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Cytophotometry of granulocytes in chronic granulocytic leukaemia patients. Part II. Effects of chemotherapy on cell kinetics and cytochemical parameters of granulocytes.

A successful course of chemotherapy (myelosan or combined regimens) results in a two to eight fold decrease of the percentage of S + G2 cells in the bone marrow and peripheral blood and in the restoration of normal morphometric and cytochemical patterns of polymorphonuclear neutrophils in the chronic phase of CGL. In those patients of the chronic phase not responding to chemotherapy and in blast crisis the proportion of S + G2 cells did not change after a course of chemotherapy. Cytostatic drugs killed cells in all phases of the cell cycle. The initial response to chemotherapy was a slight increase of the proportion of S + G2 cells in peripheral blood, apparently as a result of the block in cell cycle transition which was followed by a rapid drop of S + G2 proportion. During chemotherapy such phenomena as an appearance of tetraploid mature neutrophils and of aneuploid clones were occasionally observed.

Antineoplastic Combined Chemotherapy Protocols↗

[Frequency distribution of DNA in blood and bone marrow cells of children with acute non-lymphatic leukemia using impulse cytophotometry].

1. As compared with the peripheral blood lymphocytes of healthy subjects the cells in the mononuclear fraction of peripheral blood and in the bone marrow of children with ANLL show a significant higher S- and G2 + M-phase. 2. A high proliferative activity correlate with a bad prognosis or with reaching no haematological remission. 3. Frequently, aneuploid cell populations will occur in the morphological subtypes M 4 and M 5.

Acute Disease↗

DNA cytophotometry of voided urine sediment. Comparison with results of cytologic diagnosis and image analysis.

Measurements of nuclear DNA were performed in urothelial cells in 54 Feulgen-restained cytocentrifuge preparations of voided urine previously studied visually and with an image analysis system. The study included 30 patients with bladder tumors of various grades, 9 patients with prostatic disease and 15 control samples from normal donors. A number of additional control measurements were performed, including measurements in tissue samples of the 30 bladder tumors corresponding to the cytologic samples. It was documented that DNA can be measured in most urinary sediments. The diagnostic performance of the image analysis system reflected the DNA patterns in 47 of the 54 cases. In several instances, particularly in cases of prostatic disease, the image analysis system recognized abnormal DNA patterns in the absence of significant morphologic abnormalities in the urothelial cells. In seven cases, the image analysis findings failed to conform with the DNA patterns. The reasons for these surprising results are discussed, and future modifications of the image analysis system are proposed.

Aged↗

Extraction of cells from paraffin-embedded tissue sections for single-cell DNA cytophotometry.

A simple technique is presented for the isolation of cells from paraffin-embedded tissues for Feulgen DNA cytophotometric investigations. Tissue fragments from paraffin blocks were deparaffinized in xylene, rehydrated and refixed in a formalin solution and incubated in a solution of 0.5 pepsin in 0.25% hydrochloric acid. After filtration through a 70 micron mesh and centrifugation, the cells were smeared upon a glass slide. Comparison between the results obtained with freshly prepared imprints and with pepsin-extracted cells of the same tumor showed that the extraction technique does not influence the Feulgen reaction or the DNA distribution pattern. Investigations carried out on bladder and embryonal carcinomas have demonstrated that the method permits an analysis of histologically or histochemically identified tumor cells within individual tissue areas.

Cell Separation↗

Chromatin activation of blood lymphocytes detected by polarization microscopy and cytophotometry.

Blood lymphocytes exhibit chromatin activation upon incubation with substances to which a person is allergic. Chromatin activation can be detected by polarization microscopy. In this work, different methods of evaluating lymphocyte chromatin activation were compared in nine drug-allergic and in eight control subjects. All drug allergics had skin or mucosal involvement, ranging from localized minor herpetic lesions and purpurae through wheal and circumscribed bullous lesions to serum sickness as the most severe form. The mean path difference as the measure of nuclear birefringence was obtained by a polarization microscope using both white light and 551 +/- 7 nm monochromatic light. The equation of Brace-Köhler for converting compensation into path difference for each single cell was used in calculator programs. The values were compared with readings of a cytophotometer operated in transmission mode. Allergy "scores", deriving from the analysis of chromatin activation kinetics due to serial drug dilutions, were also compared using both methods. The results indicate a linear relationship between monochromatic and white light compensation readings and an exponential relationship between mean path difference and mean transmission values at different background amplifications. Operation of the photometer at 25.8% background amplification gave the best correlating results. The two methods gave identical results for the presence (ten tests) or absence (eight tests) of allergy.

Adolescent↗

[Determination of DNA content using cytophotometry in nuclear suspension conserved for prolonged periods (preliminary study)].

The variations in cytophotometric determination of DNA content were studied in a concentrated suspension of nuclei stored for prolonged periods. These came from normal and tumoral cells of an uterine leiomyosarcoma, fixed in formalin and included in paraffin. DNA was measured in normal and tumoral cells at days 0, 1, 7, 11, 19, 27 and 33. No significant variations in DNA measurement were observed. In a same day, there was an uniform DNA content, ploidy pattern and DNA index and an aneuploid pattern was diagnosed in all histograms. These results suggest that nuclear suspensions can be conserved for long periods and used for DNA content determination, allowing the access of remote places to these techniques.

Cytophotometry↗

[Cervix uteri cancer: determination of DNA content by means of cytophotometry].

ADN content was measured using cytofluorometry in 31 infiltrative uterine cervical carcinomas, using 22 routine cytologic cervical spreads (18 from archives) and 9 cellular spreads obtained from the fresh surgical specimen. Results were correlated to the clinical staging and morphological parameters of the tumor. Eight samples (25.8%) were diploid and 23 (74.2%) aneuploid. Women with diploid tumors were not significantly older. No correlation was found between ADN content and clinical staging, degree of histological differentiation, degree of cervical wall infiltration or the presence of vascular tumor permeations. All the cytological spreads were adequate for the cytofluorometric study. It is concluded that a high percentage of cervical carcinomas are aneuploid and that the present technique can be applied successfully to cytological samples.

Adenocarcinoma↗

[Determination of DNA content by means of cytophotometry. Technical and methodological considerations and its application in tumor pathology].

The cytofluorometric determination of ADN content in tumoral cells from 33 tumors is reported. Twenty five epithelial (2 benign and 23 malignant) and 8 non epithelial tumors (1 benign and 7 malignant) were studied. All the benign tumors had a normal ADN content (2c); on the other hand, 65% of malignant tumors had an abnormal ADN content. All non epithelial malignant tumors were aneuploid. Among aneuploid tissues, epithelial tumors were frequently tetraploid (4c) and non epithelial tumors were frequently polyploid 6c, 8c, 12c). It is concluded that this technique gives valuable information about the biological characteristics of tumoral cells and that its results could be incorporated to the routine anatomopathological study of tumoral lesions.

Aneuploidy↗

Heterogeneity of the MtTW15 mammosomatotropic tumor. I. Light microscopic evaluation of cell types by means of immunocytochemistry, morphometric quantitation, fluorescence cytophotometry and radioimmunoassay.

Large MtTW15 pituitary tumors produced 200- to 800-fold elevations in serum growth hormone (GH) and prolactin (PRL) levels. Female tumor hosts showed doubling in body weight, milk secretion, and a 2-fold hepatosplenomegaly. Pituitaries of host animals were reduced by about 50% in both weight and concentrations of GH and PRL. Large tumors were well-encapsulated, multinodular and showed variable amounts of necrosis and hemorrhage. Cytofluorometric analysis revealed a range of 100-fold in nuclear DNA content of tumor parenchymal cells which were chromophobic, pleomorphic and frequently mitotic. Concentrations of hormones in tumors were less than in normal pituitaries and highly variable with the ratio of GH/PRL ranging up to 30-fold within the same tumor. Immunostaining and linear scanning quantitation showed that about 50% of the tumor cells contained immunodetectable hormones. Comparison of immunostained adjacent sections showed that hormone-containing tumor cells were pleomorphic, unequally distributed within nodules, lacking in distinctive identifying morphological characteristics and that they contained GH or PRL but not both hormones simultaneously. Collectively our results show that large MtTW15 tumors are comprised of a markedly heterogeneous population of tumor cells and they suggest that the hormone-containing cells are monohormonal secreting tumor cells which can produce GH or PRL but not both hormones.

Animals↗