Induction of chromosome shattering by ultraviolet light and caffeine: The influence of different distributions of photolesions

Cells of synchonized and of asynchronously growing cultures of a V79 Chinese hamster line were microirradiated with a low poweer laser-UV-microbeam of wavelength 257 nm. Ultraviolet light was either focused onto a small part of the nucleus (mode I) or distributed over the whole nucleus (mode II). Following microirradiation, the cells were incubated for 7–20 h with caffeine (1–2 mM) until chromosome preparation was performed. After both modes of microirradation, shattering of the entire chromosome complement (generalized chromosome shattering, GCS) was observed. It is suggested that the probability by which GCS is induced depends on the total number lesions rather than on their distribution in the chromatin. The results are consistent with the prediction of a “factor depletion model” wich assumes that in a given cell, GCS takes place both in irradiated and non-irradiated chromosomes of the total number of daughter strand-repair sites supasses a threshold value

Induction of chromosome shattering by ultraviolet light and caffeine

Synchronized and asynchronously growing cells of a V79 sub-line of the Chinese hamster were either partial-cell irradiation (λ, 254 nm) or laser-UV-microirradiated (λ, 257 nm). Post-incubation with caffeine (1–2 mM) often resulted in chromosome shattering, which was a rare event in the absence of this compound. In experiments with caffeine, the following results were obtained. Shattering of all the chromosomes of a cell (generalized chromosome shattering, GCS) was induced by partial-cell irradiation at the first post-irradiation mitosis when the UV fluence exceeded and “threshold” valued in the sensitive phases of the cell cycle (G1 and S). GCS was also induced by laser-UV-microirradiation of a small part of the nucleus in G1 of S whereas microirradiation of cytoplasm beside the nucleus was not effective. An upper limit of the UV fluence in the non-irradiated nuclear part due to scattering of the microbeam was experimentally obtained. This UV fluence was significantly below the threshold fluence necessary to induce GCS in whole-cell irradiation experiments. In other cells, partial nuclear irradiation resulted in shattering of a few chromosomes only, while the majority remained intact (partial chromosomes shattering, PCS). G1/early S was the most sensitive phase for induction of GCS by whole-cell and partial nuclear irradiation. The frequency of PCS was observed to increase when partial nuclear irradiation was performed either at lower incident doses or at later stages of S. We suggest that PCS and GCS indicate 2 levels of chromosome damage which can be produced by the synergistic action of UV irradiation and caffeine. PCS may be restricted to microirradiated chromatin whereas GCS involves both irradiated and unirradiated chromosomes in the microirradiated nucleus

Induction of chromosome damage by ultraviolet light and caffeine: Correlation of cytogenetic evaluation and flow karyotype

Asynchrononously growing cells of a M3-1 Chinese hamster line were ultraviolet (UV) irradiated ( = 254 nm) with UV fluences up to 7.5 J/m2. After irradiation, cells were incubated with or without 2 mM caffeine for 20 hr, then mitotic cells were selected by mechanical shaking. Their chromosomes were isolated, stained with Hoechst 33258 and chromomycin A3, and measured flow cytometrically. While the fluorescence distributions of chromosomes (flow karyotypes) from cells treated with UV alone or with caffeine alone were very similar to those of untreated controls, the flow karyo-types of UV + caffeine-treated cells showed a debris continuum that increased with increasing UV fluence suggesting an increased number of chromosome fragments. Visual evaluation of metaphase plates revealed that the percentage of cells with chromosome damage also increased steadily with increasing UV fluence. A high degree of correlation was observed between the relative magnitude of the debris level from flow karyotypes and the percentage of cells with chromosome damage and with generalized chromosome shattering, respectively, as determined from metaphase spreads

Laser-UV-microirradiation of interphase nuclei and posttreatment with caffeine: a new approach to establish the arrangement of interphase chromosomes

Laser UV microirradiation of Chinese hamster interphase cells combined with caffeine post-treatment produced different patterns of chromosome damage in mitosis following irradiation of a small area of the nucleus that may be classified in three categories: I) intact metaphase figures, II) chromosome damage confined to a small area of the metaphase spread, III) mitotic figures with damage on all chromosomes. Category III might be the consequence of a non-localized distortion of nuclear metabolism. By contrast, category II may reflect localized DNA damage induced by microirradiation, which could not be efficiently repaired due to the effect of caffeine. If this interpretation is right, in metaphase figures of category II chromosome damage should occur only at the irradiation site. The effect might then be used to investigate neighbourhood relationships of individual chromosomes in the interphase nucleus

UV micro-irradiation of the Chinese hamster cell nucleus and caffeine post-treatment immunocytochemical localization of DNA photolesions in cells with partial and generalized chromosome shattering

UV micro-irradiation of a small part of the Chinese hamster nucleus and caffeine post-incubation often results in shattered chromosomes at the first post-irradiation mitosis. In some of these mitotic cells, chromosome shattering is restricted to a few chromosomes spatially related in a small area of the metaphase spread; in others, shattering includes the whole chromosome complement. These 2 types of damage have been called partial and generalized chromosome shattering (PCS and GCS). Using antisera that specifically react with UV-irradiated DNA, we identified micro-irradiated chromatin in interphase nuclei and in mitotic cells with PCS or GCS by indirect immunofluorescence microscopy. In PCS, immunofluorescence staining was found in the damaged area, while the surrounding intact chromosomes were not stained. In GCS, staining was also restricted to a small region of the shattered chromosome complement. In other experiments, cells synchronized in G1 were micro-irradiated in the nucleus, pulse-labelled with [3H]thymidine and post-incubated with caffeine. Autoradiographs of cells with GCS showed unscheduled DNA synthesis restricted to a small chromatin region. Our data present direct evidence that the distribution of DNA photolesions does not coincide with the sites of chromosomal damage in GCS. As a working hypothesis, we propose that an indirect mechanism is involved in the induction of GCS by which DNA photolesions in a small nuclear segment induce shattering of both micro-irradiated and non-irradiated chromosomes

Cellulose dissolution in an alkali based solvent: influence of additives and pretreatments

The distinction between thermodynamic and kinetics in cellulose dissolution is seldom considered in the literature. Therefore, herein an attempt to discuss this topic and illustrate our hypotheses on the basis of simple experiments was made. It is well-known that cellulose can be dissolved in a aqueous sodium hydroxide (NaOH/H2O) solvent at low temperature but it is here shown that such an alkaline solvent can be considerably improved regarding solubility, stability and rheological properties as a whole if different additives (salts and amphiphilic molecules) are used in the dissolution stage. This work probes new aqueous routes to dissolve cellulose, thereby improving the potential to commercially dissolve cellulose in an inexpensive and environmentally friendly manner.A distinção entre termodinâmica e cinética de dissolução da celulose raramente tem sido considerada na literatura. Neste trabalho, discutimos este tema e fundamentamos as nossas hipóteses recorrendo a experiências simples. É do conhecimento geral que a celulose pode ser dissolvida no solvente aquoso de hidróxido de sódio (NaOH/H2O) a baixa temperatura. Neste trabalho, demonstramos que este solvente alcalino pode ser consideravelmente melhorado em relação à sua estabilidade, solubilidade e propriedades reológicas se forem usados diferentes aditivos (sais e moléculas anfifílicas) na fase de dissolução. Este trabalho indica novos caminhos relativamente à dissolução da celulose em solventes aquosos, de uma forma mais econômica e ambientalmente amigável, aumentando o seu potencial comercial

Analysis of chromosome positions in the interphase nucleus of Chinese hamster cells by laser-UV-microirradiation experiments

Unsynchronized cells of an essentially diploid strain of female Chinese hamster cells derived from lung tissue (CHL) were laser-UV-microirradiated (=257 nm) in the nucleus either at its central part or at its periphery. After 7–9 h postincubation with 0.5 mM caffeine, chromosome preparations were made in situ. Twenty-one and 29 metaphase spreads, respectively, with partial chromosome shattering (PCS) obtained after micro-irradiation at these two nuclear sites, were Q-banded and analyzed in detail. A positive correlation was observed between the frequency of damage of chromosomes and both their DNA content and length at metaphase. No significant difference was observed between the frequencies of damage obtained for individual chromosomes at either site of microirradiation. The frequency of joint damage of homologous chromosomes was low as compared to nonhomologous ones. Considerable variation was noted in different cells in the combinations of jointly shattered chromosomes. Evidence which justifies an interpretation of these data in terms of an interphase arrangement of chromosome territories is discussed. Our data strongly argue against somatic pairing as a regular event, and suggest a considerable variability of chromosome positions in different nuclei. However, present data do not exclude the possibility of certain non-random chromosomal arrangements in CHL-nuclei. The interphase chromosome distribution revealed by these experiments is compared with centromere-centromere, centromere-center and angle analyses of metaphase spreads and the relationship between interphase and metaphase arrangements of chromosomes is discussed

Carbon Formation in the Reforming of Simulated Biomass Gasification Gas on Nickel and Rhodium Catalysts

Biomass gasification gas contains hydrocarbons that must be converted to CO and H2 prior to the utilization of the gas in a synthesis unit. Autothermal or steam reforming operating with a nickel or noble metal catalyst is a feasible option to treat the gas, but the harsh reaction conditions may lead to the formation of solid carbon. This study discusses the effects of pressure, time-on-stream, and ethylene content on the carbon formation on nickel and rhodium catalysts. The experiments were carried out with laboratory-scale equipment using reaction conditions that were closely simulated after a pilot-scale biomass gasifier. The results indicated that ethylene content above 20,000 vol-ppm and the increased pressure would increase the carbon formation, although there were differences between the rhodium and nickel catalysts. However, carbon formation was significantly more pronounced on the nickel catalyst when the reaction time was increased from 5 h to 144 h. The type of carbon was found to be primarily encapsulating and graphitic. The formation of whisker carbons (also known as carbon nanotubes) was not observed, which is consistent with the literature as the feed gas contained H2S. It was concluded that utilizing a noble metal catalyst as the front layer of the catalyst bed could lower the risk for carbon formation sufficiently to provide stable long-term operation