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D Trerè

Publications and source records attributed to D Trerè.

At least 55 records · Page 3Linked to original sources
PubMed1995

The quantity of nucleolar proteins nucleolin and protein B23 is related to cell doubling time in human cancer cells.

BACKGROUND: The quantity of the silver-stained nucleolar proteins (AgNOR proteins) measured in situ in cytohistologic preparations is related to the rapidity of cell proliferation. The term "AgNOR proteins" comprises several proteins. The relationship between the individual AgNOR protein amount and cell proliferating activity is not yet known. We studied the quantitative distribution of the individual AgNOR proteins, with specific attention to the two major AgNOR proteins, nucleolin and protein B23, in seven human cancer cell lines characterized by different cell doubling times. DESIGN: The doubling time of cancer cells was measured by counting the asynchronously growing cells at regular time intervals. The AgNOR proteins were quantified in situ, after a specific one-step staining procedure, by computerized image analysis. For the quantitative evaluation of nucleolin and protein B23, two methods were followed. Nuclear proteins after separation by SDS-PAGE were transferred onto nitrocellulose membranes and were either: 1) stained by the silver staining procedure for AgNOR proteins or 2) treated with anti-nucleolin and anti-protein B23 mAb followed by reaction with secondary Ab linked to peroxidase and revealed by chemiluminescence and autoradiography. In both cases, measurement of individual AgNOR protein and nucleolin and protein B23 amount was carried out using computerized densitometric analysis. RESULTS: Integrated density values of the silver-stained bands at 105 kDa (nucleolin) and 38 to 39 kDa (protein B23) represented, in all cell lines, more than 60% of the total silver-stained band value. A relationship was found between the densitometric values of silver-stained nucleolin and protein B23 and rapidity of cell proliferation (r = 0.85 and r = 0.86, respectively, p < 0.05). The values of nucleolin and protein B23 obtained using the Western blots were strictly related to the rapidity of cell proliferation (r = 0.93 and 0.96, respectively, p < 0.001). Finally, a good correlation was observed between the mean AgNOR protein area value, as defined in cytologic preparations in situ, and nucleolin and protein B23 amounts as evaluated in silver-stained nitrocellulose membranes (r = 0.92 and r = 0.90, respectively, p < 0.01) and in Western blots (r = 0.95 and r = 0.94, respectively, p < 0.001). CONCLUSIONS: These data indicate that the quantitative changes of AgNOR proteins observed in cytohistologic preparations in situ mainly reflect the quantitative changes of nucleolin and protein B23 and demonstrate that nucleolin and protein B23 amounts are inversely related to cell doubling time in human cancer cells.

Adenocarcinoma↗
PubMed1995

Higher reproducibility of morphometric analysis over the counting method for interphase AgNOR quantification.

In a series of 40 breast carcinomas, the reproducibility of two different methods for interphase AgNOR quantification was evaluated. Two operators independently defined on each slide the interphase AgNOR quantity both by measuring the area of the silver-stained structures using image cytometry and counting the AgNOR number directly at the microscope. The correlation between the values obtained by the two observers was statistically significant, but the correlation coefficient between AgNOR areas (r = 0.79; P < 0.001) was greater than that between AgNOR numbers (r = 0.38; P = 0.014). On the other hand, when interphase AgNOR area and number values obtained by each observer were compared, no significant correlation was found. This study has demonstrated that the two different methods for interphase AgNOR quantification are not comparable, and that morphometric analysis is more objective and reproducible than the counting method.

Humans↗
PubMed1994

Interphase AgNOR quantity is not related to DNA content in 11 established human cancer cell lines.

The relationship between interphase AgNOR quantity and DNA ploidy was evaluated in 11 cell lines derived from human tumors of different origin. Interphase AgNOR quantity was measured by morphometric image analysis and DNA content by cytofluorometry. The mean interphase AgNOR area ranged from 5.56 to 21.12 microns 2 and DNA ploidy from 2.05 to 3.30c. The interphase AgNOR value and DNA content were not significantly related (r = 0.23, P = 0.52). No significant correlation was found between DNA content and cell doubling time, whereas a strict relationship was confirmed to exist between interphase AgNOR quantity and rapidity of cell proliferation.

Cell Division↗
PubMed1994

Interphase AgNOR quantity and DNA content in endometrial adenocarcinoma.

The relationship between the amount of silver-stained nucleolar organizer regions (AgNORs) and the DNA content of interphase neoplastic cells in 63 cases of human endometrial adenocarcinoma was studied. Interphase AgNOR content was determined as the mean AgNOR area per nucleus by image cytometry in silver-stained samples from paraffin-embedded sections. DNA content was evaluated by cytofluorometry on single cells from the same specimen and expressed in c units as DNA ploidy. The mean AgNOR area ranged from 1.26 to 4.48 microns2 and the modal DNA ploidy from 1.90 to 4.00c. No significant associations were found between mean AgNOR area and modal DNA ploidy when analyzed either by linear regression (beta = 0.18; P = 0.15) or as dichotomized variables by chi 2 analysis (chi 2 = 0.82; P = 0.37). On a homogeneous subset of 45 cases of stage I adenocarcinomas we also evaluated the prognostic relevance of AgNOR content and DNA ploidy. In monovariate analysis both parameters were significantly related to patient survival. When the mean AgNOR area and the modal DNA ploidy were entered into regression analysis together with the depth of myometrial invasion and histological grading, they showed an independent, high prognostic value.

Adenocarcinoma↗
PubMed1994

Prognostic relevance of pretreatment proliferative rapidity of marrow blast cells in childhood acute lymphoblastic leukaemia.

Cell proliferation rate is a well-established prognostic factor in cancer, but it has not been considered to identify the risk group of childhood acute lymphoblastic leukaemia (ALL) at presentation. We carried out a study to demonstrate the prognostic importance of the rapidity of cell proliferation in patients with ALL. To measure the rapidity of cell proliferation we used the parameter relative to the area of silver-stained nucleolar organiser regions (AgNORs) as evaluated by morphometric analysis on smeared marrow blast cells. The mean AgNOR area of leukaemic marrow cells was measured in 119 children. By using a cut-off value of 3 microns2, we identified a group of 91 children with low proliferating blast activity (mean AgNOR value 2.11 microns2) and a group of 28 children with high proliferating activity (mean AgNOR value 3.29 microns2). The group of patients with a mean AgNOR value > 3 microns2 was characterised by a higher number of deaths, more frequent relapse and shorter time interval to relapse than the group of patients with mean AgNOR value < 3 microns2 (P < 0.01). Multivariate analysis performed to include T-cell immunophenotype, FAB morphology, leucocyte count and presence of mediastinal mass showed that the mean AgNOR value was the only independent predictor of unfavourable event-free survival probability (P > 0.01). Our results indicate that the rapidity of marrow blast cell proliferation is an important prognostic parameter in childhood ALL and should be routinely introduced in the group risk definition.

Bone Marrow↗
PubMed1994

Technical and methodological aspects of silver staining and measurement of nucleolar organizer region (NOR).

In the last few years the NOR silver-staining technique has been introduced in tumor pathology for both diagnostic and prognostic purposes, but the lack of a standardized staining protocol has frequently led to misinterpretation of actual structures evaluated by the various authors. Recent data have demonstrated that AgNOR area is closely related to the whole stained nucleolar area obtained by prolonging the silver staining reaction beyond the optimal time for selective NOR visualization. In order to compare data from different laboratories, we propose to stain the whole nucleolus by silver and to measure the dimensions of the silver-stained nucleoli by image analysis.

Breast Neoplasms↗
PubMed1994

AgNOR proteins as a parameter of the rapidity of cell proliferation.

A series of studies on the relationship between AgNOR quantity and cell kinetics have been reviewed. The results indicate that the quantity of interphase AgNORs, evaluated by morphometric analysis, increases in cycling cells from early G1-phase to the late S-phase. In cancer tissues the AgNOR value is also closely related to both the percentage of cycling cells (measured by Ki67 or PCNA immunolabelling) and S-phase cells (measured by BrdU incorporation or DNA flow cytometry). Data from cell lines cultured in vitro have clearly shown that interphase AgNOR quantity is related to cell doubling time: the faster the rapidity of cell proliferation, the greater the interphase AgNOR quantity.

Cell Cycle↗
PubMed1993

Is high AgNOR quantity in hepatocytes associated with increased risk of hepatocellular carcinoma in chronic liver disease?

AIMS: To evaluate whether high numbers of silver staining nucleolar organiser regions (AgNORs) in hepatocytes are associated with increased risk of hepatocellular carcinoma in chronic liver disease. METHODS: The quantitative distribution of AgNORs was studied in the liver biopsy specimens of 33 patients with chronic liver disease, 11 of whom developed hepatocellular carcinoma. The interval between liver biopsy and diagnosis of hepatocellular carcinoma was 26 months (range one to 61 months); the mean follow up of patients without hepatocellular carcinoma was 45 months (range 24-59 months). Quantitative evaluation of AgNORs was carried out on silver stained routine sections by morphometric analysis, using a computer assisted image analysis system. RESULTS: High interphase AgNOR values (> 3 microns2) were found in hepatocytes of nine out of the 11 (82%) patients in whom neoplastic transformation occurred. Of the remaining 22 patients, only seven (31%) had AgNOR values higher than > 3 microns2 (chi 2 4.83; p = 0.036). CONCLUSIONS: These results indicate that high numbers of interphase AgNORs are associated with increased risk of hepatocellular carcinoma in patients with chronic liver disease.

Adult↗
PubMed1993

Localization of DNA in the fibrillar components of the nucleolus: a cytochemical and morphometric study.

We studied the distribution of DNA in human circulating lymphocyte nucleoli using three different cytochemical methods for selective visualization of DNA in thin sections: the Feulgen-like osmium-ammine reaction, the NAMA-Ur procedure, and the osmium-ammine staining in glycine buffer, pH 1.5. All three methods indicated the presence of uniformly distributed, highly decondensed DNA filaments forming a large solitary agglomerate in the central part of the nucleolar area, corresponding to the solitary large fibrillar center (FC) as revealed by uranium and lead staining. We also studied the relationship between DNA agglomerates and nucleolar fibrillar components in resting and phytohemagglutinin (PHA)-stimulated lymphocytes by morphometric analysis of the areas occupied by these structures. In resting lymphocytes the mean area of the DNA agglomerates was 0.479 micron 2 +/- 0.161 SD, whereas that of FCs was 0.380 micron 2 +/- 0.149 SD, with a ratio of 1.26. In PHA-stimulated lymphocytes the mean area of the DNA agglomerates was 0.116 micron 2 +/- 0.056 SD, whereas that of the FCs was 0.075 micron 2 +/- 0.032 SD, with a ratio of 1.55. In PHA-stimulated lymphocytes we also measured the area occupied by the FCs plus the closely associated dense fibrillar component (DFC). The mean value of these two fibrillar components was 0.206 micron 2 +/- 0.081 SD. These data demonstrate that decondensed DNA filaments are uniformly distributed in the FCs and that in transcriptionally active nucleoli they are also present in the proximal portion of the DFC surrounding the FCs.

Cell Nucleolus↗
PubMed1993

Expression of alpha 2,6-sialylated sugar chains in normal and neoplastic colon tissues. Detection by digoxigenin-conjugated Sambucus nigra agglutinin.

It has previously been reported that about 90% of human colon carcinomas express increased levels of the sialyltransferase which adds sialic acid in alpha 2,6 linkage to galactose residues on N-linked chains of glycoproteins. To ascertain whether colon cancer tissues actually express increased amounts of alpha 2,6-sialylated sugar chains on their glycoconjugates, we screened tissue sections of normal colon, benign and malignant colon tumors with digoxigenin-conjugated Sambucus nigra agglutinin (SNA), a NeuAc alpha 2,6Gal/GalNAc-specific lectin. At the concentration of lectin used, epithelial cells of all the 13 normal colon specimens examined were unreactive; 3 out of 8 benign lesions showed a weak reactivity being the remainder unreactive, while 23 out of 26 carcinomas were positive at a variable degree. Qualitative differences were evident among different carcinoma specimens. In some cases a large number of intensely stained intracytoplasmatic particles was present, thus suggesting that reactivity may be associated with secretions, very likely of mucus droplets. In other specimens there was a more uniform distribution of the staining which suggest that reactivity is associated with cell membrane glycoconjugates. These data indicate that the expression of a2,6-sialylated sugar chains is remarkably increased in the majority of colon cancer specimens examined.

Adenoma↗
PubMed1993

Critical analysis of the methods commonly employed in the assessment of cell proliferation: advantages of the NOR silver-staining technique in routine cyto-histopathology.

The methods commonly employed for the assessment of cell proliferation have been critically considered. Some 'historical' techniques, such as the mitotic count and the thymidine labeling index, cannot be proposed for routine pathology because of their complexity and difficult quantification. Other techniques, such as in vitro bromodeoxyuridine incorporation or in vivo bromodeoxyuridine infusion coupled with DNA flow cytometry, permit a very sophisticated analysis of the tumour cell kinetics, but can be performed only in highly specialized centers. During the past few years the immunohistochemical detection of antigens related to the cell cycle--particularly Ki67 or proliferating cell nuclear antigen (PCNA)/cyclin--has been rapidly introduced in numerous laboratories, and, at present, certainly represents the most common method for the assessment of cell proliferation in tumour samples. However, a new marker of cell proliferation has been recently described which presents technical features particularly suitable for routine use: the quantitative evaluation of interphase silver-stained nucleolar organizer regions (AgNORs). The NOR silver-staining technique is simple, very rapidly executed, inexpensive, easily quantifiable and, most important, can be performed on routinely processed cytological and histological samples.

Animals↗
PubMed1992

Relationship between interphase AgNOR distribution and nucleolar size in cancer cells.

We have studied the relationship between interphase nucleolar organizer region (NOR) distribution and nucleolar size in cancer cells at light-microscopical level. Thirteen cases of formalin-fixed bladder cancer and fifteen cases of methacarn-fixed tumours of different origin were used. Nucleoli of the former cases were stained by Phloxine B and of the latter by Toluidine Blue. Selective visualization of interphase NORs was obtained by carrying out the one-step silver staining reaction for AgNOR proteins (Ploton et al., 1986). The area occupied by Phloxine B- or Toluidine Blue-stained nucleoli and interphase silver-stained NORs was measured by means of an automated image analyser. Both in bladder cancers and in the other tumour lesions nucleolar and interphase AgNOR areas were linearly related (r = 0.95 and r = 0.96, respectively, P < 0.001). The close relationship between the area of nucleoli and that of silver-stained nucleolar structures was maintained even if the silver-staining procedure was prolonged beyond the optimal time length for selective interphase NOR staining. In the latter case, however, single interphase AgNORs were no longer visible within the nucleolar body which was, in fact, homogeneously stained. These data indicate that evaluation of the interphase AgNOR area has the same relevance, in tumour pathology, as whole nucleolar size measurement.

Cell Nucleolus↗
PubMed1992

Ag-NOR protein distribution correlates with patient survival in stage I endometrial adenocarcinoma.

The relationship between argyrophil nucleolar organizer region (Ag-NOR) protein quantity and prognosis was studied in 33 cases of stage I endometrial adenocarcinoma. Ag-NOR protein quantity was measured by image analysis in silver-stained sections from paraffin-embedded samples of curettings. Patients had a minimum 10-year follow-up. Only 2 out of 25 patients exhibiting a mean Ag-NOR protein area of less than 3 microns2 died of cancer, whereas 5 of the 8 patients with a mean Ag-NOR protein area of more than 3 microns2 died of the disease. The present results demonstrate that the Ag-NOR protein value is closely related to patient survival in stage I endometrial carcinoma and that it is a reliable prognostic indicator in this type of carcinoma.

Adenocarcinoma↗
PubMed1991

Establishment of a human medulloblastoma cell line (BO-101) demonstrating skeletal muscle differentiation.

A permanent cell line, BO-101, was derived from a classic vermian medulloblastoma in a 9-year-old child. This line grew in vitro in adherent cultures and grew in athymic mice as serially transplantable intracranial and subcutaneous xenografts. Intracranial neoplasms grew as masses of small cells, which focally showed large cells with intense immunoreactivity for desmin, myoglobin and alpha-striated actin. The rhabdomyoblastic nature of these cells was confirmed ultrastructurally. The primary neoplasm showed immunoreactivity for synaptophysin, neuron-specific enolase and vimentin. A large panel of monoclonal antibodies and antisera against neuronal and glial antigens failed to show glial and neuronal immunoreactivity in the cell culture and xenografts. Despite the marked genotypic and phenotypic differences, the original neoplasm and the cell line share a common chromosomal marker del (12) (p 13.1). The BO-101 line differs phenotypically and genotypically from previously established medulloblastoma cell lines and further supports the heterogeneous biologic properties of the cell populations that constitute these neoplasms.

Animals↗
PubMed1991

AgNOR area in interphase nuclei of human tumours correlates with the proliferative activity evaluated by bromodeoxyuridine labelling and Ki-67 immunostaining.

The area of silver-stained proteins associated with interphase nucleolar organizer regions (AgNORs) was compared with labelling data obtained by bromodeoxyuridine (BrdU) incorporation and Ki-67 immunostaining in 25 tumours of different origins and two non-neoplastic lesions of the thyroid. Our data demonstrate a highly significant correlation between the mean area occupied by the AgNOR proteins measured by an image processing system and the proliferative indices evaluated by BrdU labelling (r = 0.89, P less than 0.001) and Ki-67 immunostaining (r = 0.86, P less than 0.001). AgNOR protein area measurement is therefore proposed as a simple, inexpensive, and reliable method of evaluating the proliferative activity in routinely processed tumour samples.

Breast Neoplasms↗
PubMed1991

Standardization of interphase Ag-NOR measurement by means of an automated image analysis system using lymphocytes as an internal control.

Using an automated image analysis system, we have developed a procedure for standardizing the measurement of silver-stained proteins of the nucleolar organizer regions (NORs) in cancer cells, irrespective of the fixative employed and the time of the staining reaction. We observed that the area of Ag-NOR proteins from the same tumour was smaller in samples fixed with formalin-containing solutions, buffered formalin, and Bouin liquid, compared with those fixed with absolute ethanol and 'methcarn' solution (1.77 +/- 0.26micron 2 and 2.36 +/- 0.35 micron2 versus 3.34 +/- 0.54 micron2 and 3.72 +/- 0.61 micron2). Increased values of the Ag-NOR area were also observed after lengthening the silver staining reaction. However, in both cases no difference was observed in the ratio between Ag-NOR area of cancer cells and that of the lymphocytes infiltrating the stroma. The value of the ratio, which is called the 'Ag-NOR index', was in fact very similar, for the same cancer, after employing different fixatives or staining times. The use of lymphocyte Ag-NOR area as an internal control for the standardization of Ag-NOR evaluation in cancer tissues was made possible by the fact that lymphocyte Ag-NOR area is almost constant in human tumours independent of sex and age.

Adult↗
PubMed1991

Importance of interphase nucleolar organizer regions in tumor pathology.

The importance of the distribution of silver-stained nucleolar organizer regions (Ag-NORs) in interphase nuclei for diagnostic and prognostic purposes in tumor pathology has been reviewed. The available data demonstrated that interphase Ag-NOR evaluation may be of help in distinguishing malignant from hyperplastic or normal cells. On the other hand, there is increasing evidence that a relationship exists between the quantity of interphase Ag-NORs and the prognosis of malignant tumors: the greater the number of interphase Ag-NORs, the worse is the prognosis. This can be explained by the observation that the interphase Ag-NOR quantity is strictly related to the cell proliferation rate. The procedures used for the measurement of the interphase Ag-NOR quantity are also critically discussed.

Cell Division↗