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Measurement of DNA ploidy can be performed either with Flow Cytometry (FCM) and Image-Analysis (IA); both methods provide prognostic information in primary breast cancer. We compared the results of quantitative DNA analysis of formalin fixed, paraffin embedded tissue from 62 invasive ductal breast cancers. For FCM nuclear suspensions from disaggregated tumor were stained with Propidium Iodide and analyzed by means of Ortho Cytoron Absolute. For IA nuclear suspensions were stained by the Feulgen method and analyzed by means of Vidas system. We found a good correlation between flow cytometry DNA Index and Histogram Type, according to Auer classification (rs = 0.65, p < 0.001) and between DNA Index and Grading of Malignancy (MG) which had been measured by Bocking's algorithm (rs = 0.38, p < 0.05). Concerning to disease free survival (DFS), flow cytometric DNA Index showed a better correlation (rs = 0.56, p < 0.001). We concluded that the two methods provide comparable results, but offer individual advantages and are complementary for analyzing DNA ploidy in breast cancer.

We evaluated breast cancer specimens from 150 patients after a follow-up more than 10 years, by means of DNA flow cytometry, immunohistochemistry (cerbB2 and Cathepsin D) and argyrophilic (AgNORs) methods. In univariate analysis all indexes studied provide prognostic information about the disease free survival, whereas in the multivariate Cox model the DNA Index is the most important prognostic factor. We conclude that the data obtained from DNA flow cytometry, associated with other parameters, can be of greatest importance for the decision an the level of aggressiveness of adjuvant therapy for the decision an the level of aggressiveness of adjuvant therapy for a individual patients.

Silver-binding nucleolar organizer regions (AgNORs) have been counted in sections of routinely processed paraffin embedded material. The AgNOR score has been correlated with proliferative activity of various neoplasms. An increase in the AgNOR score can be related to cellular growth fractions or DNA-ploidy. We have examined 301 invasive ductal breast carcinomas by this method. The AgNOR counts were also compared with DNA Ploidy and the S-phase fraction flow cytometry. A positive correlation has been found between the AgNOR score and the S-Phase fraction. These data show cell proliferation. We conclude that AgNOR counting may provide information on breast carcinoma in addition to that obtained from flow cytometric analysis.

By means of an AgNOR technique, the authors have carried out a retrospective study on proliferative activity in 170 cases of infiltrating ductal breast carcinoma (so-called carcinoma NOS). The follow-up period was 138 months. A statistically significant correlation was found between AgNOR score and outcome of the disease ("vital status"), clinical stage and OS respectively. By means of discriminant analysis a threshold value of AgNOR score was identified (= 9.5) which distinguished two risk groups among the cases under examination. Those cases with an average AgNOR score below the threshold value showed a favourable prognosis. The cases with an AgNOR score greater than 9.5 had an unfavourable outcome. The authors suggest that AgNOR score may be used as a prognostic parameter in breast carcinoma.

Background and Design: The nucleolar organizer regions (NORs) are chromosomal loops of DNA to which acidic proteins are associated that are seen by silver staining as black dots within the nucleoli (hereafter, these silver-staining NORs will be referred to as AgNORs). As their size and number reflect cell and nuclear activity, their counting in paraffin sections is regarded as a useful tool for diagnosing and prognosing malignant tumors. We counted AgNORs in 98 patients with stage I melanoma, followed up to an average of 73 months, to verify whether the number of AgNORs is of prognostic value., Results: The number of AgNORs averaged 2.792 +/- 0.901 in the 64 patients without metastases and 4.889 +/- 1.403 in the 34 with metastases. In patients with counts higher than 3.62, there was an 82% probability of metastases developing., Conclusion: The technique is fast, simple, and reproducible with easily available reagents and standard light microscopy. In our population, AgNOR counts constituted a more accurate prognostic indicator than Clark's level and Breslow's thickness. Confirmation in a new population is needed.

By means of a silver staining method, Nucleolar Organizer Region-associated proteins (NORs) have been evaluated on paraffin sections of a series of 58 ultrasound-guided liver biopsy specimens. These included 12 normal livers, 12 cirrhotic livers, 12 cases of chronic hepatitis and 22 cases of hepatocellular carcinoma. A significant difference (P less than 0.001) was found between the mean AgNOR scores of the normal and pathological biopsies, and between the non-neoplastic and the carcinomatous lesions. The authors suggest that AgNOR counts, in combination with conventional histocytological criteria, may be a useful method in the diagnosis of liver diseases.

53 cases of invasive ductal (NOS) carcinomas of the breast were studied by means of an immunostaining method with Ki-67 monoclonal antibody and an argyrophilic method for the demonstration of Nucleolar Organizer Regions (AgNORs). The percentage of cancer cells with nuclear Ki-67 immunoreactivity and the mean number of NORs for each tumour were statistically related. The data obtained showed a good correlation between Ki-67 index and NOR score (rS = 0.47 - P less than 0.001). The authors suggest that the AgNOR method--which is applicable to routinely processed material--might effectively substitute Ki-67 immunostaining as a marker of cell proliferation in ductal breast cancer.

The ultrastructural localization of glucagon in the presence of Scyliorhinus canicula was investigated. We used a post-embedding immunoelectron microscopy method on pancreatic samples fixed in glutaraldehyde and osmicated before embedding. Contrasting with uranyl acetate and lead citrate was also performed after immunolabelling, but best results were obtained with uranyl acetate only. Glucagon-like immunoreactivity was located in round granules (300-600 nm) surrounded by a limiting membrane. The matrix varied in electron density and exhibited a dense core surrounded by a less dense mantle. The granules were seen in two different cell types, which differed in the electron density of their cytoplasm. Glucagon-immunoreactive cells were the largest pancreatic cells types and were often localized near somatostatin-containing cells.

Studies on the quantitative expression of the Major Histocompatibility Complex (MCH) in hepatocytes chronically infected by Hepatitis B Virus (HBV) report that an increased expression of these antigens could be related to a good immunological response. In the present work we analyze the expression of the MCH antigens in cryostatic sections of liver biopsies taken from subjects (19 children) with various forms of HBsAg positive chronic hepatitis. A high expression of HLA class I antigens and a high degree of hepatocyte necrosis was evident in Chronic Active Hepatitis (CAH) and Chronic Lobular Hepatitis (CLH). On the contrary, subjects with histological diagnosis of Chronic Persistent Hepatitis (CPH) showed a low expression of such antigens. There was however, the difference that in subjects with high hepatic cytolysis and high expression of HLA class I antigens, serum HBV-DNA was clearly present in almost all the cases with CAH, but not detectable in all cases with CLH. The expression of HLA class II antigens and of Beta2 microglobulin was the same in all 19 cases. All cases with HBV-DNA positivity with high class I antigen expression had active hepatitis which seems to suggest that all attempts at viral clearance on the part of the immune system have been in vain. We hope our paper will be an additional parameter for evaluating the course of hepatitis during Interferon treatment.

Nucleolar Organizer Region-associated proteins (NORs) have been evaluated on paraffin sections of a series of cutaneous malignant melanomas, stage IA at the time of the initial diagnosis. A significant difference (P less than 0.001) was found between the numbers of AgNORs in the cases which developed metastases and those without metastases: increasing values of AgNORs representing increased risk of metastases.

Carrier mouse erythrocytes, i.e., red cells, subjected to a dialysis technique involving transient hypotonic hemolysis and isotonic resealing were treated in vitro in three different ways: (a) energy depletion by exposure for 90 min at 42 degrees C; (b) desialylation by incubation with neuroaminidase; and (c) oxidative stress by incubation with H2O2 and NaN3. Procedure (c) afforded maximal damage, as shown by analysis of biochemical properties of the treated erythrocytes. Reinfusion in mice of the variously manipulated erythrocytes following their 51Cr labeling showed extensive fragilization as indicated by rapid clearance of radioactivity from the circulation. Moreover, both the energy-depleted and the neuraminidase-treated erythrocytes showed a preferential liver uptake, reaching 50 and 75%, respectively, within 2 h. On the other hand, exposure of erythrocytes to the oxidant stress triggered a largely splenic removal, accounting for almost 40% of the reinjected cells within 4 h. Transmission electron microscopy of liver from mice receiving energy-depleted erythrocytes demonstrated remarkable erythrocyte congestion within the sinusoids, followed by hyperactivity of Kupffer cells and by subsequent thickening of the perisinusoidal Disse space. Concomitantly, levels of serum transaminase activities were moderately increased. Each of the three procedures of manipulation of carrier erythrocytes may prove applicable under conditions where selective targeting of erythrocyte-encapsulated chemicals and drugs to either the liver or the spleen has to be achieved.

A case of aggressive angiomyxoma (AAM) of the pelvis and perineum is reported, whose clinical and pathological findings are consistent with the literature. This is a clinicopathological entity recently described by Rosai and colleagues (1983) who stressed its locally infiltrative nature and its tendency to recur. Its differential diagnosis includes a series of benign and malignant soft tissue tumours.

By means of a silver staining method, Nucleolar Organizer Region-associated proteins (NORs) have been evaluated on paraffin sections of two series of benign and malignant melanocytic skin lesions (represented by 30 intradermal naevi and 30 malignant melanomas). A significant difference (P less than 0.001) was found between the numbers of Ag-NORs in the nuclei of benign lesions and those of malignant lesions. It is suggested that this method could be of value in assessing the biological potential of cutaneous melanocytic neoplasms.