A morphological and cytological study of Petunia hybrida exposed to UV-B radiation

The aim of this study was to investigate whether the cytoskeleton, and in particular the microtubular system, is affected by enhanced levels of ultraviolet-B (280–320 nm, 9 kJ m⁻² day⁻¹ biologically effective UV-B radiation) radiation in epidermal cells of Petunia x hybrida Vilm, isolated from leaves of plants grown under UV-B radiation and visible light. In addition, morphological changes during development were monitored. In a previous study microtubules were depolymerized and delays in the different stages of the cell cycle were found when protoplasts of Petunia were irradiated with UV-B radiation (Staxén et al. 1993. Protoplasma 173: 70–76). Thus it was of interest to ascertain whether the cytoskeleton would be similarly affected in an intact system. Assuming an effect of UV-B radiation on the microtubular system, we wished to determine whether this could be correlated to concomitant changes in leaf morphology. Plants of Petunia hybrida were grown in greenhouse conditions in the presence or absence of UV-B radiation. During the course of the experiment, samples were taken from young, expanding leaves and from older, fully expanded leaves and prepared for localization and analysis of microtubules from the adaxial epidermal cells. Morphology rather than the cytoskeleton was affected by UV radiation, despite the fact that the epidermal cytoskeleton would most likely be affected, since it is located in the cells which form the first intercepting layer for incident radiation.
Morphological changes under UV-B radiation, as compared to those under control conditions, were reflected in earlier flowering and an increase in leaf number. Cell division was thus stimulated as was also evidenced from the increased leaf area. Our results indicate that the number of stomata differentiated on a leaf area basis was not altered although the number of stomata per epidermal cell was reduced.     

Radioprotection of hematopoietic tissues in mice by indomethacin

We recently reported that indomethacin, an inhibitor of prostaglandin (PG) synthesis, increased the radioresponse of PG-producing murine tumors, but it protected the hematopoietic system from radiation damage [Furuta et al., Cancer Res. 48, 3008-3013 (1988)]. Here we have investigated possible mechanisms responsible for the radioprotective effect of indomethacin. In the exogenous spleen colony assay, bone marrow cells from indomethacin-treated mice showed a similar radioresponse to those from mice not treated with indomethacin, thus excluding true radioprotection as a mechanism. Also, neither the total number of bone marrow cells nor the number of stem cells in bone marrow were affected by the treatment with indomethacin. However, indomethacin induced significant splenomegaly, which was associated with an increased number of both nucleated cells and hematopoietic stem cells in the spleen. The latter was determined by the exogenous spleen colony assay. Thus indomethacin protected hematopoietic tissue indirectly through stimulation of hematopoietic cells in the spleen. When indomethacin was combined with WR-2721, which is a true radioprotector, we obtained a greater radioprotective effect than with either used alone according to the endogenous spleen colony assay.     

Peripheral-type benzodiazepine receptors in human blood cells of patients affected by migraine without aura

The kinetic parameters at equilibrium of peripheral benzodiazepine receptors in platelets, lymphocytes and granulocytes of 15 patients affected by migraine without aura were tested using [3H]PK 11195, a specific radioligand for this receptor and compared with the same number of healthy controls: a statistically significant increase (platelets 212%, lymphocytes 203%, granulocytes 171%, as absolute percentage) in the maximal number of binding sites (B(max)) in all three patient samples, compared with healthy controls was detected; on the contrary, the values of the dissociation constant (K(d)) at equilibrium do not show any statistically significant variations between the two groups. These data further confirm the presence of peripheral biochemical alterations in migraine without aura. As peripheral benzodiazepine receptors appear to be involved in the regulation of the mitochondrial respiratory chain, the observed increase in B(max) might be related to the mitochondrial anomalies found in migraine disorders.     

Effect of electromagnetic microwave radiation on the growth of Ehrlich ascites carcinoma

Daily exposure of mouse recipients of Ehrlich ascites carcinoma to electromagnetic radiation of the microwave range leads to a change in the dynamics of tumor growth by decreasing the total number of cells. The number of tumor cells with blebbing morphological signs after microwave radiation increases gradually with tumor growth. The maximum content of tumor cells in the state of blebbing is observed during active proliferation in tumor-recipient mice of the control group (without irradiation).     

Effect of UV Radiation on the Bacterivory of a Heterotrophic Nanoflagellate

The effects of UV-B radiation on the heterotrophic nanoflagellate Bodo saltans (Kinetoplastida) were examined under controlled conditions with artificial UV sources and also under natural solar radiation in an oligotrophic lake. In both types of experiments, the characteristic elongated cell morphology of this flagellate changed into a spherical one. This effect was due to UV-B but also to UV-A radiation, and after 4 h of exposure at 0.5 m of depth, 99% (UV-B plus UV-A plus photosynthetically active radiation) and 69% of the cells (UV-A plus photosynthetically active radiation) were spherical. At 6 m of depth where only 10% of the UV-B (305 nm) at the surface was measured, no significant effect was observed. The spherical cells were nonmotile, but before the morphological change took place, the swimming speed was ca. 3.5 times lower in the plus-UV-B treatment. The negative relation between the abundance of spherical cells and the average ingestion of fluorescently labeled bacteria per cell indicates that these cells are not able to feed upon bacteria. In bacterivory experiments lasting for 6 h, the total number of grazed bacteria was up to 70% lower in the plus-UV-B treatment than in the control without UV-B. This resulted in a positive feedback between UV-B and bacterial growth. The high sensitivity of B. saltans to solar UV-B and UV-A radiation strongly reduces its ability to live near the surface at times of high UV radiation.     

Correlation between glucocorticoid receptor binding parameters, blood pressure, and body mass index in a healthy human population

Correlation between the glucocorticoid receptor (GR) number and affinity for the ligand, as well as the relationship between these equilibrium binding parameters and body mass index, blood pressure, and age were examined in peripheral blood mononuclear cells (PBMC) of healthy human subjects. It was found that the only statistically significant correlation was that between the GR number per cell and equilibrium dissociation constant, K(d) (r = 0.84, p < 0.0001). This observation implies the existence of a compensatory mechanism providing for lower GR affinity in individuals that have more receptor sites in circulating mononuclear cells and vice versa. This compensatory phenomenon together with considerable interindividual variation (GR number per cell ranging from 1391 to 15133, CV = 58.62%; and K(d) from 2.5 to 98.6 nM, CV = 80.87%), reflects plasticity of the glucocorticoid system. The results pose the question of whether this compensatory mechanism observed in healthy human subjects persists in pathophysiological states associated with glucocorticoid hormone actions and suggest that tissue sensitivity to glucocorticoids could be better predicted by the sign and magnitude of the correlation between the two receptor equilibrium binding parameters than by each of them separately.     

Global Stability of a System of Two Interacting Species with Convective and Dispersive Migration

Using Liapunov's direct method, effects of convective and dispersive migration on the global stability of the equilibrium state of a system of two interacting species are investigated. It is shown that the stable equilibrium state without dispersal remains so with dispersal. Further, it is pointed out that stability or instability of the equilibrium state of the system is not affected by convective migration. These results are justified in cases of a system of mutualistic interactions of species and a prey-predator system with functional response.     

The role of Bcl-X(S) in radiation sensitivity

Bcl-X(S) is a pro-apoptosis member of the Bcl2 family that has been shown to induce cell death and enhance chemosensitivity. We have investigated the effect of Bcl-X(S) overexpression on radiation sensitivity. Using a tetracycline-repressible system, we found that removal of tetracycline for 16 h induced Bcl-X(S) and reduced the surviving fraction of NIH 3T3 cells to 25%. However, radiation sensitivity was not significantly affected by Bcl-X(S) expression; the mean inactivation doses for Bcl-X(S) repressed and Bcl-X(S) induced cells were 2.7 +/- 0.3 and 2.3 +/- 0.1 Gy, respectively. We conclude that Bcl-X(S) induces cell death without affecting radiation sensitivity. These results suggest that mitochondrial pathways to apoptosis may not have a significant role in survival after irradiation.     

Dose-rate effect on proliferation suppression in human cell lines continuously exposed to γ rays

Irradiation time and dose rate are important factors in the evaluation of radiation risk for human health. We previously proposed a novel dose-rate effect model, the modified exponential (MOE) model, which predicts that radiation risks decline exponentially as the dose rate decreases. Here we show that, during the early phase of exposure, up to 1000 h, the proliferation of cells continuously exposed to γ rays at a constant dose rate is gradually suppressed, even as the total dose increases. This trend holds for a number of cell lines including tumor cells, nontransformed fibroblasts and leukocytes. The accumulation of total dose by longer exposure times does not increase this suppressive effect even in cells with a defective DNA repair system, suggesting that risk is determined solely by dose rate in the later phase. The dose-rate effect in the early phase follows the MOE model in DNA repair-proficient cell lines, while cells with impaired DNA-PK or ATM show no dose-rate effect. In the later phase, however, a certain dose-rate effect is observed even in mutant cell lines, and suppression of cell proliferation no longer follows the MOE model. Our results suggest that a distinct mechanism that can operate in the absence of intact DNA-PK or ATM influences the dose-rate effect in the later phase of continuous radiation exposure.     

Control of Ehrlich cell division by zinc

The nutritional requirement for zinc in the proliferation of normal and malignant cells has been demonstrated in a number of animal studies. A distinction is made between the effect of zinc status upon the host during carcinogenesis and tumor growth. The present studies focus on the Ehrlich ascites tumor in mice fed a semipurified zinc-deficient diet along with defined concentration of zinc in the drinking water. This model of zinc deficiency is compared with others in which chelating agents are used to create zinc-deficient conditions or the microorganismEuglena gracilis is examined in a defined zinc-deficient medium. It is reported here that Ehrlich cells remain quiescent for several weeks in severely deficient mice, suggesting their restriction to a G₁ or G₀ state of the cell cycle. The kinetics of thymidine and uridine uptake and incorporation into DNA and RNA in Zn-normal and Zn-deficient tumors is consistent with the inhibition of thymidine kinase and DNA polymerase in the Zn-deprived system, but with little effect on RNA synthesis. The concentration of metabolites of these labeled nucleosides in Ehrlich cells is also consistent with a primary effect upon thymidine kinase. Although the ascites fluid Zn is depressed in Zn deficiency, total cellular zinc and its distribution among cell fractions is not significantly affected. It is suggested that these effects are specific in nature and not the result of a general lack of zinc for zinc metalloproteins and other binding sites in the cell.     

Radiation-induced damage to DNA in drug- and radiation-resistant sublines of a human breast cancer cell line.

An Adriamycin-resistant subline of a human breast cancer cell line, MCF-7 ADRR, has been shown to exhibit radioresistance associated with an increase in the size of the shoulder on the radiation survival curve. In the present study, damage to DNA of MCF-7 sublines WT and ADRR by 60Co gamma radiation was measured by filter elution techniques. The initial amount of DNA damage, measured by both alkaline and neutral filter elution, was lower in ADRR cells, suggesting that these cells are resistant to radiation-induced single- and double-strand DNA breaks. In the case of double-strand breaks the difference between WT and ADRR cells was significant only at the lower radiation doses studied (up to 100 Gy). In cells depleted of glutathione (GSH) by L-buthionine sulfoximine (BSO) treatment, ADRR cells were sensitized to radiation-induced DNA damage, while WT cells were unaffected. The rate of repair of single- and double-strand DNA breaks following radiation was the same for both sublines, and repair of radiation damage was not affected by BSO treatment in either cell line. The relative resistance of ADRR cells to initial DNA damage by radiation is the only difference so far detected at the molecular level which reflects radiation survival, and it is possible that other factors are involved in the resistance of ADRR cells to killing by radiation. Sensitization of ADRR cells to radiation-induced DNA damage by GSH depletion, although not likely to involve inhibition of GSH-dependent detoxification enzymes per se (irradiation was done at 4 degrees C), suggests that at the molecular level radioresponse in this subline is related to maintenance of GSH/GSSG redox equilibrium.     

Yeast survival following gamma irradiation: delayed colony formation and determination of the proportion of undamaged cells

The experimental method is proposed to determine both a relative number of cells without damages and the effect of the delayed appearance of colonies of haploid and diploid yeast after exposure to ionizing radiation.     

Culture of human mast cells from peripheral blood progenitors

This protocol details a method for obtaining a considerable number of human mast cells from peripheral blood cells from normal donors without using stem cell mobilization treatment. By using the magnetic cell sorting system, 10(4)-10(5) cells are retrieved in the CD34+ fraction when 100 ml of blood is drawn from a healthy donor. When these cells are cultured using methylcellulose medium supplemented with stem cell factor and interleukin 6, 10(3)-10(4) mast cell colonies are formed in 6 weeks. The total mast cell number at 6 weeks of culture will be 10(6)-10(7).     

Reduction of transferrin receptor expression by interferon gamma in a human cell line sensitive to its antiproliferative effect

Interferon gamma (IFN gamma) reduced 125I-transferrin binding to WISH cells which are sensitive to its antiproliferative effect. IFN gamma did not affect transferrin binding to Daudi cells or phytohemagglutinin-stimulated human lymphocytes, neither of which respond to its antigrowth action. Scatchard analyses of the equilibrium binding of 125I-transferrin to WISH cells exposed to IFN gamma revealed a decrease in the number of cell surface receptors but no change in the apparent association constant compared with control cells. When 125I-transferrin binding was measured using detergent-extracted cells, the IFN-induced reduction of binding was smaller than with intact cells. This suggests that in WISH cells, IFN gamma not only reduced the total number of transferrin receptors, but also modified the process of receptor internalization and recycling. Labeling of newly synthesized receptors with [35S]-methionine indicated that a reduction in the biosynthesis might account for the decrease in the total number of transferrin receptors in IFN gamma-treated cells. Our results suggest that the antigrowth effect of IFN gamma is at least partly due to its inhibitory action on transferrin receptor expression leading to iron starvation.     

Effects of antibiotics that inhibit the bacterial peptidoglycan synthesis pathway on moss chloroplast division

Moss chloroplasts should prove useful for studying the cyanobacteria-derived system in chloroplasts. To determine the effects of antibiotics that inhibit bacterial peptidoglycan synthesis, the numbers of chloroplasts in treated Physcomitrella patens cells were counted. Ampicillin and D-cycloserine caused a rapid decrease in the number of chloroplasts per cell. Fosfomycin affected half of the cells, while vancomycin affected a few cells. Conversely, bacitracin had no effect. With the decrease in chloroplast number, macrochloroplasts appeared in antibiotic-treated cells. Removal of the antibiotics resulted in the recovery of chloroplast number, suggesting that the decrease in number was directly dependent on the antibiotic treatment. Microscopic observations showed that the decrease in the number of chloroplasts resulted from cell division without chloroplast division. These results suggest that enzymes derived from the bacterial peptidoglycan synthesis pathway are related to moss chloroplast division.     

The intestinal microflora of persons subjected to a radiation lesion]

The content of large intestine has been studied in persons exposed to radiation injury in consequence of the accident at the Chernobyl Atomic Power Plant. It is stated that bifidobacteria (10(7)-10(10) cells in 1 g of feces) prevailed (as in healthy people), Bifidobacterium indicum being a dominating species. Amount of lactic-acid bacteria in 1 g of defecations of examined patients was within the range of 10(6)-10(9) cells and in certain persons it reached 10(10) cells (primarily fecal Enterococci). A considerable amount of patients with acute radiation sickness of the 3d degree had in their intestine 10(9)/g of lactic-acid bacteria, Lactobacillus casei and L. plantarum prevailing there. The frequency of yeast isolation from defecations of patients constituted 83%, while the number of cells in 1 g of feces--from 10 to 10(4). Yeast of the Candida genus, mainly Candida parapsilosis, prevailed. The species composition of isolated microorganisms has no substantial differences from microcenosis of healthy people. The content of intestine of persons suffered from radiation is characterized only by greater amount of lactic-acid bacteria and enterococci as compared with healthy adults.     

Fatal disease and demographic Allee effect: population persistence and extinction

If a healthy stable host population at the disease-free equilibrium is subject to the Allee effect, can a small number of infected individuals with a fatal disease cause the host population to go extinct? That is, does the Allee effect matter at high densities? To answer this question, we use a susceptible-infected epidemic model to obtain model parameters that lead to host population persistence (with or without infected individuals) and to host extinction. We prove that the presence of an Allee effect in host demographics matters even at large population densities. We show that a small perturbation to the disease-free equilibrium can eventually lead to host population extinction. In addition, we prove that additional deaths due to a fatal infectious disease effectively increase the Allee threshold of the host population demographics.     

Effects of γ radiation on the reproduction and enteroendocrine cells of the spruce bark beetle <Emphasis Type="Italic">Ips typographus</Emphasis> and prospects for its control

The European spruce bark beetle Ips typographus is one of the most important pests of spruce forests in Europe. The present study investigated the feasibility of a sterile insect technique (SIT) for this pest control. Laboratory-reared males were exposed to various doses of γ radiation (0, 10, 30, 50, 70 and 90 Gy) and allowed to mate with laboratory-reared untreated females. The radiation significantly affected the number of viable offspring. Although females oviposited on average 16.0–18.8 eggs per maternal tunnel in all treatments, the number of larval tunnels per maternal tunnel observed after 4 weeks of development decreased from 17.3 in the control (0 Gy) to 4.5 in the 90-Gy group. Numerous oviposition sites without larval tunnels were observed in galleries when males received high doses (70 and 90 Gy) of γ radiation. Comparison of the sperm viability in the control and irradiated males did not reveal any statistically significant differences. Side effects of the irradiation were examined by immunostaining of the enteroendocrine cells. Two distinct types of cells were revealed with antibodies to neuropeptides allatostatin A and tachykinin in the midgut. The alimentary tracts of males receiving a high dose of radiation (90 Gy) showed a significantly decreased number of tachykinin-like immunoreactive enteroendocrine cells. This observation suggests potential radiation-induced damage of the digestive system, which could lead to a reduction in male fitness. The implications of these findings for successful use of SIT in spruce bark beetle control are discussed.     

Superovulatory responses of Holstein cows

Approximately 1000 registered cows and heifers were superovulated one to 10 times. Nonsurgical embryo recoveries were performed on all donors which exhibited estrus. Healthy donors produced more total ova and cleaving embryos and had a higher ovum recovery rate, fertilization rate and pregnancy rate from embryos transferred than did cows classified as infertile. While ovum number was not affected during 10 repeated superovulations, fertilization rate and embryo number decreased. The number of ova recovered from healthy cows was affected by season, and from infertile cows by the day of the estrous cycle on which FSH was started and by the number of days since calving. More ova were recovered from infertile cows synchronized with prostaglandins prior to superovulation than following a natural estrous cycle. The number of embryos recovered from infertile cows was affected by age and from healthy cows by daily milk production. Fertilization rates in both healthy and infertile cows were affected by age, time since calving, daily milk production, day of cycle FSH was injected and season. There was no effect of the day of recovery on the number of ova or embryos recovered from healthy or infertile cows.