Sensitivity to cisplatin-induced mutations and elevated chromosomal aberrations in lymphocytes from sickle cell disease patients

Sickle cell disease (SCD) is an inherited disorder caused by a single nucleotide substitution in the β-globin gene. The clinical heterogeneity observed in SCD patients has been attributed to environmental and genetic factors. The patients are subjected to increased oxidative stress, particularly during vaso-occlusive crises and acute chest pain. Another possible cause of oxidative stress in SCD is the high concentration of iron in the patients’ plasma. The increase in oxidative stress could be a relevant risk factor for mutagenesis and carcinogenesis. Studies on the frequency of basal chromosomal aberrations in cultured lymphocytes from SCD patients have not been reported so far. In order to contribute to the understanding of the role of the different biomarkers and their relationship with the extremely variable clinical manifestation of SCD, we investigated the frequency of chromosome damage in peripheral lymphocytes from sickle cells patients and healthy controls. We found an increased frequency of chromosome damage and percentage of aberrant metaphases in these patients when compared with control subjects, even at basal values (p < 0.05). In the cytogenetic sensitivity assay, the results showed that these patients presented a marked decrease in the mitotic index values compared with healthy controls. Cisplatin-induced chromosomal damage in lymphocytes from these patients was significantly higher than the frequency measured in healthy controls. The results obtained in the present study showed that more investigations are needed in order to elucidate the susceptibility to genomic instability of SCD patients.     

二氧化硫体内衍生物诱发中国仓鼠肺细胞微核的效应

目的 为了解二氧化硫（SO2）对哺乳类细胞的遗传毒理效应。方法 以SO2在体内的衍生物－－亚硫酸氢钠和亚硫酸钠处理培养的中国仓鼠肺成纤维细胞（CHL）,并对其诱发微核（MN）形成的作用进行了分析。结果 对照盐水组和1、2、5、10mmol/L的SO2衍生物处理组CHL的MN率（％）分别为8.17、9.83、10.33、12.83、13.33及14.17,从2mmol／L的SO2衍生物处理组开始 与对照组差异有显著性,且呈剂量依赖关系。无论是否有S9混合物存在,SO2衍生物均可诱发CHL的MN率显著增高,且SO2衍生物处理细胞时间愈长,引起细胞遗传损伤所需的最低浓度就愈底。结论 SO2 是不需要体内酶促转化的直接的细胞染色体断裂剂和基因毒性因子。同时也启示,长期接触环境低浓度SO2污染,有引起人体细胞遗传物质损伤的潜在危险。     

Role of ascorbic acid on mercuric chloride-induced genotoxicity in human blood cultures

Efforts are made to find therapeutic agents capable of minimizing genotoxicity of various natural and man-made compounds. The genotoxicity induced by mercury compounds remains controversial. Therefore we have investigated the genotoxic effect of mercuric chloride (MC; HgCl₂) at three concentrations (1.052, 5.262 and 10.524 μ ) and role of ascorbic acid (vitamin C) at a concentration of 9.734 μ on MC-treated short-term human leucocyte cultures. We assessed the proliferative rate index (PRI), sister chromatid exchange (SCE) and chromosomal aberrations (CAS) in control and MC-treated cultures with and without vitamin C supplementation. The results showed that MC has no effect on cell-cycle kinetics, but the frequency of SCE/cell was significantly higher in a dose-dependent manner than control values. HgCl₂ also significantly induced C-anaphases (abnormal mitosis) in blood cultures. These effects were prevented by the addition of vitamin C to MC-treated cultures. The data indicate the mutagenic activity of MC and the protective role of vitamin C on mercury-induced genotoxicity in human blood cultures is probably due to its strong antioxidant and nucleophilic nature.     

生物可吸收性涂层补片的细胞毒性和遗传毒性研究

生物可吸收性涂层补片在应用于临床前必须进行一系列生物学评价以保证其安全性,细胞毒性和遗传毒性研究是其体外筛选部分。细胞毒性研究以补片浸提液接触受试细胞,观察细胞的毒性反应。生物可吸收性涂层补片浸提液1.5 cm2/ml时,细胞毒性反应1级;生物可吸收性涂层补片浸提液3 cm2/ml时,细胞毒性反应2级。遗传毒性研究包括两个试验,细菌回复突变试验结果表明生物可吸收性涂层补片的浸提液对鼠伤寒沙门氏菌受试株无致突变性;体外哺乳动物细胞染色体畸变试验中,任一处理组的细胞生长率均与阴性对照处于同一水平,表明没有诱导哺乳动物体细胞产生染色体畸变。     

氧化型染发剂DNA交联作用的研究

采用溴乙锭荧光法测定氧化型染发剂的主要成分对苯二胺（ＰＰＤ）、过氧化氢（Ｈ２Ｏ２）和ＰＰＤ－Ｈ２Ｏ２混合物对小牛胸腺ＤＮＡ的双链间交联形成作用。结果表明：用０～６％的对苯二胺及０～３％的Ｈ２Ｏ２对小牛胸腺ＤＮＡ直接染毒，３７℃３０ｍｉｎ时无双链间ＤＮＡ交链的形成；而ＰＰＤ和Ｈ２Ｏ２发生氧化反应后则具有双链ＤＮＡ交联形成作用并具有剂量反应关系（ｒ＝０．８２９，Ｐ＜０．０１），在受试剂量为０～０．８％（以对苯二胺含量计）时，氧化型对苯二胺的ＤＮＡ交联作用增加了４．２倍。研究结果提示，ＤＮＡ交联作用可能是氧化型染发剂遗传毒作用过程中ＤＮＡ损伤的一种方式。     

Biological activity of N-nitrosodiethanolamine and of potential metabolites which may arise after activation by alcohol dehydrogenase in Salmonella typhimurium,in mammalian cells,and in vivo

Summary The potent carcinogen N-nitrosodiethanolamine (NDELA) which is nonmutagenic in standard modifications of the S. typhimurium/mammalian microsome assay, can be activated effectively by alcohol dehydrogenase/NAD (ADH/NAD) to intermediates which are directly mutagenic in strains TA 98 and TA 100. The expected metabolites N-nitroso-2-hydroxymorpholine (NHMor), N-nitroso-(2-hydroxyethyl)-glycine (NHEG), N-nitrosoiminodiacetic acid (NIDA), and glycolaldehyde were assayed for their direct mutagenic activities in S. typhimurium TA 1535, TA 98, and TA 100. All compounds were clearly mutagenic in TA 100, but different specifities were observed for the other strains. NDELA and its putative mutagenic metabolites were also tested for induction of genotoxic activities by determination of DNA single strand breaks in primary rat hepatocytes. In these cells, NDELA and NHMor were clearly genotoxic, whereas NHEG and NIDA were inactive. In contrast, when assayed for the induction of selective DNA amplification NDELA and its metabolites were not found to induce SV40 DNA synthesis in SV40-transformed Chinese Hamster cells. The compounds were also assayed for induction of DNA single strand breaks in the liver after a single oral application to rats. NDELA and NHMor were about equally active in this in vivo test, whereas NHEG, NIDA and glycolaldehyde were inactive. Differences in biological activity in the cultivated cells, as compared to hepatocytes or to the in vivo situation may most probably be due to differences in metabolism and/or pharmacokinetics.Dedicated to Professor Dietrich Schmähl on the occasion of his 60th birthday     

Micronucleus induction in cells co-exposed in vitro to 50 Hz magnetic field and benzene, 1,4-benzenediol (hydroquinone) or 1,2,4-benzenetriol

The generation, transmission (e.g. power lines, transformers, service wires, and electrical panels), and use (e.g. home appliances, such as electric blankets, shavers, and televisions) of electrical energy is associated with the production of weak electric and magnetic fields (EMF) which oscillate 50 (Europe) or 60 (USA) times per second (power-line frequency), falling in the extremely-low frequency (ELF) region of the electromagnetic spectrum. Epidemiological reports suggest a possible association between exposure to ELF-EMF and an increased risk of cancer (e.g. childhood acute leukaemia). Benzene is an established human leukomogen. This xenobiotic, which is unlikely to be the ultimate carcinogen, is metabolized in the liver to its primary metabolite phenol, which is hydroxylated to hydroquinone (1,4-benzenediol) and 1,2,4-benzenetriol. In this in vitro approach, to test the genotoxic and / or co-genotoxic potency of ELF-EMF, the cytokinesis block micronucleus (MN) method with Jurkat cells has been used. A 50 Hz magnetic field (MF) of 5 mT field strength was applied for different length of time (from 1 to 24 h), either alone or with benzene, 1,4-benzenediol, or 1,2,4-benzenetriol. Our preliminary results show that, after 24 h exposure, the frequency of micronucleated cells in MF-exposed cultures is 1.9 fold higher than in sham-exposed (control) cultures. Benzene exposure does not show any cytogenetic activity, whereas 1,4-benzenediol or 1,2,4-benzenetriol alone significantly affect the number of MN in Jurkat cells, as compared to untreated cultures. Moreover, co-exposure to ELF-MF does not seem to affect the frequency of micronuclei induced by benzene, 1,4-benzenediol, or 1,2,4-benzenetriol.     

Passive smoking at work: biochemical and biological measures of exposure to environmental tobacco smoke

Summary Several biochemical and biological measures of tobacco smoke intake were used to evaluate exposure of restaurant personnel to environmental tobacco smoke as compared with active smokers and non-exposed non-smokers. All of the measured parameters — carboxyhaemoglobin (COHb), thiocyanate (SCN) and cotinine in plasma, cotinine and mutagenicity in urine, total white blood cell count (WBC), and sister chromatid exchange (SCE) frequency in cultured lymphocytes — were significantly elevated in the smoker group (n = 22) compared to the non-exposed group (n = 20). Work-related passive exposure (n = 27) was seen most clearly in the cotinine values, both from plasma (mean P-cot in passive smokers 10 ng/ml vs 5.2 ng/ml in non-exposed) and from urine (mean U-cot in passive smokers 56 ng/ml vs 8.3 ng/ml in non-exposed), but significant increases were also seen in the thiocyanate levels (mean P-SNC in passive smokers 58 mol/1 vs 46 mol/1 in non-exposed) and, as a preliminary finding, in total leucocyte count (in passive smokers 8.0 × 10⁹/1 vs 6.8 x 10⁹/1 in non-exposed). The results demonstrate that environmental tobacco smoke may be an occupational health hazard.     

N-Nitrosodiethanolamine is activated in the rat to an ultimate genotoxic metabolite by sulfotransferase

Summary Inhibition of sulfotransferase by 2,6-dichloro-4-nitrophenol (DCNP) has been found to completely abolish the genotoxic potential of N-nitrosodiethanolamine (NDELA) in rat liver as indicated by induction of DNA single strand breaks. The DNA strand breaking potential of N-nitroso-2-hydroxymorpholine (NHMOR), a metabolite of NDELA formed by alcohol dehydrogenase-mediated oxidation, was also almost quantitatively abolished. In contrast to these -hydroxylated nitrosamines, the effectiveness of N-nitrosodiethylamine (NDEA) remained unaffected by DCNP with respect to its DNA damaging potential. N-Nitrosoethylethanolamine (NEELA) was the most potent genotoxic agent of this series of nitrosamines and its strand breaking activity was only partialy inhibited by DCNP. A new activation mechanism for NDELA is proposed: NDELA is transformed at first by alcohol dehydrogenase into the cyclic hemiacetal NHMOR. This cyclic -hydroxynitrosamine appears to be a substrate for sulfotransferase. The resulting sulfate conjugate is suggested to be the ultimate genotoxic electrophile. However, the results do not exclude the possiblity that NDELA itself undergoes sulfate conjugation.Dedicated to Professor Dietrich Schmähl on the occasion of his 60th birthday     

Black bean (Phaseolus vulgaris L.) as a protective agent against DNA damage in mice

This study was designed to evaluate the toxicogenetic or protective effect of cooked and dehydrated black beans (Phaseolus vulgaris L.) in bone marrow and peripheral blood cells of exposed mice. The frequency of micronuclei detected using the bone marrow erythrocyte micronucleus test and level of DNA lesions detected by the comet assay were chosen as end-points reflecting mutagenic and genotoxic damage, respectively. Initially, Swiss male mice were fed with a 20% black bean diet in order to detect mutagenic and genotoxic activity. However, no increase in the frequency of bone marrow micronucleated polychromatic erythrocytes (MN PCEs) or DNA lesion in leukocytes was observed. In contrast, received diets containing 1, 10 or 20% of black beans, a clear, but not dose-dependent reduction in the frequency of MN PCEs were observed in animals simultaneously treated with cyclophosphamide, an indirect acting mutagen. Similar results were observed in leukocytes by the comet assay. Commercial anthocyanin was also tested in an attempt to identify the bean components responsible for this protective effect. However, instead of being protective, the flavonoid, at the highest dose administered (50 mg/kg bw), induced primary DNA lesion, as detected by the comet assay. These data indicate the importance of food components in preventing genetic damage induced by chemical mutagens, and also reinforce the role of toxicogenetic techniques in protecting human health.