Effect of estradiol and ethynylestradiol on microtubule distribution in Chinese hamster V79 cells.

The effects of estradiol (E2) and ethynylestradiol (EE2) on the chromosome number and cellular microtubule architecture of Chinese hamster V79 cells were studied using fluorescent anti-tubulin antibody. Treatment with 20 microM E2 for 48 h induced only a small amount of tetraploid cells, but the normal microtubule network was disrupted completely by only 3 h of treatment. This data reveals that E2 has higher microtubule-disruptive activity than diethylstilbestrol in V79 cells.     

Enhanced cytotoxicity in simultaneous and sequential drug-heat treatments of cultured Chinese hamster V79 cells

Under hyperthermal conditions, some genotoxic drugs such as bleomycin, paraquat, and N-alkyl-N-nitrosoureas exhibit increased cytotoxicity toward cultured Chinese hamster V79 cells. Sequential combinations of heat and drug treatments, regardless of whether drug-exposure precedes or follows hyperthermia, also induce synergistic cytotoxicity to some extent. This may be attributed not only to the relationship of temperature and chemical injury as defined by the Arrhenius law, but also to a lethal interaction between the biological consequences of chemical injury and thermal damage. Ethanol, dimethylsulfoxide (DMSO), ethylenediaminetetraacetic acid (EDTA), and urea, which are known to affect cell membrane and protein, also exert synergistic cytotoxicity at 43 degrees C at a dose range that is nontoxic at 37 degrees C. When used sequentially with thermal treatment, they also proved to be synergistic. Glycerol, however, protected cells against thermal damage when used in a simultaneous chemical-thermal combination. But when treatments were carried out sequentially, glycerol proved destructive.     

Photogenotoxicity of fluoroquinolones in Chinese hamster V79 cells: dependency on active topoisomerase II

The Chinese hamster V79 lung cell in vitro micronucleus assay was adapted to detect and quantify phototoxicity and photogenotoxicity of fluoroquinolones. Using this assay, the quinolones were ranked in terms of decreasing phototoxicity: clinafloxacin > lomefloxacin, sparfloxacin > trovafloxacin, nalidixic acid, ofloxacin, ciprofloxacin > enoxacin, norfloxacin. This rank order agrees well with published studies utilizing various other phototoxicity models and establishes this approach as a fast and sensitive way to characterize the phototoxic potential of quinolones. Nearly complete inhibition of phototoxicity was observed if the cells were pretreated for as little as 1 min with 10-20 mM sodium azide prior to the addition of quinolone. An identical azide effect was seen in unirradiated quinolone- and etoposide-treated cells. These findings are consistent with a model in which sodium azide renders DNA topoisomerase II catalytically inactive. In this state, topoisomerase II cannot initiate DNA strand cleavage and the DNA/topoisomerase complex becomes insensitive to quinolones and other topoisomerase II inhibitors. The fact that azide reduces both UV-dependent and UV-independent toxicity and clastogenicity strongly suggests a common mechanism of toxicity dependent on the formation of topoisomerase-induced DNA double-strand breaks.     

Multiplex polymerase chain reaction analysis of UV-A- and UV-B-induced delayed and early mutations in V79 Chinese hamster cells

We previously reported that approximately 10% of V79 Chinese hamster fibroblast populations clonally derived from single cells immediately after irradiation with either ultraviolet B (UV-B, 290-320 nm, mainly 311 nm) or ultraviolet A (UV-A, 320-400 nm, mainly 350-390 nm) radiation exhibit genomic instability. The instability is revealed by relatively high mutation frequencies in the hypoxanthine phosphoribosyl transferase (hprt) gene up to 23 cell generations after irradiation. These delayed mutant clones exhibited higher levels of oxidative stress than normal cells. Therefore, persistently increased oxidative stress has been proposed as a mechanism for UV-induced genomic instability. This study investigates whether this mechanism is reflected in the deletion spectrum of delayed mutant clones. Eighty-eight percent of the delayed mutant clones derived from UV-A-irradiated populations were found to have total deletion of the hprt gene. Correspondingly, 81% of UV-A-induced early mutations (i.e. detected shortly after irradiation) also had total deletions. Among delayed UV-B-induced mutant clones, 23% had total deletions and 8% had deletion of one exon, whereas all early UV-B events were either point mutations or small deletions or insertions. In conclusion, the multiplex polymerase chain reaction deletion screen showed that there were explicit differences in the occurrence of large gene alterations between early and delayed mutations induced by UV-B radiation. For UV-A radiation the deletion spectra were similar for delayed and early mutations. UV-A radiation is, in contrast to UV-B radiation, only weakly absorbed by DNA and probably induces mutation almost solely via production of reactive oxygen species. Therefore, the present results support the hypothesis that persistent increase in oxidative stress is involved in the mechanism of UV-induced genomic instability.     

Biological activities of phthalocyanines--III. Photoinactivation of V-79 Chinese hamster cells by tetrasulfophthalocyanines

Abstract— Tetrasulfophthalocyanine and a series of its metal chelates were tested for their ability to photoinactivate V-79 Chinese hamster cells. Incubation of cells for 1 h with tetrasulfophthalocyanine at 5 μM effectively sensitized cells towards red light. At the 1% survival level, the dye was 4 x more efficient than hematoporphyrin, efficiency being defined in terms of drug concentration in the medium and incident light fluence rather than on the basis of quanta absorbed. Chelation of the dye with metal ions resulted in most cases in a greatly diminished photosensitizing effect, except for cerium. The cerium complex was about 5 x more effective for cell killing than the metal free tetrasulfophthalocyanine and 20 x more efficient as compared to hematoporphyrin. Hypoxic conditions resulted in total loss of photoactivity indicating the involvement of oxygen in the action mechanism. The inactivation by near-UV light by these drugs was also investigated. The potential of sulfonated phthalocyanines as novel photosensitizers for photodynamic cancer therapy is discussed.     

Changes in cell cycle distribution of V79 Chinese hamster fibroblasts after irradiation at different wavelengths

Changes in cell cycle distribution of V79 Chinese hamster fibroblasts were investigated at different wavelengths between 254 and 313 nm. The fluences applied led to surviving fractions of 0.61. In all cases, the S fraction was temporarily increased within 8-12 h after irradiation, whereas the G1 fraction was decreased. The maximum deviations from the initial values did not significantly depend on the wavelength.     

Photosensitization of Chinese hamster cells by water-soluble phthalocyanines

Abstract— Phthalocyanines are efficient photosensitizers of cultured mammalian cells and are considered for use in photodynamic therapy. The photobiological properties of chloroaluminum phthalocyanine sulfonate (AIPCS) were compared to those of the unsubstituted, water-insoluble derivative (AIPC). The development of photosensitization after addition of the dye into growth medium is ca. 8 times more rapid for AlPC than for AIPCS. Conversely, the loss of photosensitization when cells are incubated in a dye-free growth medium following a period of dye uptake, is also faster for AIPC. The dye uptake followed a kinetic behavior similar to the development of photosensitivity, but the loss of dye was too slow for both AlPC and AIPCS to explain loss of photosensitivity. When cells are incubated prior to illumination with AIPCS in phosphate buffered saline instead of growth medium, shorter time and smaller amount of dye are required to achieve the same level of photosensitization. The dependence of photosensitivity on dye concentration is linear for both AIPC and AIPCS. As already found for AIPC, photosensitization by the water-soluble derivative is also not enhanced in D₂O, suggesting that singlet oxygen is not involved in the cytotoxic response. Sodium salicylate, which was found to enhance the effect of AlPC was also effective with AIPCS. This effect is quite specific since the meta and para isomers had no effect. The metal atom complexed with the phthalocyanine ring is significant for the photobiological activity. Among the compounds tested, those containing Al or Zn are most active.     

Recovery of Chinese hamster cells following photosensitization by zinc tetrahydroxyphthalocyanine

The phthalocyanine dyes are attractive sensitizers for photodynamic therapy of cancer. The light fluence response curves for photocytotoxicity of zinc tetrahydroxyphthalocyanine were constructed using the colony-forming ability of Chinese hamster cells as an end-point. The survival curve of cells photosensitized to white light by this dye has a pronounced shoulder followed by an exponential decline. Postillumination hypertonic treatment (0.5 M NaCl for 20 min at 37 degrees C) enhanced log-phase killing, although to a lesser extent than after exposure to ionizing radiation. While such an enhancement usually indicates that the cells are able to repair potentially lethal damage, delayed trypsinization of photosensitized cells in plateau-phase failed to show a significant increase in cell survival. Thus, the repair of such a damage in plateau-phase is apparently absent. Experiments with split light fluence indicated that log-phase cells can repair sublethal damage during a 24 h interval, as evidenced by the reappearance of the shoulder on the split-dose survival curve.     

Resistance to induction of micronuclei,chromosomal aberrations and apoptosis by <Superscript>60</Superscript>Co gamma-ray in a cell strain M5, derived from Chinese hamster V79 cells

The paper aims to investigate cytogenetic and apoptotic responses of γ-irradiation in a radio-resistant cell strain designated as M5. Induced micronuclei, chromosomal aberrations, nuclear fragmentation and nucleosomal ladders by γ-irradiation were less at equal doses in M5 cells in comparison with that obtained in the parental Chinese hamster V79 cells. However, at equal survival, there were no differences in the end points studied. Results indicate that the residual damages that lead to reproductive cell death also resulted in the cytogenetic and apoptotic responses. We speculate that the repair efficiency in M5 cells was more efficient and increased DNA repair could be the cause of radiation resistance observed in M5 cells.     

Lethal Effect Induced in Pseudomonas aeruglnosa Exposed to Ultraviolet-A Radiation

Ultraviolet-A (365 nm, 120 kJ/m²/h) exposure caused cell death in Pseudomonas aeruginosa at doses at which Escherichia coli cell viability was not affected. We have not found that UVA induced growth delay or any other sublethal effect. Irradiated suspensions of P. aeruginosa showed a marked reduction in membrane-bound succinate dehydrogenase (SDH) and lactate dehydrogenase (LDH) activities. Succinate-driven respiration and several nutrient transport systems were also inhibited. Whereas SDH and LDH activities were independent of the irradiation conditions, cell viability, respiration and transport systems were protected when irradiation was performed in an N₂ atmosphere. A similar protective effect was observed when cells were grown in media containing glycerol or when preirradiation bacterial growth was carried out at 30°C (instead of 37°C). Results suggest that UVA induces a differential damaging effect on several biochemical functions of P. aeruginosa. The UVA induced photodamage may fall into two categories: indirect damage mediated by oxygen (cell killing and inhibition of respiration and transport systems) and direct damage to SDH and LDH (apparently not oxygen dependent). These enzymes and leucine transport appear not to be involved in the lethal effect described herein because they were altered despite viability-preserving conditions.     

量子电动学诱导电磁辐射研究纳米催化剂

Gold has been regarded as a poor heterogeneous catalyst because it is generally considered a nonreactive metal. But as nanocatalysts,gold and other metals somehow significantly enhance reactivity. It is generally thought chemical bonds of reactants are weakened by adsorption to nanocatalysts thereby allowing reactions to proceed more rapidly,but how this reaction proceeds to completion is not well understood. Here gold nanocatalysts are treated as unsupported nanoparticles (NPs) in a solution of reactant molecules from which extensions are made to gold NPs supported on titanium dioxide. Whether the NPs are supported or unsupported,enhanced catalytic reactivity depends on absorbed thermal kT (k is Boltzmann's constant and T is absolute temperature) energy accumulated from prior collisions of reactant molecules. The accumulated kT energy is treated as electromagnetic thereby allowing frequency up-conversion by quantum electrodynamics (QED) to the confinement frequency of the NP,typically beyond the vacuum ultraviolet (VUV). By this theory,the chemical reaction of reactant molecules having bonds weakened by adsorption is completed by QED induced VUV photolysis.     

Separation of flavins and nicotinamide cofactors in Chinese hamster ovary cells by capillary electrophoresis

Simultaneous extraction, separation and quantitation of reduced nicotinamide adenine dinucleotide (NADH), reduced nicotinamide adenine dinucleotide phosphate (NADPH), flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) in Chinese Hamster Ovary (CHO) cells were investigated. The separation of flavins and nicotinamide cofactors was performed by capillary electrophoresis with laser-induced fluorescence detection at the excitation wavelength of 325 nm. The separation protocol was established by investigating the excitation wavelength, high voltage and effects of buffer nature, pH and concentration. All endogenous fluorophores riboflavin, FAD, FMN, NADH and NADPH show wide linear range of quantitation. The limits of detection for the five compounds ranged from 4.5 to 23 nM. Extraction conditions were optimized for high-efficiency recovery of all endogenous fluorophores from CHO cells. To account for the complex matrix of cell extracts, a standard addition method was used to quantify FAD, FMN, NADH and NADPH in CHO cells. The quantitative results should be useful to reveal the metabolic status of cells. The protocols for extraction, separation and quantitation are readily adaptable to normal and cancer cell lines for the analysis of endogenous fluorophores.     

Structural and Functional Changes in Catalase Induced by Near-UV Radiation

Part one of this study shows that exposure of purified beef liver catalase in buffered solutions to BL lamps that provide a mixture of 99% UVA and 1% UVB (to be labeled UV_A) alters its chemistry and enzymatic activity. Thus, its spectral absorbance lost detail, it aggregated and exhibited a lower isoelectric point and its enzymatic activity was substantially reduced. These photochemically induced changes were increased by irradiation in phosphate buffer or in physiological medium (minimal essential medium) containing riboflavin and tryptophan. Neither α-tocopherol nor de-feroxamine were protective against these UV_A-induced changes in pure catalase. We further investigated the effect of UV_A radiation on the activity of catalase in cultured lens epithelial cells and the protective effects of antioxidants. Cultured lens epithelial cells of rabbits and squirrels were exposed to near-UV radiation with representation in the UVA region of 99% and 1% UVB. Catalase assays were done on ho-mogenate supernatants of cells kept dark or UV exposed. In some instances, cells were cultured in medium containing a-tocopherol or deferoxamine prior to UV radiation. Comparisons were made between UV-exposed lens cell catalase activity when exposure was done with or without the antioxidants. The UV_A radiation was strongly inhibitory to both rabbit and squirrel lens epithelial cell catalase activities. The range of fluxes of near UV radiation was compatible with that which could reach the lens from the sunlit environment. Catalase inactivation was lessened in cells preincubated with a-tocopherol and deferoxamine. This suggests that both singlet oxygen and hydroxyl radical formation may be involved in near-UV damage to lens epithelial cell catalase. Such inhibition of catalase by near-UV would enhance H₂O₂ toxicity and stimulate SH oxidation so as to damage the lens.     

Chinese hamster ovary cells sensitivity to localized electrical stresses

Application of an external electric field on a cell suspension induces an alteration in the membrane structure giving free access to the cell cytoplasm. Under mild pulsation conditions, permeabilization is a reversible process which weakly affects cell viability while drastic electrical conditions lead to cell death. The field pulse must be considered as a complex stress applied on the cell assembly. This study is a systematic investigation of the stress effects of field strength, pulse duration and number of pulses, at given joule energy. The loss in cell viability is not related to the energy delivered to the system. At a given joule energy, a strong field during a short cumulated pulse duration affects more viability than using a weak field associated with a long cumulated pulsation. At a given field strength and for a given cumulated pulse duration an accumulation of short pulses is also observed to be very damaging for cells. A control by the delay between the pulses suggests a memory effect. The field effect appears also to be vectorial in line with the known asymmetry of the membrane organization. These results suggest that processes at a cellular level are involved, either an activation of cell death or damage in cellular functions.     

Repair replication in Chinese hamster cells after damage from ultraviolet light

Abstract— After irradiation with u.v. light, Chinese hamster cells perform repair replication of their DNA. Small numbers of nucleosides are inserted into DNA, such that when BrUdR is used there is no detectable change in density. Repair replication begins immediately after irradiation, but it decelerates steadily and at least half is complete within 4 hr. Repair replication saturates above 200 ergs/mm² at a level which represents 0.055 per cent replacement of all thymine sites in 4 hr. Repair replication in mammalian cells, in contrast to that in microorganisms, does not appear to replace pyrimidine dimers excised from DNA in acid soluble form, and neither repair nor semiconservative replication discriminates between BrUdR and TdR.     

Kale extract increases glutathione levels in V79 cells, but does not protect them against acute toxicity induced by hydrogen peroxide

This study aims to evaluate the antioxidant potential of extracts of Brassica oleracea L. var. acephala DC. (kale) and several materials of Pieris brassicae L., a common pest of Brassica cultures using a cellular model with hamster lung fibroblast (V79 cells) under quiescent conditions and subjected to H?O? induced oxidative stress. Cytotoxicity was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and glutathione was determined by the 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB)-oxidized glutathione (GSSG) reductase recycling assay. The phenolic composition of the extracts was also established by HPLC-DAD. They presented acylated and non acylated flavonoid glycosides, some of them sulfated, and hydroxycinnamic acyl gentiobiosides. All extracts were cytotoxic by themselves at high concentrations and failed to protect V79 cells against H?O? acute toxicity. No relationship between phenolic composition and cytotoxicity of the extracts was found. Rather, a significant increase in glutathione was observed in cells exposed to kale extract, which contained the highest amount and variety of flavonoids. It can be concluded that although flavonoids-rich extracts have the ability to increase cellular antioxidant defenses, the use of extracts of kale and P. brassicae materials by pharmaceutical or food industries, may constitute an insult to health, especially to debilitated individuals, if high doses are consumed.     

Effects of BAPTA-AM and Forskolin on Apoptosis and Cytochrome c Release in Photosensitized Chinese Hamster V79 Cells

The mechanism of caspase-3-dependent apoptosis induced by photodynamic therapy (PDT) of cultured Chinese hamster V79 cells with pheophorbide a (PPa) was investigated. The PPa-PDT induced rapid apoptosis within 30 min after irradiation of cells. This apoptosis was inhibited by the 1O2 quencher N3- and caspase-3 inhibitor acetyl-Asp-Glu-Val-Asp-aldehyde, suggesting that 1O2 activated caspase-3 and then caused apoptosis. The intracellular calcium [Ca2+]i chelator (acetoxymethyl)-1,2-bis(o-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA-AM) and the cyclic adenosine monophosphate (cAMP)-increasing agent forskolin also inhibited not only the PPa-PDT-induced activation of caspase-3 but also apoptosis in V79 cells. Furthermore, PPa-PDT-induced cytochrome c release from mitochondria was found to be inhibited by the treatment with BAPTA-AM but not forskolin. These results indicated that [Ca2+]i and cAMP independently serve as regulators for PPa-PDT-induced apoptosis in the upstream of caspase-3.     

紫外辐射诱导C60聚合的扫描隧道显微研究

C60的聚合反应是近年来C60研究的热点．Yeretzian等[1]首先在质谱中观察到了C60在激光作用下的聚合反应，我们也曾以激光真空溅射技术制备了丰富的C60与C70的聚合物[2]．进一步研究还发现，C60在紫外光照射下也能聚合成膜[3]．为更深入地了解C60的聚合过程，本文以扫描隧道显微（STM）技术跟踪观察了C60在紫外光辐射下所发生的形态变化．1实验部分C60吸附在Au（111）面上时能够铺展成单分子层[4]．因此，我们选择Au（111）面作基底，将C60配成一定浓度的甲苯溶液滴在金表面上，待甲苯挥发后制得C60的单分子层．实验所用的紫外光源为…