Evaluating genotoxicity associated with microcystin-LR and its risk to source water safety in Meiliang Bay, Taihu Lake

In recent years cyanobacteria blooms have become a severe problem in Taihu Lake, a large shallow eutrophic lake in China. Microcystins produced by certain genera of cyanobacteria can affect public health in this area because of their acute and chronic toxic effects. In this study, samples of cyanobacteria were collected and extracted by two solvent systems. The extracts were tested with three short-term genotoxicity assays, the ara test, the Ames test, and the SOS/umu test. In addition, temporal variation in the concentrations of microcystin-LR in the water samples was determined and monitored by an ELISA assay. Then the concentration of microcystin-LR in the drinking water was estimated. The risk of microcystin-LR exposure by drinking water was assessed according to tolerable daily intake (TDI). The three genotoxicity assays showed negative results regardless of the solvent system used, and there were clear inconsistencies in the spatiotemporal profiles of genotoxic potential and microcystin concentrations in Taihu Lake. Risk assessment showed that the drinking water from Taihu Lake was not safe from the end of July to the beginning of November because of a high concentration of microcystin-LR. Our study indicated the drinking water from Taihu Lake posed a risk because of the microcystin-LR, although it was neither genotoxic nor associated with genotoxicity of the lake water.     

The delayed genotoxic effect of N-nitroso N-propoxur insecticide in mammalian cells.

The N-nitroso derivative of an extensively used insecticide, propoxur, consistently induced dose-responsive chromosome aberrations and sister-chromatid exchanges (SCEs) in Chinese hamster ovary (CHO-W8) cells. Further investigations indicated that post-treatment incubation with a regular 1.5-cell-cycle period did not offer an unbiased estimation of the genotoxicity of N-nitroso carbamate insecticides. The scale of chromosome aberration induction increased with extension of the post-treatment incubation period. Comparable phenomena were not found in CHO-AGT cells proficient for O(6)-methylguanine-DNA-methyltransferase. In CHO-W8 cells, pulsed-treatment of the insecticide in the 1st replication cycle showed higher SCE induction than in the 2nd cycle. Similar phenomenon was also found in SCE induced by N-nitroso derivatives from other carbamate insecticides including aldicarb, carbofuran and methomyl. Treated cells did not show significantly perturbed cell cycle progression until 12 h after treatment removal. Based on the above observations, the O(6)-methylguanine-DNA adduct is suggested to be the major lesion caused by the delayed genotoxic effect of N-methyl carbamate insecticides as described in this report.     

Predicting the carcinogenic potential of environmental nitropyrenes

Nitrated polycyclic aromatic hydrocarbons (nitroarenes) constitute a large group of anthrapogenic environmental contaminants. Some members of the group are mutagenic and genotoxic in a wide spectrum of systems while others are either mutagenic only in Salmonella typhimurium or devoid of activity altogether. Additionally, some nitroarenes are uniformly carcinogenic in rodents while others are not. In view of the logistic and economic problems involved in preparing large quantities of individual nitroarenes in pure (greater than 99.5%) form and assaying them for carcinogenicity in animals, we have analyzed the results that nitropyrenes and related chemicals yield in short-term tests by CPBS, the Carcinogen Prediction, and Battery Selection method, and demonstrate that CPBS classifies them correctly with respect to carcinogenicity.     

Intestinal inflammation induces genotoxicity to extraintestinal tissues and cell types in mice

Chronic intestinal inflammation leads to increased risk of colorectal and small intestinal cancers and is also associated with extraintestinal manifestations such as lymphomas, other solid cancers and autoimmune disorders. We have previously found that acute and chronic intestinal inflammation causes DNA damage to circulating peripheral leukocytes, manifesting a systemic effect in genetically and chemically induced models of intestinal inflammation. Our study addresses the scope of tissue targets and genotoxic damage induced by inflammation-associated genotoxicity. Using several experimental models of intestinal inflammation, we analyzed various types of DNA damage in leukocyte subpopulations of the blood, spleen, mesenteric and peripheral lymph nodes and in intestinal epithelial cells, hepatocytes and the brain. Genotoxicity in the form of DNA single- and double-stranded breaks accompanied by oxidative base damage was found in leukocyte subpopulations of the blood, diverse lymphoid organs, intestinal epithelial cells and hepatocytes. The brain did not demonstrate significant levels of DNA double-stranded breaks as measured by γ-H2AX immunostaining. CD4(+) and CD8(+) T-cells were most sensitive to DNA damage versus other cell types in the peripheral blood. In vivo measurements and in vitro modeling suggested that genotoxicity was induced by increased levels of systemically circulating proinflammatory cytokines. Moreover, genotoxicity involved increased damage rather than reduced repair, as it is not associated with decreased expression of the DNA double-strand break recognition and repair protein, ataxia telangiectasia mutated. These findings suggest that levels of intestinal inflammation contribute to the remote tissue burden of genotoxicity, with potential effects on nonintestinal diseases and cancer.     

微核试验和彗星试验检测朱砂的遗传毒性

目的:研究朱砂对大小鼠染色体的损伤作用,比较短期给药与长期给药对遗传毒性检出的影响,探讨结合生殖毒性Ⅰ段试验进行遗传毒性研究的可行性。方法:18只小鼠分3组灌胃给药10,5.02,.5 g.kg-1(分别相当于人临床最高等效剂量约100,50,25倍)朱砂悬浊液,2 d后处死;结合生殖毒性Ⅰ段大鼠(每组6只)连续灌胃给药1.0,0.3,0.1 g.kg-1(分别相当于人临床最高等效剂量的20,6.4,2.0倍),雄性42 d以上,交配成功后处死;雌性20 d以上,妊娠第15天处死,取骨髓细胞做微核试验和彗星试验。结果:小鼠各剂量组灌胃给药的微核率分别为0.175%,0.108%,0.092%,与阴性对照组比较差异有显著意义,但彗星试验结果阴性。结合生殖毒性Ⅰ段试验的微核试验中雄性和雌性大鼠的微核率与阴性对照组比较,差异无显著意义,但有随剂量增高而增高的趋势;而彗星试验结果显示雄性的中、高剂量和雌性的高剂量的拖尾阳性率为27.6%,42.8%,22.3%,与阴性对照组比较,差异有显著和极显著意义。结论:①朱砂短期内大剂量灌胃给药或长期小剂量给药可能引起染色体损伤;②利用生殖毒性Ⅰ段试验多次给药后取材做微核试验和彗星试验在方法上是可行的,在剂量设计上更符合中药长期低剂量给药方式;③微核试验和彗星试验的组合在评价药物的体内遗传毒性中具有互补性。     

Cytotoxic and genotoxic effects of paraquat in Hordeum vulgare and human lymphocytes in vitro

Two phylogenetically distant types of test‐systems—root tip meristems of barley (Hordeum vulgare) and human lymphocytes in vitro were used to detect genotoxicity and cytotoxicity induced by the herbicide paraquat (PQ) in the concentration range (10^?6 to 5 × 10^?4 mol/l). As an endpoint for cytotoxicity the mitotic index (MI) was evaluated. The frequency of chromosome aberrations (CA) and the frequency of micronuclei (MN) were used as endpoints for genotoxicity. A dose‐dependent increase of CA and MN was observed in both test systems, although the values for PQ‐induced MN were somewhat lower. The increase of the genotoxic effect corresponds to a decrease of mitotic activity. The structurally reconstructed barley karyotype MK14/2034 allowed the allocation of the PQ‐specific features of aberration distribution patterns and gave information about which chromosome segments in different chromosomal positions were involved in induced aberrations. Paraquat produced preferably isochromatid breaks and “aberration hot spots” in a restricted number of heterochromatin‐containing segments. The comparative analysis of susceptibility in the used test‐systems to PQ with respect to its cytotoxic and clastogenic effect showed that the human lymphocytes were more sensitive than Hordeum vulgare. © 2009 Wiley Periodicals, Inc. Environ Toxicol, 2010.     

Genotoxicity of retroviral hematopoietic stem cell gene therapy

Introduction: Retroviral vectors have been developed for hematopoietic stem cell (HSC) gene therapy and have successfully cured X-linked severe combined immunodeficiency (SCID-X1), adenosine deaminase deficiency (ADA-SCID), adrenoleukodystrophy, and Wiskott-Aldrich syndrome. However, in HSC gene therapy clinical trials, genotoxicity mediated by integrated vector proviruses has led to clonal expansion, and in some cases frank leukemia. Numerous studies have been performed to understand the molecular basis of vector-mediated genotoxicity with the aim of developing safer vectors and safer gene therapy protocols. These genotoxicity studies are critical to advancing HSC gene therapy.

Areas covered: This review provides an introduction to the mechanisms of retroviral vector genotoxicity. It also covers advances over the last 20 years in designing safer gene therapy vectors, and in integration site analysis in clinical trials and large animal models. Mechanisms of retroviral-mediated genotoxicity, and the risk factors that contribute to clonal expansion and leukemia in HSC gene therapy are introduced.

Expert opinion: Continued research on virus–host interactions and next-generation vectors should further improve the safety of future HSC gene therapy vectors and protocols.     

Assessment of the potential in vivo ecotoxicity of Double-Walled Carbon Nanotubes (DWNTs) in water,using the amphibian Ambystoma mexicanum

Because of their specific properties (mechanical, electrical, etc), carbon nanotubes (CNTs) are being assessed for inclusion in many manufactured products. Due to their massive production and number of potential applications, the impact of CNTs on the environment must be taken into consideration. The present investigation evaluates the ecotoxic potential of CNTs in the amphibian larvae (Ambystoma mexicanum). Acute toxicity and genotoxicity were analysed after 12 days of exposure in laboratory conditions. The genotoxic effects were analysed by scoring the micronucleated erythrocytes in the circulating blood of the larvae according to the French standard micronucleus assay. The results obtained in the present study demonstrated that CNTs are neither acutely toxic nor genotoxic to larvae whatever the CNTs concentration in the water, although black masses of CNTs were observed inside the gut. In the increasing economical context of CNTs, complementary studies must be undertaken, especially including mechanistic and environmental investigations.     

In vitro Evaluation of Genotoxicity of Avocado (Persea americana) Fruit and Leaf Extracts in Human Peripheral Lymphocytes

Persea americana is much sought after both for the nutritional value of its fruit and the medicinal values of its various plant parts. A chromosomal aberration assay was undertaken to evaluate the potential genotoxicity of crude extracts from avocado fruits and leaves. Chromosomal aberrations were observed in cultured human peripheral lymphocytes exposed to separately increasing concentrations of 50% methanolic extracts of Persea americana fruit and leaves. The groups exposed to leaf and fruit extracts, respectively, showed a concentration-dependent increase in chromosomal aberrations as compared to that in a control group. The mean percentage total aberrant metaphases at 100 mg/kg, 200 mg/kg, and 300 mg/kg concentrations of leaf extract were found respectively to be 58 ± 7.05, 72 ± 6.41, and 78 ± 5.98, which were significantly higher (p < 0.0001 each) than that in the control group (6 ± 3.39). The mean percentage total aberrant metaphases at 100 mg/kg, 200 mg/kg, and 300 mg/kg concentrations of fruit extract were found to be 18 ± 5.49, 40 ± 10.00, and 52 ± 10.20, respectively, which were significantly higher (p = 0.033, p < 0.0001, and p < 0.0001, respectively) than that for control (6 ± 3.39). Acrocentric associations and premature centromeric separation were the two most common abnormalities observed in both the exposed groups. The group exposed to leaf extracts also showed a significant number of a variety of other structural aberrations, including breaks, fragments, dicentrics, terminal deletion, minutes, and Robertsonian translocations. The group exposed to leaf extract showed higher frequency of all types of aberrations at equal concentrations as compared to the group exposed to fruit extract.     

Genotoxic assessment and toxicity evaluation of peginesatide in CByB6F1 hybrid mice

Peginesatide is a PEGylated, investigational, peptide-based erythropoiesis-stimulating agent (ESA) that was designed and engineered to stimulate specifically the erythropoietin receptor dimer that governs erythropoiesis. Clinical use of peginesatide is anticipated to result in chronic dosing in chronic kidney disease (CKD) patients, and the nonclinical data to support development should include an evaluation of carcinogenic potential evaluation. Peginesatide was not mutagenic or clastogenic in a standard genotoxicity battery of tests. Doses for a rasH2 transgenic mouse carcinogenicity assay were defined in a 28-day study in the wild-type littermates of the rasH2 transgenic mouse strain, using intravenous doses of 1–25?mg/kg on days 1 and 22. The findings were consistent with exaggerated pharmacology, including polycythemia, with associated increases in hemoglobin level and extramedullary hematopoiesis and bone marrow hypercellularity.