Flow cytometric DNA measurements in paraffin-embedded bladder carcinoma tissue before and after precystectomy radiotherapy.

From 1976 through 1983, 48 patients with T2/T3 bladder carcinomas were treated with pre-operative radiotherapy (46 Gy) and total cystectomy. DNA histograms were recorded by flow cytometry (FCM) from all pre-treatment paraffin-embedded tumour biopsies and, if still present after radiotherapy, from tumour tissue from the cystectomy specimen. Five patients were excluded from the study because no DNA histograms could be recorded due to extensive destruction of the cell nuclei during preparation. Thus, 43 patients are completely evaluable. Before the start of radiotherapy, 32 bladder carcinomas were interpreted as non-diploid, whereas 11 were diploid. Non-diploidy of the pre-treatment biopsy was associated with radiotherapy-induced stage reduction (p = 0.13). The survival rates in patients with diploid and non-diploid tumours were 65% and 45% respectively at 4 years (p = 0.13). Although not statistically significant, our results suggest that DNA-FCM may provide clinically relevant information about the tumour biology and response to radiotherapy concerning bladder carcinomas.

[Mutability of the liver cells in the progeny of irradiated male rats]. / Mutabil'nost' kletok pecheni potomstva obluchennykh samtsov krys.

The descendants of the first generation of radiation exposed rat males exhibited an increased sensitivity of liver cells to the mutagenic effect of ionizing radiation as was determined by the incidence of chromosome aberrations. It is assumed that the stability of the hereditary apparatus of these animal cells is diminished due to the presence therein of radiation induced injuries of mutation and epigenomic origin at a heterozygous condition.

[Cytofluorimetric study of ploidy in the preleukemic thymus in C57BL/Ka mice under fractionated irradiation]. / Etude cytofluorimétrique de la ploïdie dans le thymus préleucémique chez la Souris C57B1/Ka soumise à une irradiation fractionnée.

The ploidy of the thymus was studied in C57B1/Ka mice irradiated with 4 weekly X-Ray doses of 1.75 Gy. The determination of nuclear DNA content was performed by flow cytometry of intact thymocytes labeled with propidium iodide in presence of a mixture of chicken and rainbow trout red blood cells as internal reference standards. The method has been tested by detecting the sex difference in DNA content of G0/G1 of normal thymic mouse cells. The mean value was 2.9% higher in female mice. The thymus of almost 60% of irradiated male mice present a slight hypoploidy of 2.6% one month after the last irradiation.

Growth kinetics as a function of ploidy in diploid, tetraploid, and octaploid smooth muscle cells derived from the normal rat aorta.

The smooth muscle cell population in major arteries of humans and experimental animals is heterogeneous with regard to cellular DNA content. A proportion of cells has polyploid DNA content and this proportion increases with normal aging and with hypertension. We have isolated pure populations of rat aortic smooth muscle cells containing 2C, 4C, and 8C DNA content by cloning of cultures of cells previously subjected to flow cytometric cell sorting. Karyologic analysis of these clonal populations revealed them to be pure diploid, tetraploid, and octaploid populations, respectively, containing 2N (= 42), 4N, and 8N chromosomes. Cell attachment area and nuclear size appeared to increase with the level of ploidy. Studies of the proliferative characteristics of the cells revealed that the growth rate and ultimate cell densities achieved decreased as the ploidy level increased. The intrinsic cellular radiosensitivity of these clones did not vary with ploidy. Increased smooth muscle cell ploidy is, therefore, associated with a decreased rate of proliferation. The emergence of smooth muscle cells with polyploid DNA content under normal and pathologic conditions is probably due to mitotic polyploidization without net cell proliferation and may be related to the need for expression of differentiated functions.

Longterm effect of neonatal whole-body X-ray exposure on ploidy development in mouse liver cells.

Mice of the strains C3H, DBA, NZB, and NMRI were X-irradiated with 0.475 Gy and 2.85 Gy (or 3.80 Gy) at newborn stage or at the age of six months. Their livers were prepared at the age of one to maximum 18 months and the hepatocytes isolated. The DNA content of both whole cells and separated nuclei was measured by flow cytometry and the liver ploidy pattern was determined applying a correcting calculation. Irradiation with 0.475 Gy of newborns of strain C3H results in a relative decrease of octoploid cells and an increase of tetraploid nuclei during lifetime (significance level of 0.001). After irradiation with 3.80 Gy a reversed reaction was observed. However, if the animals were irradiated as adults with a dose of 3.80 Gy the 8c cells decreased significantly three months later. Similarly, newborns of the outbred strain NMRI irradiated with 2.85 Gy, showed an increase of octoploid cells from the third month on, but an oscillating pattern of the 4c nuclear DNA content. In strains DBA and NZB, significantly altered ploidy values could not be found, perhaps due to a too great variance of experimental values.

Predicting radiocurability.

Several predictors of tumor radiocurability are already integrated into clinical practice, e.g., tumor size, gross morphologic features (i.e., infiltrative or exophytic), histologic type, and grade. These are relatively imprecise, however, and none is specific. The aim of research into predictive assays is not only to refine the discrimination of existing predictors but also to suggest specific experimental approaches for overcoming tumor radioresistance in individual patients. Two broad categories of predictive assays can be defined: direct and indirect measurements of tumor cell survival and/or repair capability following irradiation and measurement of cellular and extracellular parameters affecting radiosensitivity. Examples from ongoing research at the University of Texas M. D. Anderson Hospital using one technique from each category (the micronucleus assay and flow cytometric analysis of tumor cell proliferation kinetics and ploidy) are used to illustrate potential methods of selecting patients for fast neutron radiotherapy.

[Variability of cultures of Pullularia pullulans with different ploidy levels]. / Izmenchivost' kul'tur Pullularia pullulans, imeiushchikh raznyi uroven' ploidnosti.

As the level of ploidy rises in Pullularia pullulans, this causes an increase in the frequency of spontaneous and UV-induced auxotrophic mutants as well as mutants with a modified respiration activity while the frequency of morphological mutants decreases. The latter can arise as a result of recessive and dominant mutations. A higher frequency of morphological mutants in the haploid may be result of recessive mutations. It is likely that the frequency of dominant mutations increases in cultures with a higher level of ploidy since, as the difference between the frequency of UV-induced mutants and the frequency of spontaneous morphological mutants increases.

Decreased acquisition of glucocorticoid resistance in tetraploid human lymphoid cells.

Resistance to dexamethasone (1 microM) was measured in glucocorticoid-sensitive diploid and tetraploid clones of the human leukaemic cell line CCRF-CEM (clone C7) during continuous culture and after X-ray or chemical mutagenesis. In continuous culture resistant diploid cells accumulated at a rate of about one cell per 10(5) divisions, while the rate for tetraploid cells was less than one per 10(7) divisions. Chemical and X-ray mutagenesis caused a marked increase in the number of resistant diploid cells but had very little effect on tetraploid cells. These results are consistent with a mutational basis for the acquisition of the glucocorticoid-resistant phenotype in human lymphoid cells.

Variance of ploidy in Candida albicans.

Determination of ploidy was performed on isolates of Candida albicans from clinical sources by measuring nuclear DNA content with fluorescent microscope photometry. By this criterion and UV irradiation survival experiments, haploid, diploid, and tetraploid strains were identified in this organism. The dimensions of nucleus-associated organelles (equivalent to spindle pole bodies) in these strains increased as a function of ploidy number.

[Quantitative analysis of yeast cell inactivation under the action of UHF radiation]. / Kolichestvennyi analiz inaktivatsii drozhzhevykh kletok pod deistviem SVCh-radiatsii.

A particular case of the earlier suggested probability model of the action of electromagnetic irradiation on the cell is described. The model has been used for the quantitative analysis of inactivation forms and survival curves of yeast cells of different strains exposed by electromagnetic field of SHF-range. The results of calculation and experimental data show that the probability model is suitable for both qualitative and quantitative description of different cell responses to SHF-irradiation.