In this study, serum and tissue sialic acid levels in various histopathological types of thyroid cancer were measured and were found to be significantly higher than those measured in the control group. The amount of sialic acid in tissue was 2.8 +/- 1.42 mg/100 mg protein while the serum level was 79.2 +/- 6.6 mg per dl. The rise in tissue and serum sialic acid levels was relatively non-specific with respect to type of cancer as they depend on the tumor burden only. Neither were they related to age, sex or duration of cancer. The serum sialic acid levels increased during the few days following thyroidectomy and returned to normal thereafter. The sialic acid determination appeared to be an important criterion in patient follow-up and evaluating therapeutic response.

S-100 protein positivity in breast carcinomas: a potential pitfall in diagnostic immunohistochemistry.

S-100 protein, originally isolated from neural tissues, has also been identified in various normal and neoplastic cells, including malignant melanomas. A systematic immunohistochemical investigation of this antigen was performed on formalin-fixed paraffin-embedded samples of benign and malignant breast tissues with use of the avidin-biotin-peroxidase complex immunoperoxidase technique and a polyclonal antiserum that recognizes both the alpha and beta subunits of S-100 protein. In benign breast tissue, S-100 protein was present in both epithelial and myoepithelial cells of terminal ducts and lobules; the staining was cytoplasmic and sometimes nuclear. Of 100 randomly selected invasive breast carcinomas, 48 per cent contained S-100 protein-positive tumor cells. Lobular and medullary carcinomas (60 per cent and 80 per cent, respectively) were more frequently positive than ductal carcinomas (45 per cent). Dendritic cells, most likely Langerhans' cells, were present in some carcinomas and were also positive for S-100. There was no relationship of S-100 positivity to histologic differentiation, recurrence interval, or the expression of various tumor markers. The presence of S-100 protein positivity in metastatic breast carcinomas may lead to the erroneous diagnosis of malignant melanoma. Our observations underscore the importance of testing for a broad panel of tumor markers rather than relying on single antigens in evaluating metastatic malignancies of undetermined origin.

Type II estrogen receptors in the papillary cystic tumor of the pancreas.

Two cases of papillary cystic tumor (PCT) of the pancreas were investigated for the presence of estrogen receptors (ERs) and progesterone receptors (PgRs). Both PCT and normal pancreas are able to specifically bind 3H-estradiol. This binding almost exclusively results from the presence of high levels of type II ER, whereas type I ERs were absent or present at very low levels. Both normal and neoplastic pancreas studied immunohistochemically for the presence of nuclear ER had negative results. This could be explained assuming that anti-ER antibodies are specific for type I binding sites. In conclusion, the presence of specific estrogen as well as progesterone binding may explain the sex and age predilection of PCT and suggest a possible hormone sensitivity for this tumor.

Dendritic/Langerhans cells and prognosis in patients with papillary thyroid carcinomas. Immunocytochemical study of 106 thyroid neoplasms correlated to follow-up data.

Paraffin sections of 106 primary thyroid carcinomas were the subject of an immunocytochemical study to determine the density of infiltrates of S-100 protein-positive dendritic/Langerhans cells (LC), lysozyme-positive histiocytes, and LCA-positive lymphocytes. Evidence of dense infiltrates of LCs was found only in the majority of papillary thyroid carcinomas (PCs). The determination of the quantity of LCs proved to be a highly effective means of assessing the prognosis of these tumors. Irrespective of other morphologic and clinical features, no single instance of death resulting from cancer occurred among 23 PCs with dense LC infiltrates (including 6 tumors of stage pT4), while 9 of 53 (17%) of the remaining patients ultimately died from thyroid cancer. On the other hand, the degree of histiocytic and lymphocytic infiltrations was not associated with a distinct biologic behavior neither among PC nor among the remaining thyroid carcinomas. These findings suggest that LCs may play an important role in the immunologic defense mechanisms of the host against the tumor only in the papillary type of thyroid cancer.

Expression of keratin 19 distinguishes papillary thyroid carcinoma from follicular carcinomas and follicular thyroid adenoma.

Keratin expression with the use of chain-specific monoclonal antikeratin antibodies was investigated in normal thyroid tissue (n = 4), colloid nodules (n = 19), follicular thyroid adenomas (n = 18), follicular carcinomas (n = 10), and papillary carcinomas (n = 12). Frozen sections were stained with monoclonal antibodies M20 (keratin 8), M9 (keratin 18), and LP2K (keratin 19) with the use of the indirect immunoperoxidase technique. The immunohistochemical findings showed that the expression of keratins 8 and 18 was equally extensive in all normal, benign, and malignant lesions tested. In contrast, different staining patterns were observed with the use of monoclonal antibody to keratin 19. Follicular carcinomas were only focally stained with this antibody or were not reactive at all. Keratin 19, however, was present in all the tumor cells of papillary tissues and in a moderate amount of cells of nonneoplastic thyroid lesions and follicular adenomas. In papillary carcinoma, an identical homogeneous expression of keratin 19 was observed in both papillary and follicular structures, which suggests a common cellular origin. These results show that immunohistochemical staining with the use of monoclonal antibody against keratin 19 is useful to distinguish papillary thyroid carcinomas from follicular adenomas and follicular thyroid carcinomas.

Comparison of nuclear DNA content in primary and metastatic differentiated thyroid carcinoma.

Nuclear DNA content of 20 cases of primary differentiated thyroid carcinoma, 27 corresponding cervical lymph node metastases, and 2 local recurrent tumors was determined by flow cytometry. Evidence of DNA aneuploidy was found in either the primary tumor or in the corresponding metastases in 10 (63%) of the 16 papillary carcinomas, in all 3 follicular carcinomas, and in the case of medullary carcinoma studied. In all but one case, the aneuploid stemlines found in the regional metastases were also found in the primary or in the recurrent tumor. In four cases diploid metastatic tissue was found to originate from a tumor with DNA aneuploidy, and in three other cases two stemlines of tumor cells with different DNA indices could be shown in the primary tumor, further indicating clonal heterogeneity in differentiated thyroid carcinomas. None of the 11 patients with either diploid or aneuploid primary tumor with a DNA index less than 1.2 evaluated for 4-12 years died from thyroid cancer, whereas 5 of the 7 patients with primary tumor DNA index greater than 1.2 died from thyroid cancer (P less than 0.01).

Thyrotropin binding and adenylate cyclase stimulation in thyroid neoplasms.

Experiments were performed to determine whether thyroid-stimulating hormone (TSH) receptors coupled to adenylate cyclase are present in both benign and differentiated malignant thyroid tumors, and to identify abnormalities in the TSH receptor-adenylate cyclase system in neoplastic thyroid tissue. TSH binding and adenylate cyclase assays were performed under the same in vitro cyclase conditions. 125I-bovine TSH was incubated with an 8,000 X g thyroid or thyroid tumor particulate fraction from both normal and most neoplastic tissue two receptor sites were identified. The association constant for the high-affinity receptor from normal thyroid was 10.5 +/- 3.2 nM (mean +/- standard error) with a capacity of 0.8 +/- 0.3 pM/mg particulate protein. The association constant of the high-affinity receptor found in the particulate fraction from the thyroid adenomas was 3.5 +/- 0.5 nM with a capacity of 0.4 +/- 0.1 pM/mg particulate protein; for the thyroid carcinomas the association constant was 1.4 +/- 0.3 nM and the capacity 0.2 +/- 0.1 pM/mg particulate protein. The maximum adenylate cyclase response to TSH for particulate fractions from both benign and malignant tumors was greater (P less than 0.05) than in the particulate fraction from normal adjacent thyroid tissue. There was also a positive correlation between the equilibrium constants for 125I-bTSH binding and adenylate cyclase activation suggesting that binding sites are coupled to cyclase in the neoplastic tissue and that the binding observed in these experiments is of biologic consequence.

DNA content in radiation-associated thyroid cancer.

DNA content has been reported to be of prognostic significance in differentiated thyroid carcinoma. Since malignant tumors with irradiation as an initiator often contain DNA aberrations, the DNA content of well-differentiated thyroid carcinoma in patients with a prior history of low-dose head and neck irradiation was determined and compared with similar nonradiation-associated lesions. The DNA content of thyroid cancers from 53 patients was determined with use of flow cytometry. Sixteen radiation-associated thyroid carcinomas (11 papillary, 3 follicular, and 2 medullary) all were diploid. In a group of 37 nonradiation-associated tumors, 10 were aneuploid (10 of 29 papillary carcinomas and 0 of 2 follicular or 6 medullary carcinomas). This difference in DNA content is significant (p less than 0.02, Fisher's exact test). These findings were unexpected and suggest that if the initiating irradiation causes a DNA aberration, this aberration is not reflected in DNA content as measured by means of flow cytometry.

Co-existent anaplastic and well differentiated thyroid carcinomas: a nuclear DNA study.

Clonal transformation of well differentiated follicular or papillary carcinomas has been suggested as a mechanism by which anaplastic carcinomas of the thyroid might arise. Of 126 cases of anaplastic (giant cell) carcinomas, 17 (13.5%) contained histologically well differentiated tumour foci within or adjacent to the high grade malignant anaplastic tumour. Cytophotometric DNA analysis after Feulgen staining was performed on 11 cases in order to evaluate ploidy of the anaplastic and the well differentiated tumour cells. The majority of these co-existent carcinomas (9/11) were papillary. All 11 anaplastic carcinomas demonstrated an aneuploid DNA pattern which correlated with a poor clinical outcome (7 of 11 died of disease in less than 6 months). In contrast six co-existent papillary and one co-existent follicular tumours were diploid. These data show that the co-existence of anaplastic and well differentiated carcinoma occurs only rarely and when it occurs only one third of the well differentiated tumours contain aneuploid tumour cells. This suggests that in the majority of cases of anaplastic thyroid carcinoma the malignant cells arise de novo rather than through clonal transformation of well differentiated carcinomas.

Correlation of an estrogen receptor-related phosphoprotein with histopathological features in breast cancer.

A series of 65 cases of different histological types of breast carcinoma was investigated for the immunohistochemical location of the estrogen receptor-related, 29 kD phosphoprotein using the ER-D5 monoclonal antibody. The ER-D5 response is heterogeneous in relation to some therapeutic limitations and is correlated with histopathological features of the tumors and survival. The main parameters for evaluation of breast cancers are reviewed, both those that are statistically correlated and those that are not apparently always correlated but are known to have considerable biological meaning, such as the ER-status of tumors.

Comparison of computerized analysis of nuclear DNA changes in uterine cervix dysplasia and in urothelial non-invasive papillary carcinoma.

The nuclear DNA content was measured in preneoplastic lesions of the uterine cervix and in papillary carcinomas of the bladder. Three groups of features were calculated from the raw data: basic DNA, DNA deviation and DNA distribution. The basic DNA features, concerning both the cervix and the bladder, showed a progressive increase in the mean DNA content, a decrease in the percentage of diploid nuclei and steadily increasing values of polyploid and aneuploid nuclei. Among the DNA deviation features, the malignancy grade value was zero in the normal cervical epithelium and in the normal urothelium. An increase in this value was evident in moderate dysplasia and in urothelial papillary carcinoma of grade 2, the highest value being in CIS and in grade 3. Concerning the DNA distribution features, the values of the 15th and 95th percentiles and their difference were progressively higher both in the cervix and in the bladder, expressing a continuous shift and spread of the DNA content measurements in the different diagnostic categories, with respect to normal epithelium and urothelium. The statistical analysis showed that the strongest correlation is between 2c D.I. and % of polyploid nuclei in the cervix and between M.I. and % of aneuploid nuclei greater than 4c in the bladder. In the cervix the most discriminating feature is the Malignancy Grade, whereas in the bladder it is the percentage of diploid nuclei. The comparison between the results of the three groups of features showed that: 1) Mild dysplasia of the cervix and urothelial papillary carcinoma of grade 1 showed similar changes in DNA features. Both were basically characterized by increased proliferative activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Keratin, luminal epithelial antigen and carcinoembryonic antigen in human urinary bladder carcinomas. An immunohistochemical study.

14 urinary bladder carcinomas of all main types were investigated with antisera to "broad spectrum keratin" (aK), "luminal epithelial antigen" (aLEA) and carcinoembryonic antigen (aCEA), using an indirect immunoperoxidase method on formalin fixed paraffin embedded sections. Keratin and LEA were both present in normal transitional epithelium, papilloma and carcinoma in situ whereas CEA was absent. Transitional cell carcinomas reacted with both aK and aLEA whereas CEA was seen only in a few foci. In squamous metaplasia and squamous carcinoma reaction with aK was particularly strong, while LEA was almost lacking and CEA was present in necrotic centres. In adenocarcinomas aK and aLEA reacted equally while aCEA reacted only on the surface.

Gastric cancer with special references to WHO and Laurén's classifications: glycogen and triacylglycerol concentrations in the tumor.

In eighty patients with histologically verified gastric carcinoma the concentrations of glycogen and triacylglycerols were evaluated in specimens taken endoscopically from the tumor and the surrounding unchanged gastric mucosa. The results were analyzed in relation to the histological type of carcinoma according to WHO's and Laurén's classifications. The control group consisted of sixteen patients with superficial chronic gastritis. An elevated glycogen concentration was found in tumors of all types of gastric carcinoma; its level in the neoplasm was significantly higher also in relation to unchanged gastric mucosa surrounding the tumor. A particularly high glycogen level was present in the slow growing well-differentiated cancers, e.g. papillary and tubular adenocarcinomas or intestinal-type carcinoma. Reversely, in the fast growing and poorly differentiated cancers, e.g. undifferentiated or diffuse-type carcinoma, the glycogen contents were lower. Also triacylglycerol concentrations in the tumors as well as in the surrounding unchanged gastric mucosa were significantly higher than those in the control gastric mucosa specimens; no significant difference in triacylglycerol concentrations was observed between groups of patients with various types of carcinoma. It was concluded that (1) glycogen concentrations in the neoplastic tissue are cancer-growth related and characteristic for each kind of carcinoma, (2) an elevated triacylglycerol content in the tumor is probably a result of general lipid changes in the host.

Intracystic papillary carcinoma of the male breast. Immunohistochemical and ultrastructural study.

We report a case of mammary intracystic papillary carcinoma occurring in a 75-year-old man. The tumor was present on the left pectoral area for five years. Grossly, the neoplasm was a cystic structure 10 cm in diameter, with multiple intramural filiform papillae and small foci of cyst wall invasion. By transmission electron microscopy the tumor cells had the normal complement of organelles and also multiple electron-dense, membrane-bound secretory granules. These granules were also demonstrated with multiple stains for argyrophilia and with periodic acid-Schiff. Immunoperoxidase stains were negative for neuron-specific enolase, S100 protein, vasoactive intestinal peptide, corticotropin, calcitonin, lactalbumin, and bombesin, and positive for human heart factor (myoepithelial cells) and carcinoembryonic antigen. We believe that this rare neoplasm represents a variant of mammary adenocarcinoma and not a neuroendocrine (carcinoid) neoplasm.

DNA measurements on cell nuclei of normal, proliferating and neoplastic thyroid tissues in rats.

Nuclear DNA content was measured in 3 normal, 9 hyperplastic and 16 neoplastic rat thyroid glands. Thyroid hyperplasia and tumor growth were induced after treatment of the animals with X-rays and methylthiouracil. In the control animals only diploid thyroid epithelial cells were observed. At the stages of diffuse and nodular thyroid hyperplasia, the total DNA content per nucleus indicated for a diploid chromosome number and only a few cells were hyperdiploid. In the thyroid adenomas and carcinomas a scattering of the diploid region and an increase in the number of hyperdiploid cells was found. Among the various types of thyroid tumors neither difference in the number of hyperdiploid cells, nor typical pattern of distribution of these cells in the histogram was found. The increased number of hyperdiploid cells in the hyperplastic and neoplastic thyroids suggest an increase in the proportion of the cells entering the cell cycle and does not indicate for appearance of a neoplastic stemline.