Genotoxic and mutagenic effects of erythrosine B,a xanthene food dye,on HepG2 cells

Erythrosine (ErB) is a xanthene and an US Food and Drug Administration approved dye used in foods, drugs and cosmetics. Although its utilization is permitted, ErB is described as inhibitor of enzymes and protein–protein interactions and is toxic to pituitary and spermatogenesis processes. However, the genotoxicity and mutagenicity of ErB is inconclusive in the literature. This study aimed to analyze the genotoxicity of this dye using the alkaline comet assay and is the first investigation to evaluate ErB mutagenicity using the cytokinesis block micronucleus cytome (CBMN-Cyt) assay in HepG2 cells. These cells were chosen because they produce phase I and phase II enzymes that can mimic in vivo metabolism. The cells were treated with seven concentrations (0.1–70.0 μg mL⁻¹) of ErB, and the results showed genotoxicity at the two highest concentrations and mutagenicity at six concentrations. Furthermore, as micronuclei result from clastogenic and aneugenic processes, while comet assay is often considered more sensitive and detects DNA single strain breaks, we suggest that an aneugenic is responsible for the observed damage. Although ErB is approved for use in the food, cosmetic and pharmaceutical industries, it must be used carefully because it damages the DNA structure.     

Effects of orally administered antioxidants on micronuclei and sister chromatid exchange frequency in workers professionally exposed to antineoplastic agents

The widespread use of antineoplastic drugs in cancer treatment increased concern about possible hazard to workers involved in the preparation and administration of these drugs. In the present study, the effects of commercial antioxidative drug Oligogal Se® on genome protection were analyzed in 15 nurses handling the antineoplastic drugs at the Oncology Department in comparison to twenty healthy volunteers. The nurses took antioxidant mixture Oligogal Se®, consisting of vitamins C, E, A and selenium, one capsule per day, over a period of 6 months. Genome damage was measured in peripheral blood lymphocytes by usage of sister chromatid exchange test and the cytokinesis-block micronuclei test. The frequency of sister chromatid exchange (SCE) and micronuclei (MN) in the exposed group was significantly higher when compared to the control group (SCE, p < 0.05; MN, p < 0.01 respectively). After antioxidant supplementation, the frequency of sister chromatid exchange and micronuclei decreased (p < 0.05) when compared with the values from the beginning of the study, but were still above the values of the control group. The effects of confounding factors such as cigarette smoking and cytostatics exposure time were also evaluated. The data indicated that Oligogal Se® contributed to the decreasing of genome damages in workers handling the cytostatics.     

Lack of cytotoxic and genotoxic effects of Minthostachys verticillata essential oil: Studies in vitro and in vivo

Minthostachys verticillata (peperina) is an aromatic and medicinal plant with several uses and ethnobotanical properties. Numerous studies have demonstrated that its essential oil (Mv-EO) presents antimicrobial capacity and shows immunomodulating and anti-allergic properties in human cell lines. Thus, the goal of this study was to investigate the main chemical composition, analyzed by GC–FID, and the cyto-genotoxic effects of Mv-EO, using Vero cells, human PBMCs and mice bone marrow cells. The Mv-EO was rich in pulegone 60.5% and menthone 18.2%. Our results clearly show that Mv-EO is not cyto-genotoxic in vitro nor in vivo. It not induced cytotoxic effects, as indicated by trypan blue dye exclusion and NRU assays both in Vero cells and human PBMCs. In addition, Mv-EO (100–1000 μg/mL) not induced apoptotic effects on human PBMCs, as indicated by Hoechst staining and DNA fragmentation analysis by agarose gel electrophoresis. The in vivo assay showed that Mv-EO (25–500 mg/kg) not increased the frequency of micronucleus in bone marrow cells of mice. Further, the ratio of polychromatic/normochromatic erythrocytes was not modified. These findings suggest that Mv-EO appears to be safe as a therapeutic agent.     

Protective effect of bixin on cisplatin-induced genotoxicity in PC12 cells

Bixin is the main carotenoid found in annatto seeds (Bixa orellana L.) and is responsible for their reddish-orange color. The antioxidant properties of this compound are associated with its ability to scavenge free radicals, which may reduce damage and protect tissues against toxicity caused by anticancer drugs such as cisplatin. In this study, the genotoxicity and antigenotoxicity of bixin on cisplatin-induced toxicity in PC12 cells was assessed. Cytotoxicity was evaluated using the MTT assay, mutagenicity, genotoxicity, and protective effect of bixin were evaluated using the micronucleus test and comet assay. PC12 cells were treated with bixin (0.05, 0.08, and 0.10μg/mL), cisplatin (0.1μg/mL) or a combination of both bixin and cisplatin. Bixin was neither cytotoxic nor genotoxic compared to the controls. In the combined treatment bixin significantly reduced the percentage of DNA in tail and the frequency of micronuclei induced by cisplatin. This result suggests that bixin can function as a protective agent, reducing cisplatin-induced DNA damage in PC12 cells, and it is possible that this protection could also extend to neuronal cells. Further studies are being conducted to better understand the mechanisms involved in the activity of this protective agent prior to using it therapeutically.     

In vitro micronucleus screening of pharmaceutical candidates by flow cytometry in Chinese hamster V79 cells

We previously reported a high concordance of in vitro micronucleus (MNvit) results obtained by flow cytometry to the known cytogenetic activity often commercially available compounds mentioned as validation compounds in an early draft of the OECD MNvit TG487 [Bryce et al., 2010; Organization for Economic Co-operation and Development(OECD), 2007]. The current study investigated this method in Chinese hamster V79 cells with pharmaceutical compounds of unknown genotoxic potential. Twenty-five compounds from several therapeutic areas such as oncology, neuroscience and immunological research were tested in the flow cytometry assay, and for comparison using the cytokinesis-block microscopy assay. Five of these 25 compounds were considered positive for micronucleus induction by the microscopy assessment. In all cases, the results from the flow cytometry assess ment matched the results of the microscopy assay. Thus, flow cytometry is a viable method for assessing the aneugenic/clastogenic potential of pharmaceutical drug candidates. The flow method offered several advantages over traditional microscopy. For instance, the ratio of micronuclei (MN) to 10,000 nuclei was evaluated in less than 2 min vs.15 min to manually assess 600 binucleate cells. Evaluation by flow cytometry can be automated,freeing resources and eliminating scorer fatigue.The assay may also provide for mechanistic understanding of MN formation based on size and the ratio of nuclei with sub-2N DNA content, allowing for discrimination between aneugenic and clastogenic compounds.     

迷迭香酸对γ射线致小鼠骨髓嗜多染红细胞微核的保护作用

目的研究迷迭香酸对γ射线致小鼠骨髓嗜多染红细胞微核的保护作用。方法以骨髓嗜多染红细胞微核形成率为观测指标,C57BL/6J小鼠经60Coγ射线1～3 Gy照射24 h后观察骨髓嗜多染红细胞微核发生率的变化以选择合适的照射剂量;C57BL/6J小鼠经60Coγ射线2Gy照射后不同时间观察骨髓嗜多染红细胞微核发生率的变化以选择照射后取骨髓的时间;2 Gy照射前经迷迭香酸处理的C57BL/6J小鼠照后24 h观察骨髓嗜多染红细胞微核发生率的变化,以确定迷迭香酸对小鼠微核保护作用的剂量效应与时间效应。结果 2 Gy照射24 h后小鼠骨髓嗜多染红细胞微核发生率增加比较明显,适合作为小剂量照射条件;照射后24 h或48 h骨髓嗜多染红细胞微核发生率增加比其他时间明显;经迷迭香酸100 mg/kg连续7次处理后的受照射小鼠骨髓细胞中,微核发生率（6.50‰）明显低于单纯照射组（23.65‰）。结论迷迭香酸对γ射线致骨髓嗜多染红细胞微核具有保护作用。     

铜陵市150例放射人员淋巴细胞微核及染色体畸变情况调查

目的探讨放射工作人员外周血淋巴细胞微核和染色体畸变的变化情况,为当前放射人员的卫生防护工作提供依据。方法用常规方法检测150名放射工作人员的外周血淋巴细胞染色体畸变和微核情况并分析。结果医务组与工业组微核阳性检出率和染色体畸变阳性检出率分别为8.5%、2.9%和4.5%、2.9%,随工龄的增加而增高。结论低剂量电离辐射对放射工作人员遗传物质有一定影响,不同暴露条件下的机体的损伤程度有所不同。     

Mutagenicity studies with N-acetyl-l-aspartic acid

Analytical studies have reported that N-acetyl-l-aspartic acid (NAA) is present at low concentrations in many foods. The current studies were conducted to assess the mutagenicity of NAA using standard OECD guideline in vitro bacterial and in vivo mammalian mutagenicity studies. For comparison and control data, mutagenicity studies were also conducted with its constituent amino acid l-aspartate (ASP) because NAA is metabolized to ASP. The combination of an in vitro method for assessing point mutations in bacteria and an in vivo method to assess clastogenicity in an animal model provided adequate evidence for mutagenicity hazard assessment of NAA. No evidence of mutagenicity was observed in either test system with either NAA or ASP. The results from the current studies demonstrate that the presence of NAA in foods is not likely to represent a risk for mutagenicity.     

中子辐射诱发人外周血染色体畸变、微核HPRT基因突变的比较研究

目的研究中子辐射诱发离体人外周血淋巴细胞染色体畸变率、微核率和HPRT基因突变率作为中子辐射的生物剂量计的可能性。方法采用14MeV中子等剂量率照射离体人外周血,吸收剂量范围0~1Gy,秋水仙素阻断法检测染色体畸变率,CB细胞法和常规培养法检测微核率,多核细胞法检测HPRT基因突变率,建立相应的剂量效应曲线,比较各指标间的相关性。结果在该剂量范围内,各指标与剂量关系符合现行平方模型,染色体畸变、微核呈过离散分布,三者有线性相关性。染色体畸变率灵敏度最高,其次是CB细胞法微核率,再次是常规培养法微核率,HPRT基因突变率最低。结论中子辐射事故可采用准确灵敏的染色体畸变率和CB细胞法微核率作为生物剂量计进行辐射种类判定和生物剂量估算。     

宇宙辐射对民航飞行人员淋巴细胞微核率的影响

目的探讨宇宙辐射对民航飞行人员细胞遗传学指标的影响,为辐射防护提供依据。方法用胞质分裂阻滞微核法观察宇宙辐射对外周血淋巴细胞微核率的影响。采集150名健康飞行人员和32名地面对照人员的外周静脉血,在含植物血凝素的RPMI1640培养基中37℃培养38～44h,加入细胞松弛素-B继续培养至72h。经低渗、固定、涂片、染色,每例观察500个双核淋巴细胞中的微核。结果（1）飞行人员双核淋巴细胞微核率（MNF）（16．15±0．33）‰、微核细胞率（MNCF）（13．66±0．21）‰与地面人员MNF（11．48±0．50）‰、MNCF（10．42±0．57）‰相比,差异有显著性（P〈0．01）;（2）MNF、MNCF与飞行人员的工龄、累积飞行小时和年龄呈反相变化：（3）飞行人员经1个月的休假后,MNF、MNCF均低于连续飞行组,与连续飞行组比较,两组间差异有显著性（P〈0．05）,但近1个月未飞行组的MNF、MNCF仍显著高于地面组;（4）在去除年龄、性别等影响因素后,未发现吸烟增加飞行人员的微核;（5）无论飞行组还是地面组,男女性别间微核差异无显著性（P〉0．05）。结论宇宙辐射能引起民航飞行人员遗传学指标变化;胞质分裂阻滞微核法能够灵敏地反映近期内飞行负荷的差别,可用于监测辐射损伤。