Functional genomics of UV radiation responses in human cells

Functional foods are "foods and beverages with claimed health benefits based on scientific evidence". Health claims need to be substantiated scientifically. The future of functional foods will heavily rely on proven efficacy in well-controlled intervention studies with human volunteers. In order to have the maximum output of human trials, improvements are needed with respect to study design and optimization of study protocols. Efficacy at realistic intake levels needs to be established in studies with humans via the use of suitable biomarkers, unless the endpoint can be measured directly. The human body is able to deal with chemical entities irrespective of their origin, and the pharmaceutical terms "absorption, distribution, metabolism and excretion" have their equivalent when biomarkers are concerned. Whereas only "diurnal variation" or "circadian rhythm" is sometimes considered, little attention is paid to "kinetics of biomarkers". "Kinetics of biomarkers" comprises "formation, distribution, metabolism and excretion". However, this is at present neither an established science nor common practice in nutrition research on functional foods. As a consequence, sampling times and matrices, for example, are chosen on the basis of historical practice and convenience (for volunteers and scientists) but not on the basis of in depth insight. The concept of kinetics of biomarkers is illustrated by a variety of readily comprehensible examples, such as malaria, cholesterol, polyphenols, glutathione-S-transferase alpha, F2-isoprostanes, interleukin-6, and plasma triacylglycerides.     

Mutagenic action of heavy ions on Escherichia coli cells

Induction of direct mutations in the lactose operon of E. coli cells by gamma-radiation and accelerated heavy ions with different LET was studied. The experiments were performed with the wild-type PolA and LexA strains. A quadratic dependence of the mutation rate on the dose of different radiations for the wild-type strain and the PolA mutant was observed. However, different types of radiation showed different relative genetic effectivenesses (RGE). The dependence of RGE on LET for the wild-type and PolA strain has a maximum. A LexA strain showed much reduced mutation rates and a linear dose response. The RGE decreased with increasing LET of ionizing radiation.     

Mutagenic adaptive response to high-LET radiation in human lymphoblastoid cells exposed to X-rays

The ability of cells to adapt low-dose or low-dose rate radiation is well known. High-LET radiation has unique characteristics, and the data concerning low doses effects and high-LET radiation remain fragmented. In this study, we assessed in vitro the ability of low doses of X-rays to induce an adaptive response (AR) to a subsequent challenging dose of heavy-ion radiation. Lymphoblastoid cells (TK6, AHH-1, NH32) were exposed to priming 0.02-0.1Gy X-rays, followed 6h later by challenging 1Gy heavy-ion radiation (carbon-ion: 20 and 40keV/μm, neon-ion: 150keV/μm). Pre-exposure of p53-competent cells resulted in decreased mutation frequencies at hypoxanthine-guanine phosphoribosyl transferase locus and different H2AX phosphorylation kinetics, as compared to cells exposed to challenging radiation alone. This phenomenon did not seem to be linked with cell cycle effects or radiation-induced apoptosis. Taken together, our results suggested the existence of an AR to mutagenic effects of heavy-ion radiation in lymphoblastoid cells and the involvement of double-strand break repair mechanisms.     

Mutagenic action of cadmium on the sex cells of male mice

Possible mutagenic effect of cadmium chloride was studied by determining the frequency of dominant lethal mutations induced in germ cells of male mice. Water solution of CdCl2 was injected intraperitoneally to male mice at doses of 1.0, 2.0 and 4.0 mg/kg. The results obtained did not reveal any mutagenic effect of this compound. The dose of 4.0 mg/kg CdCl2 resulted in the death of spermatocytes and spermatogonia and the sterility of male mice. Cadmium chloride at a dose of 2.0 mg/kg did not affect the frequency of dominant lethal mutation induced by gamma-rays 60Co at a dose of 450 r in germ cells of male mice.     

Mutagenic adaptive response to high-LET radiation in human lymphoblastoid cells exposed to low doses of heavy-ion radiation

Adaptive response (AR) and bystander effect are two important phenomena involved in biological responses to low doses of ionizing radiation (IR). Furthermore, there is a strong interest in better understanding the biological effects of high-LET radiation. We previously demonstrated the ability of low doses of X-rays to induce an AR to challenging heavy-ion radiation [8]. In this study, we assessed in vitro the ability of priming low doses (0.01Gy) of heavy-ion radiation to induce a similar AR to a subsequent challenging dose (1-4Gy) of high-LET IR (carbon-ion: 20 and 40keV/μm, neon-ion: 150keV/μm) in TK6, AHH-1 and NH32 cells. Our results showed that low doses of high-LET radiation can induce an AR characterized by lower mutation frequencies at hypoxanthine-guanine phosphoribosyl transferase locus and faster DNA repair kinetics, in cells expressing p53.     

The effect of monochromatic radiation in the range 350 to 750 nm on the carotenogenesis in Verticillium agaricinum

An action spectrum for carotenogenesis in V. agaricinum has maxima at 395, 433, 660 and 737 nm. In a previous study it had been shown that a light-minus-dark difference spectrum of a crude extract from V. agaricinum had maxima at 390 and 420 nm, and furthermore a red, far-red interaction suggesting phytochrome involvement has been proposed. All these data suggest that there may be at least two photoreceptor systems operating in the photoinduction process here; one for the near-ultraviolet (UV-A)-mediated carotenogenesis, presumably a novel pigment, and the other for the red, far-red region, most likely phytochrome.     

Mutagenic action of nitroso compounds on Escherichia coli cells]

An attempt to induce some forward and back mutations in two Escherichia coli strains (his- and HfrH requiring thiamine) under the action of the carcinogenic nitrosamines--dimethylnitrosamine (DMN) and diethylnitrosamine (DEN)--is described. For this purpose the cells of E. coli were treated with 5% DMN or 1% DEN for 1 hour at 37 degrees C in 0.14 M NaCl. It was shown that the sensitivity of both strains to both nitrose compounds was not the same. DEN was 5-fold as toxic as DMN for the E. coli cells. DMN and DEN induced neither mutations of resistance to 10(-3) M valine, nor reversions in histidine-dependent strain. These mutations were obtained after the cells were treated with 0.1 M NaNO2. Lethal effects of DMN increased more than in 5 times and the toxicity of DEN did not change in hydroxylating mixture, in which nitrosamines derived to active compounds. Under these conditions both carcinogenes showed a mutagenic activity. DEN proved to be about twice as strong mutagenically as DMN. Thus, in our experiments we could see that DMN and DEN could induce both forward and back mutations in E. coli.     

阳光紫外辐射对两种微藻类光化学效率的影响

近年来人类活动释放的含氟、氯等有机物对臭氧层造成破坏,使得到达地球表面的紫外线B（Ultra-violet B（UV—B）．280-315nm）辐射增强。为此,生物活动受到了多方面的影响。以往的研究发现,紫外辐射（Ultra-violet radiation．UVR）可以抑制藻类的生长及光合固碳和游动性（有鞭毛的种类）。并损伤细胞色素和遗传物质DNA。     

Mutagenic fingerprint of ozone in human cells

Ozone is an important factor in urban pollution and represents a major concern for human health. The chemical reactivity of ozone toward biological targets and particularly its genotoxicity supports a possible link between exposure and cancer risk, but no molecular data exist on its mutagenic potential in human cells. Using a shuttle vector, we showed that ozone is indeed a potent mutagen and we characterized the mutation spectrum it produced in human cells. Almost all mutations are base substitutions, essentially located at G:Cs (75%), typical of reactive oxygen species (ROS), but occurring in a specific pattern, i.e. a similar extent of GC:TA (28%), GC:CG (23%) and GC:AT (23%). The targeted distribution of mutations and identification of hotspot sequences define the first molecular fingerprint of mutations induced by ozone in human cells. Possible applications derived from our results with respect to ozone genotoxicity should help determining quantifiable biomarkers of ozone exposure in human health, especially for carcinogenesis.     

UV radiation induces delayed hyperrecombination associated with hypermutation in human cells

Ionizing radiation induces delayed genomic instability in human cells, including chromosomal abnormalities and hyperrecombination. Here, we investigate delayed genome instability of cells exposed to UV radiation. We examined homologous recombination-mediated reactivation of a green fluorescent protein (GFP) gene in p53-proficient human cells. We observed an approximately 5-fold enhancement of delayed hyperrecombination (DHR) among cells surviving a low dose of UV-C (5 J/m2), revealed as mixed GFP+/- colonies. UV-B did not induce DHR at an equitoxic (75 J/m2) dose or a higher dose (150 J/m2). UV is known to induce delayed hypermutation associated with increased oxidative stress. We found that hypoxanthine phosphoribosyltransferase (HPRT) mutation frequencies were approximately 5-fold higher in strains derived from GFP+/- (DHR) colonies than in strains in which recombination was directly induced by UV (GFP+ colonies). To determine whether hypermutation was directly caused by hyperrecombination, we analyzed hprt mutation spectra. Large-scale alterations reflecting large deletions and insertions were observed in 25% of GFP+ strains, and most mutants had a single change in HPRT. In striking contrast, all mutations arising in the hypermutable GFP+/- strains were small (1- to 2-base) changes, including substitutions, deletions, and insertions (reminiscent of mutagenesis from oxidative damage), and the majority were compound, with an average of four hprt mutations per mutant. The absence of large hprt deletions in DHR strains indicates that DHR does not cause hypermutation. We propose that UV-induced DHR and hypermutation result from a common source, namely, increased oxidative stress. These two forms of delayed genome instability may collaborate in skin cancer initiation and progression.     

Measurement of solar UV radiation in Antarctica with collagen sheets

Collagen sheets were used in a unique evaluation method to examine skin damage caused by ultraviolet (UV) light of short wavelength during a season of the Antarctic ozone hole. The collagen sheets were exposed outdoors for 25 and 50 d, in the spring when the ozone hole was formed and in the ozone-hole-free autumn. Extracts from the exposed collagen sheets were analyzed for total protein and terminal amino acid concentrations as an index of collagen fragmentation. The results show that the amount of extractable collagen and terminal amino acid concentration in the spring exposure were approximately double and five times higher, respectively, when compared with those in the autumn exposure. During the ozone hole occurrence, the terminal amino acid concentration of the extracted collagen was about five times higher when exposure lasted 50 d from mid-September to the end of October compared to when exposure lasted 25 d from mid-September to early October. This result could be attributed to a limited amount of short-wavelength UV radiation reaching the ground surface as a result of the low height of the sun in September, when the ozone hole occurred. In fact, UV radiation measurements taken at Syowa Station indicate that short-wavelength UV radiation in the range 290-295 nm was not detected until approximately 1-2 months after the beginning of the ozone hole occurrence.     

Inactivation of nuclear polyhedrosis virus (Baculovirus subgroup A) by monochromatic UV radiation

Monochromatic radiation at wavelengths of 290, 300, 310, and 320 nm inactivated occluded nuclear polyhedrosis virus of the Douglas-fir tussock moth, Orgyia pseudotsugata (McDunnough). Data indicate that all of the wavelengths are capable of causing virus inactivation; much greater fluences are needed for virus inactivation as the wavelength increases.     

Thickness of mammalian cells in culture, the absorption of UV radiation in the cells and the related shift in action spectra

The thickness of SPEV cells in monolayer with different cell densities was measured by means of 3-dimensional reconstruction from serial vertical sections. No significant changes in the mean cell thickness were detected despite the wide range of volume and cell density variations. UV absorption at different wave-lengths was measured in various sites of single cells. It is shown that the well known shift between peaks of the UV action spectrum for mammalian and bacteria cells may result from the cell self-shielding at short wavelengths.     

Mutagenic effect of cadmium on tetranucleotide repeats in human cells

Cadmium is a human carcinogen that affects cell proliferation, apoptosis and DNA repair processes that are all important to carcinogenesis. We previously demonstrated that cadmium inhibits DNA mismatch repair (MMR) in yeast cells and in human cell-free extracts (H.W. Jin, A.B. Clark, R.J.C. Slebos, H. Al-Refai, J.A. Taylor, T.A. Kunkel, M.A. Resnick, D.A. Gordenin, Cadmium is a mutagen that acts by inhibiting mismatch repair, Nat. Genet. 34 (3) (2003) 326–329), but cadmium also inhibits DNA excision repair. For this study, we selected a panel of three hypermutable tetranucleotide markers (MycL1, D7S1482 and DXS981) and studied their suitability as readout for the mutagenic effects of cadmium.

We used a clonal derivative of the human fibrosarcoma cell line HT1080 to assess mutation levels in microsatellites after cadmium and/or N-methyl-N-nitro-N-nitrosoguanidine (MNNG) exposure to study effects of cadmium in the presence or absence of base damage. Mutations were measured in clonally expanded cells obtained by limiting dilution after exposure to zero dose, 0.5 μM cadmium, 5 nM MNNG or a combination of 0.5 μM cadmium and 5 nM MNNG.

Exposure of HT1080-C1 to cadmium led to statistically significant increases in microsatellite mutations, either with or without concurrent exposure to MNNG. A majority of the observed mutant molecules involved 4-nucleotide shifts consistent with DNA slippage mutations that are normally repaired by MMR. These results provide evidence for the mutagenic effects of low, environmentally relevant levels of cadmium in intact human cells and suggest that inhibition of DNA repair is involved.     

The role of solar and UV radiation in environmental stress assessment

The purpose of this study was to evaluate the role of ultraviolet (UV) radiation in environmental assessment. Analysis of two meteorological databases, from Israel and New Zealand, used in this study revealed that the UV radiation component in the environmental stress assessment was not significant. However, because of the health hazards, an independent UV index should be implemented and used for preventing acute and chronic injuries. A prediction of UV radiation from solar radiation measurement would be a great benefit for such an index.     

Mutagenic action of the dimethyl ester of terephthalic acid on murine somatic cells in vivo

Mutagenic activity of dimethyl terephthalate (DMtP) was evaluated using the bone marrow micronucleus test in mice. Clear clastogenic effect with the highest response in 24 h after a single i.p. injection was obtained at all concentrations used (0.2-1.0 mM/kg). The time-course for the micronuclei induced by DMtP was in agreement with the literature data on fast excretion of phthalates from mammal body. The dose-response curve for DMtP-induced micronuclei was linear in form with the logarithmic component. The emergence of the latter was related to the elevation of the chemical's concentration to the level at which DMtP starts to exert toxic influence on bone marrow erythropoietic function. The comparison of the effect induced by DMtP with that of methyl nitrosourea indicated that DMtP could not be considered as a strong mutagenic compound. Susceptibility of the micronucleus test was compared with that of Drosophila dominant lethal test in terms of the concentrations at which equal clastogenic effect was seen. This comparison made it possible to conclude that the micronucleus test in mice was able to respond to much lower phthalate concentrations, as compared with the test in Drosophila. The results provided the evidence of capacity of dimethyl terephthalate to cause alterations of genetical structures in both somatic and germinal cells of two highly organized species in vivo.     

Lethal and mutational effects of solar and UV radiation on Staphylococcus aureus

Strains of Staphylococcus aureus, an opportunistic pathogen commonly found on human skin, were exposed to sunlight and UV C radiation, and the lethal and mutational effects measured. Sunlight killed cells with an inactivation constant of 3×10^-5 per joule per square metre; UV C was much more lethal, giving an inactivation constant of approximately 0.1 per joule per square metre. Some strains tested showed a sensitivity to sunlight that was dependent on the growth phase of the cells, exponentially growing cells showing a greater sensitivity. Mutational effects of irradiation were measured by the appearance of mutants sensitive to methicillin following irradiation of a multiresistant strain. Mutants appeared at a frequency of 10^-3; this high frequency of mutation in the region of the mec gene has also been observed when multiresistant strains are subjected to nutritional or thermal stress. Mutants showed the same chromosomal alteration (seen in pulse-field gel electrophoresis of Smal-digested DNA) whether induced by solar or UV C irradiation.     

铜绿微囊藻对紫外辐射的生理代谢响应

采用紫外(UV)滤膜过滤日光UV以及紫外灯添加UV的方法,研究了UV辐射对铜绿微囊藻Microcystis aeruginosa单细胞藻株PCC7806和群体藻XW01生长及生理代谢的影响。结果显示,在室内条件下低剂量UV辐射可促进群体微囊藻XW01生长;室外条件下与滤除了UV的光照相比,含有UV的完全日光更有利于微囊藻生长;而相同的UV辐射强度均导致单细胞株死亡,群体株显示了较强的UV抗性;日光中的UV可促进XW01合成抗氧化相关的超氧化物歧化酶(SOD)和过氧化氢酶(CAT)、促进胞外多糖的产生并形成较大的群体、促进UV屏障物质类菌孢素氨基酸(MAAs)和伪枝藻素(Scy)积累。这些生理代谢的改变,消除了阳光辐射中UV对微囊藻的伤害。研究的结果提示,自然条件下阳光中的UV有助于群体微囊藻生长。