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W Gorczyca

Publications and source records attributed to W Gorczyca.

At least 55 records · Page 3Linked to original sources
PubMed1993

Presence of DNA strand breaks and increased sensitivity of DNA in situ to denaturation in abnormal human sperm cells: analogy to apoptosis of somatic cells.

The presence of sperm cells characterized by increased sensitivity of DNA in situ to denaturation and by abnormal morphology (shape) in semen is associated with low fertility. Such cells often appear in increasing numbers following exposure to toxic or mutagenic agents. In the present study, the sensitivity of DNA in situ to acid-induced denaturation was assayed in sperm cells of 25 patients attending a human fertility center. The same samples were also subjected to analysis of DNA strand breaks. A new method for detecting DNA strand breaks in situ by labeling 3'-OH termini with biotinylated dUTP in a reaction employing exogenous terminal deoxynucleotidyl transferase has been applied. Both methods utilized flow cytometry. A good correlation was observed in all samples between the percentage of sperm cells with DNA strand breaks and the percentage of cells characterized by an increased sensitivity of DNA to denaturation (r = 0.87; P < = 0.05). It is likely, thus, that the sperm cells characterized by increased DNA sensitivity to denaturation have extensive DNA strand breakage. The above properties of abnormal sperm cells, which in all probability are dead in terms of their reproductive capacity, resemble properties of apoptotic somatic cells. Activation of the endogenous endonuclease which causes extensive DNA breakage, thus, is characteristic to both the programmed death of somatic cells (apoptosis) and functional elimination (of possibly defective) germ cells from the reproductive pool.

Apoptosis↗
PubMed1993

p145 expression during the cell cycle in HL-60 cell line and normal human lymphocytes: effects of camptothecin, vinblastine, cycloheximide, actinomycin D, retinoic acid and DMSO.

Bivariate flow cytometric analysis of nucleolar antigen p145 was performed on quiescent and phytohemagglutinin-stimulated human lymphocytes and on a promyelocytic cell line (HL-60). Data were acquired on a FACScan flow cytometer and analyzed using LYSYS II. Quiescent lymphocytes did not express p145. PHA-stimulated proliferating lymphocytes expressed p145 maximally after 48 h, similarly to HL-60 cells in exponential growth. Antigen expression in G1 was notably heterogeneous in both cell types. The ratio of p145/DNA was highest in early S and decreased during mid and late S and remained low in G2M. p145 expression was lowest in M-phase cells treated for 6 h with vinblastine. Cycloheximide and actinomycin D had similar effects on p145 in HL-60 cells: expression of p145 gradually decreased from 1 to 6 h incubation in all phases of the cell cycle. Camptothecin did not decrease p145 expression and apoptotic cells from CAM-treated cultures still expressed p145. Retinoic acid and DMSO induced differentiation in HL-60 cells, and as this process progressed, p145 levels gradually fell until they approached isotype antibody control levels at 9 and 6 days, respectively. However, after 5 days treatment with 2 nM retinoic acid apoptotic cells appeared which still expressed p145. The data on drug treatment suggest that p145 exists in undifferentiated and proliferating cells and may not be a specific marker for malignancy, but may prove useful as a monitor of chemotherapeutic effects in cancer treatment.

Antigens, Nuclear↗
PubMed1993

Apoptosis of S-phase HL-60 cells induced by DNA topoisomerase inhibitors: detection of DNA strand breaks by flow cytometry using the in situ nick translation assay.

DNA topoisomerase I inhibitor camptothecin (CAM), topoisomerase II inhibitors teniposide (TN) and amsacrine (m-AMSA) induce apoptosis of HL-60 cells. One of the early events of apoptosis is DNA degradation, which occurs as a result of activation of the specific endonuclease. DNA strand breaks generated during this process were revealed, in the present study, by the in situ nick translation assay which was adapted to flow cytometry. In this assay, the incorporation of biotinylated dUTP by apoptotic cells was detected by the use of fluorescinated avidin, whereas simultaneous staining of DNA with propidium iodide made it possible to correlate the appearance of DNA strand breaks with cell position in the cell cycle. The breaks were detected as early as 90 min after the initial cell contact with CAM, and they were limited to cells in the S phase of the cell cycle. At that early stage of apoptosis DNA was not yet extractable from the cells; the loss of DNA from S-phase cells could not be seen, by flow cytometry, during the initial 2 h of incubation with CAM. DNA strand breaks induced by TN and m-AMSA also occurred preferentially in S-phase cells. The data indicate that DNA strand breaks resulting from activation of endonuclease in HL-60 cells treated with DNA topoisomerase I or II inhibitors can be conveniently measured using the in situ nick translation assay. This assay has certain advantages over other methods of identification of apoptotic cells by flow cytometry, such as providing direct evidence of DNA damage and offering the opportunity to correlate DNA damage with cell position in the cell cycle. The method may be of interest in clinical oncology where testing tumor response (by DNA degradation) to DNA topoisomerase inhibitors or other treatments may be of prognostic value.

Amsacrine↗
PubMed1993

Pituitary microadenomas: early enhancement with dynamic CT--implications of arterial blood supply and potential importance.

In a search for early contrast medium enhancement, which can indicate the presence of direct arterial supply, a retrospective review of dynamic computed tomographic (CT) scans was performed in 260 patients with a pituitary microadenoma smaller than 10 mm in diameter. Fifty patients underwent examination with dynamic CT for nonendocrinologic disease as a control group to establish the normal pattern of pituitary gland enhancement. One hundred seventy microadenomas (65.4%) displayed the usual dynamic CT pattern (ie, they did not show early enhancement before that of the portal system of the pituitary gland: those pituitary microadenomas appeared less enhanced than the normal pituitary gland during the entire examination). On the other hand, in 90 microadenomas (34.6%), early partial or complete enhancement was seen within the microadenoma before the normal portal enhancement of the gland. Therefore, analysis with dynamic CT yields two groups of pituitary microadenomas separable on the basis of blood supply: those with portal blood supply only and those with partial or predominantly direct arterial blood supply; in theory, the second group avoids hypothalamic control.

Adenoma↗
PubMed1993

Proliferating cell nuclear antigen in archival surgical specimens of malignant lymphoma and metastatic carcinoma: immunohistochemical and flow cytometric analysis.

Immunohistochemical and flow cytometric multiparameter analysis of proliferating cell nuclear antigen (PCNA) was performed on fifteen formalin fixed, paraffin embedded lymph nodes with malignant lymphoma (eleven non-Hodgkin's lymphomas, four Hodgkin's lymphomas), and fifteen lymph nodes with metastatic carcinomas. A general concordance between PCNA measurement by both methods has been observed: the percentage of positively stained cells in tissue sections correlated well with the percentage of cells expressing this antigen in cell suspensions (r = 0.76). Both diploid and aneuploid tumors expressed PCNA, and a correlation between PCNA and the percent cells in S-phase was evident in both: in PCNA-positive tumors the mean percent of cells in S-phase was 16.5%, and in PCNA-negative tumors, 5.9%. The data indicate that PCNA can be detected in formalin-fixed tissues by either classic immunohistochemical analysis or by flow cytometry.

Adolescent↗
PubMed1993

[Programmed death of cells (apoptosis)].

Apoptosis is a mode of cell death defined by characteristic morphological, biochemical and molecular changes. It was first described as a "shrinkage necrosis", and then this term was replaced by apoptosis to emphasize its role opposite mitosis in tissue kinetics. During apoptosis the cell decrease in size, loose contact with neighboring cells, and loose specialized surface elements such as microvilli and cell-cell junctions. A shift of fluid out of the cells causes cytoplasm condensation, which is followed by convolution of the nuclear and cellular outlines. In later stages of apoptosis the entire cell becomes fragmented, forming a number of plasma membrane-bounded apoptotic bodies which contain nuclear and or cytoplasmic elements. The ultrastructural appearance of necrosis is quite different, the main features being mitochondrial swelling, plasma membrane breakdown and cellular disintegration. Apoptosis occurs in many physiological and pathological processes. It plays an important role during embryonal development as programmed cell death and accompanies a variety of normal involutional processes in which it serves as a mechanism to remove "unwanted" cells. Apoptosis is associated with prostate atrophy after castration or atrophy of the adrenal cortex and thymus after administration of glucocorticoids. Apoptosis is involved in elimination of CD4 T lymphocytes in the course of HIV infection. The interest in apoptosis in oncology stems from the fact that it occurs in tumors, spontaneously as well as triggered by different antitumor drugs, radiation or after withdraw of growth factors. Spontaneous apoptosis may play a role in evolution of tumor malignancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Apoptosis↗
PubMed1993

Induction of DNA strand breaks associated with apoptosis during treatment of leukemias.

A new flow cytometric method is described to detect DNA strand breaks associated with apoptosis, by labeling the 3'-OH termini in the breaks with biotinylated dUTP in a reaction employing exogenous terminal deoxynucleotidyl transferase. The method has been applied in studies on leukemic HL-60 and MOLT-4 cell lines to reveal whether it is specific to apoptotic cells, and whether it can be used in the clinic to detect DNA breakage in leukemic cells during chemotherapy. There was labeling of mononuclear cells in peripheral blood of all 11 patients studied during chemotherapy for acute lymphoblastic, acute myelogenous, or chronic myelogenous leukemia (ALL, AML, or CML) in blastic crisis, indicating induced DNA damage; the number of labeled cells increased from 1-8% before treatment up to 80% during the course of treatment. The DNA topoisomerase inhibitors mitoxantrone, VP-16 (etoposide), and m-AMSA (amsacrine) were more effective in inducing DNA breaks than was hydroxyurea or cytosine arabinoside (AraC). Cells with DNA breaks were identified in peripheral blood for up to 5 days following administration of Mitoxantrone and VP-16. In the case of DNA aneuploid leukemias, the DNA breaks were predominant in the aneuploid cell subpopulations, whereas presumably non-neoplastic diploid cells were unlabeled. In one case of ALL there were two distinct subpopulations of aneuploid cells: one responded to the treatment (by DNA breakage) and the other was non-responding. Thus, cells undergoing apoptosis can be detected by this method of labeling DNA strand breaks and the technique is applicable for analysis of response of leukemic cells to chemotherapy. With this method it may be possible to identify tumor cell sensitivity or resistance to particular drugs early in the course of treatment.

Adult↗
PubMed1992

Cell cycle-related expression of p120 nucleolar antigen in normal human lymphocytes and in cells of HL-60 and MOLT-4 leukemic lines: effects of methotrexate, camptothecin, and teniposide.

Expression of the proliferation-associated nucleolar antigen p120 was studied by flow cytometry in human quiescent and phytohemagglutinin-stimulated lymphocytes, as well as in human lymphocytic (MOLT-4) and promyelocytic (HL-60) cell lines. Bivariate analysis of p120 and DNA content made it possible to correlate p120 expression with cell position in the cycle. Proliferating lymphocytes and MOLT-4 and HL-60 cells had a similar pattern of p120 expression. Populations of G1 cells, in all three cell types, were very heterogenous with respect to p120, and a threshold in G1 was observed. The cells with a p120 level below the threshold value did not enter S phase. An increase in p120 was observed during progression through S phase, and the antigen was maximally expressed in G2 cells. The p120/DNA content ratio, however, was highest in late G1 cells (G1B) and was declining during S and G2. The data thus suggest that p120 may be degraded during mitosis and that the postmitotic cells inherit little, if any, of this protein; the antigen then accumulates predominantly during G1, and must reach a threshold level to enable the cells to enter S phase. Antigen p120 could not be detected in noncycling lymphocytes nor in HL-60 cells induced to myeloid differentiation by growth in the presence of dimethyl sulfoxide. Treatment of MOLT-4 cells with pharmacological concentrations of methotrexate, camptothecin, or teniposide induced cell arrest in S or G2; expression of p120 in the arrested cells was unchanged from that of untreated MOLT-4 controls at the same phase of the cycle. The level of p120 was minimal in MOLT-4 or HL-60 cells arrested in M phase by vinblastine, but vinblastine had no effect on p120 fluorescence of interphase cells. Camptothecin or teniposide induced apoptosis selectively in S phase of HL-60 cells; apoptotic cells from camptothecin-treated cultures, however, despite the marked nucleolysis, still expressed p120. The data on the drug-treated cells indicate that the p120 level in tumors of patients may be used as a marker of tumor/malignancy even in clinical samples obtained during treatment.

Antigens, Neoplasm↗
PubMed1992

Effect of ionic strength in immunocytochemical detection of the proliferation associated nuclear antigens p120, PCNA, and the protein reacting with Ki-67 antibody.

This study was aimed at revealing whether or not ionic interactions between the epitope of the antigen detected by Ki-67 antibody, or the proliferation-associated proteins PCNA or p120, and neighboring cellular constituents impede detectability of these antigens in HL-60 cells by indirect immunofluorescence assay. To this end, the ionic strength (NaCl concentration) of the solutions in which cells were suspended during their fixation with 0.5% paraformaldehyde was increased, to up to 1.65 M NaCl, to weaken the intra- and/or intermolecular ionic interactions during the process of crosslinking, and the cells were then immunostained. Fluorescence of cells reacting with Ki-67 antibody was maximally increased after their treatment with 1.15 M NaCl; the average increase was nearly 110% above the level seen with the standard methodology utilizing 0.15 M NaCl. The increase was greater for cells in the G1 phase of the cell cycle compared to cells in S or G2. Fluorescence of cells stained with the PCNA antibody was maximally enhanced after cell treatment with 0.65 M NaCl. The enhancement, however, varied depending on the source of the antibody; it was nearly 200% in the case of the antibody provided by Boehringer and over 100% by DAKO. Detection of the nucleolar antigen p120 was not significantly affected by 0.65-1.65 M NaCl. The data indicate that ionic interactions between cellular constituents indeed play a role in masking the epitope of PCNA and the antigen detected by Ki-67.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Cycle↗
PubMed1992

Features of apoptotic cells measured by flow cytometry.

The present review describes several methods to characterize and differentiate between two different mechanisms of cell death, apoptosis and necrosis. Most of these methods were applied to studies of apoptosis triggered in the human leukemic HL-60 cell line by DNA topoisomerase I or II inhibitors, and in rat thymocytes by either topoisomerase inhibitors or prednisolone. In most cases, apoptosis was selective to cells in a particular phase of the cell cycle: only S-phase HL-60 cells and G0 thymocytes were mainly affected. Necrosis was induced by excessively high concentrations of these drugs. The following cell features were found useful to characterize the mode of cell death: a) Activation of an endonuclease in apoptocic cells resulted in extraction of the low molecular weight DNA following cell permeabilization, which, in turn, led to their decreased stainability with DNA-specific fluorochromes. Measurements of DNA content made it possible to identify apoptotic cells and to recognize the cell cycle phase specificity of the apoptotic process. b) Plasma membrane integrity, which is lost in necrotic but not apoptotic cells, was probed by the exclusion of propidium iodide (PI). The combination of PI followed by Hoechst 33342 proved to be an excellent probe to distinguish live, necrotic, early- and late-apoptotic cells. c) Mitochondrial transmembrane potential, assayed by retention of rhodamine 123 was preserved in apoptotic but not necrotic cells. d) The ATP-dependent lysosomal proton pump, tested by the supravital uptake of acridine orange (AO) was also preserved in apoptotic but not necrotic cells. e) Bivariate analysis of cells stained for DNA and protein revealed markedly diminished protein content in apoptotic cells, most likely due to activation of endogenous proteases. Necrotic cells, having leaky membranes, had minimal protein content. f) Staining of RNA allowed for the discrimination of G0 from G1 cells and thus made it possible to reveal that apoptosis was selective to G0 thymocytes. g) The decrease in forward light scatter, paralleled either by no change (HL-60 cells) or an increase (thymocytes) of right angle scatter, were early changes during apoptosis. h) The sensitivity of DNA in situ to denaturation, was increased in apoptotic and necrotic cells. This feature, probed by staining with AO at low pH, provided a sensitive and early assay to discriminate between live, apoptotic and necrotic cells, and to evaluate the cell cycle phase specificity of these processes. i) The in situ nick translation assay employing labeled triphosphonucleotides can be used to reveal DNA strand breaks, to detect the very early stages of apoptosis.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Aspiration cytology in the diagnosis of malignant tumors in children.

On the basis of 65 fine-needle aspiration biopsies (FNAB) the efficiency of aspiration cytology combined with immunocytochemistry in preoperative diagnosis of malignant tumors in children was evaluated. All FNAB were performed under ultrasound or fluoroscopy guidance. There were no false-positive diagnoses. Morphological type of the tumor was properly diagnosed preoperatively in 50 patients. In 10 aspirates, the tumor type could not be diagnosed on the basis of light microscopy and immunocytochemistry. Due to technical reasons, five false-negative diagnoses (7.7%) were made. This study showed that FNAB is a safe, rapid, and effective method, which enables preoperative diagnosis of children's tumors in most cases.

Adolescent↗
PubMed1992

Aspiration biopsy of the breast tumors. Analysis of diagnostically difficult cases.

In this paper the authors described those of the breast fine needle aspirates which were difficult for interpretation e.g. clear-cut differentiation between benign and malignant character of the lesion could not be made. There were 40 such cases (1.96%) out of 2332 breast tumors biopsied between 1987-1989. In the cases that proved to be malignant on histology the authors described cytological features which speak in favour of malignancy. In the group of smears which proved to be benign on histology the authors described the features that imitate atypia and should not be misdiagnosed as malignant. The described criteria may be of some help in cytological diagnosis of breast masses.

Adenofibroma↗
PubMed1992

Binding properties and expression of the Fc gamma receptors on guinea pig peritoneal macrophages treated with inhibitors of glycosylation.

The effect of inhibition of glycosylation in guinea pig peritoneal macrophages on interaction of their surface Fc gamma receptors with guinea pig and rabbit IgG was studied. The inhibitors used were tunicamycin and monensin. The cells treated with tunicamycin incorporated markedly less [3H]mannose, bound less peanut (PNA) and wheat germ (WGA) lectins and showed diminished ability of binding IgG in comparison with control cells. Treatment of the cells with monensin resulted in an increased incorporation of [3H]mannose, increased binding of PNA but a decreased binding of WGA. Monensin affected binding of guinea pig IgG2 and rabbit IgG to macrophages and had no effect on guinea pig IgG1 binding. Analysis of binding parameters showed that although the number of IgG-binding sites on treated cells was significantly lower, especially in the case of tunicamycin, the apparent association constants were 2-4 times higher than in control cells. The effect of tunicamycin and monensin on parameters of binding of guinea pig IgG2 or rabbit immunoglobulins was much more pronounced than in the case of binding guinea pig IgG1. The results showed that glycosylation modulates expression and IgG binding ability of the Fc gamma receptors on the surface of guinea pig peritoneal macrophages.

Animals↗
PubMed1991

Effect of glycosylation inhibitors on binding properties of the Fc gamma receptors from guinea pig peritoneal macrophages.

Guinea pig peritoneal macrophages possess on their surfaces receptors for the Fc region of IgG immunoglobulins (Fc gamma R). The cells treated with the glycosylation inhibitors tunicamycin or monensin interacted with IgG with a higher affinity and showed a lower number of IgG-binding sites in comparison with control cells. This indicates that the interaction of IgG with the macrophage Fc gamma receptor depends on the degree of glycosylation of the cell surface glycoconjugates and that glycosylation of the macrophage Fc gamma receptor is important for the expression of the mature form of the receptor.

Animals↗
PubMed1991

Storiform neurofibroma (Bednar tumor). A case report.

Clinical data and morphological features of the storiform neurofibroma, presenting as an exophytic tumor of the trunk in 30 years old white man are described. This benign tumor, is microscopically similar to dermatofibrosarcoma protuberans and differs from the letter by the presence of melanin containing cells.

Adult↗
PubMed1990

[Immunocytochemical evaluation of estrogen receptors in histological specimens of primary breast cancer].

Immunocytochemical methods (ER-ICA) by using monoclonal antibodies were applied to determine the presence of estrogen receptors (ER) in 44 primary breast cancers in women. Of this 48% of the tumours were classified as receptor positive. In these tumours the nuclei had a clearly positive heterogenous colouration. In three cases a positive reaction was also found in benign epithelial cells of the breast. ER determinations by ER-ICA method were compared with quantitative analysis carried out by using radioligand and immunoenzymatic methods. There was a strong correlation between immunocytochemical ER evaluations and quantitative methods. We also found a correlation of the menopause state and patients age and ER content.

Adult↗
PubMed1990

Shedding of the guinea pig peritoneal macrophage Fc gamma receptor. Effect on apparent association constant and on the number of IgG-binding sites on the cells.

During our studies on the structure and properties of guinea pig peritoneal macrophage Fc gamma receptor we observed that these cells spontaneously release an IgG-binding material into supernatant. Although the shedding was accompanied by a decrease in IgG-binding ability of macrophages, the number of IgG-binding sites/cell before and after shedding was similar. However, macrophages after the shedding interacted with IgG with a lower apparent association constant, Ka. Therefore, we assume that the decrease of IgG-binding ability of cells was an effect of decrease in value of Ka. Experiments with protein synthesis inhibitors showed that the shed receptor is replaced by "de novo" synthesized receptor molecules.

Animals↗