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Acute myeloid leukaemia (AML) is associated with exposure to benzene and treatment with chemotherapeutic agents. It is thought to arise from damage to specific regions of DNA, resulting in chromosome rearrangements or loss. For instance, a deletion on the long arm of chromosome 5 [e.g. del(5q31)] is common in AML patients previously treated with alkylating agents, such as melphalan, or exposed to benzene. Translocations of the MLL gene at 11q23 are frequently observed in AML arising from treatment with topoisomerase II inhibitors, such as etoposide. Our goal was to determine whether or not breakage at 5q31 and 11q23 is selectively induced by these chemical agents. To address this question, the comet assay combined with fluorescence in situ hybridization (comet-FISH) was used to detect DNA breakage in the specific chromosomal regions in an in vitro model. TK6 lymphoblastoid cells were exposed to melphalan, etoposide or the benzene metabolite, hydroquinone (HQ), at various concentrations. HQ, melphalan and etoposide induced DNA breaks at both 5q31 and 11q23 chromosome regions in a dose-dependant manner. However, HQ produced significantly more DNA damage at 5q31 than at 11q23. Etoposide produce slightly more DNA damage at 11q23 and melphalan had a somewhat greater effect at 5q31, but not significantly so. Thus, HQ and melphalan act similarly, perhaps explaining some similarities between benzene- and alkylating agent-induced AML. Comet-FISH also appears to be a useful approach for detecting and comparing damage to specific chromosome regions of significance in leukaemogenesis.

Benzene is an established cause of adult leukemia, but whether it is associated with childhood leukemia remains unclear. We conducted a meta-analysis in which we reviewed the epidemiologic literature on this topic and explored causal inference, bias, and heterogeneity. The exposure metrics that we evaluated included occupational and household use of benzenes and solvents, traffic density, and traffic-related air pollution. For studies of occupational and household product exposure published from 1987 to 2014, the summary relative risk for childhood leukemia was 1.96 (95% confidence interval (CI): 1.53, 2.52; n = 20). In these studies, the summary relative risk was higher for acute myeloid leukemia (summary relative risk (sRR) = 2.34, 95% CI: 1.72, 3.18; n = 6) than for acute lymphoblastic leukemia (sRR = 1.57; 95% CI: 1.21, 2.05; n = 14). The summary relative risk was higher for maternal versus paternal exposure, in studies that assessed benzene versus all solvents, and in studies of gestational exposure. In studies of traffic density or traffic-related air pollution published from 1999 to 2014, the summary relative risk was 1.48 (95% CI: 1.10, 1.99; n = 12); it was higher for acute myeloid leukemia (sRR = 2.07; 95% CI: 1.34, 3.20) than for acute lymphoblastic leukemia (sRR = 1.49; 95% CI: 1.07, 2.08) and in studies that involved detailed models of traffic pollution (sRR = 1.70; 95% CI: 1.16, 2.49). Overall, we identified evidence of associations between childhood leukemia and several different potential metrics of benzene exposure.

Formaldehyde (FA) is a human leukemogen and is hematotoxic in human and mouse. The biological plausibility of FA-induced leukemia is controversial because few studies have reported FA-induced bone marrow (BM) toxicity, and none have reported BM stem/progenitor cell toxicity. We sought to comprehensively examine FA hematoxicity in vivo in mouse peripheral blood, BM, spleen and myeloid progenitors. We included the leukemogen and BM toxicant, benzene (BZ), as a positive control, separately and together with FA as co-exposure occurs frequently. We exposed BALB/c mice to 3 mg/m3 FA in air for 2 weeks, mimicking occupational exposure, then measured complete blood counts, nucleated BM cell count, and myeloid progenitor colony formation. We also investigated potential mechanisms of FA toxicity, including reactive oxygen species (ROS) generation, apoptosis, and hematopoietic growth factor and receptor levels. FA exposure significantly reduced nucleated BM cells and BM-derived colony-forming unit-granulocyte-macrophage (CFU-GM) and burst-forming unit-erythroid (BFU-E); down-regulated GM-CSFRα and EPOR expression; increased ROS in nucleated BM, spleen and CFU-GM cells; and increased apoptosis in nucleated spleen and CFU-GM cells. FA and BZ each similarly altered BM mature cells and stem/progenitor counts, BM and CFU-GM ROS, and apoptosis in spleen and CFU-GM but had differential effects on other end points. Co-exposure was more potent for several end points. Thus, FA is toxic to the mouse hematopoietic system, including BM stem/progenitor cells, and it enhances BZ-induced toxic effects. Our findings suggest that FA may induce BM toxicity by affecting myeloid progenitor growth and survival through oxidative damage and reduced expression levels of GM-CSFRα and EPOR.

Ecosystem valuation can serve as a basis of scientific support for decision-making. So far, the most popular methods for ecosystem valuation are ecosystem service valuation (ESV), which is based on the utility of ecosystem to human beings rather than on the objective value of the ecosystem. After more than 10 years' application, it has been found that all losses of ESV were about 10% of the benefits of human activities. In this paper, the ecosystem intrinsic value (EIV) is defined. EIV is an objective value that emerges from the existence, structures, functions and processes of ecosystem, but independent with man, man's will and preferences. The valuating approach and methods used for EIV were developed by using energy approach and the methods of emergy analysis and eco-exergy analysis. The EIV calculated by emergy from the substance, energy and information of ecosystem and by eco-exergy from the structure and function of ecosystem represents the existent value and the externally working capacity of ecosystem, respectively. The approach and methods of EIV evaluation were applied to Xiamen Bay, China. The results showed that the marine EIV in Xiamen Bay was 209 billion RMB, including 116 billion RMB of emergy and 92.4 billion RMB of eco-exergy in 2010, nearly 30 times of ESV and 8.5 times of the GDP of Xiamen marine industry in 2010. The EIV in unit area of Xiamen Bay is more than 10 times higher than the average global ESV in estuaries. It implied a potential undervaluation to ecosystem value by ESV calculation, which may mislead decisions-making processes.

Characterizing variability in the extent and nature of responses to environmental exposures is a critical aspect of human health risk assessment. Chemical toxicants act by many different mechanisms, however, and the genes involved in adverse outcome pathways (AOPs) and AOP networks are not yet characterized. Functional genomic approaches can reveal both toxicity pathways and susceptibility genes, through knockdown or knockout of all non-essential genes in a cell of interest, and identification of genes associated with a toxicity phenotype following toxicant exposure. Screening approaches in yeast and human near-haploid leukemic KBM7 cells, have identified roles for genes and pathways involved in response to many toxicants but are limited by partial homology among yeast and human genes and limited relevance to normal diploid cells. RNA interference (RNAi) suppresses mRNA expression level but is limited by off-target effects (OTEs) and incomplete knockdown. The recently developed gene editing approach called clustered regularly interspaced short palindrome repeats-associated nuclease (CRISPR)-Cas9, can precisely knock-out most regions of the genome at the DNA level with fewer OTEs than RNAi, in multiple human cell types, thus overcoming the limitations of the other approaches. It has been used to identify genes involved in the response to chemical and microbial toxicants in several human cell types and could readily be extended to the systematic screening of large numbers of environmental chemicals. CRISPR-Cas9 can also repress and activate gene expression, including that of non-coding RNA, with near-saturation, thus offering the potential to more fully characterize AOPs and AOP networks. Finally, CRISPR-Cas9 can generate complex animal models in which to conduct preclinical toxicity testing at the level of individual genotypes or haplotypes. Therefore, CRISPR-Cas9 is a powerful and flexible functional genomic screening approach that can be harnessed to provide unprecedented mechanistic insight in the field of modern toxicology.

Some previous studies have suggested that home pesticide exposure before birth and during a child's early years may increase the risk of childhood leukemia. To further investigate this, we pooled individual level data from 12 case-control studies in the Childhood Leukemia International Consortium. Exposure data were harmonized into compatible formats. Pooled analyses were undertaken using multivariable unconditional logistic regression. The odds ratio (ORs) for acute lymphoblastic leukemia (ALL) associated with any pesticide exposure shortly before conception, during pregnancy and after birth were 1.39 (95% confidence interval [CI]: 1.25, 1.55) (using 2,785 cases and 3,635 controls), 1.43 (95% CI: 1.32, 1.54) (5,055 cases and 7,370 controls) and 1.36 (95% CI: 1.23, 1.51) (4,162 cases and 5,179 controls), respectively. Corresponding ORs for risk of acute myeloid leukemia (AML) were 1.49 (95% CI: 1.02, 2.16) (173 cases and 1,789 controls), 1.55 (95% CI: 1.21, 1.99) (344 cases and 4,666 controls) and 1.08 (95% CI: 0.76, 1.53) (198 cases and 2,655 controls), respectively. There was little difference by type of pesticide used. The relative similarity in ORs between leukemia types, time periods and pesticide types may be explained by similar exposure patterns and effects across the time periods in ALL and AML, participants' exposure to multiple pesticides, or recall bias. Although some recall bias is likely, until a better study design can be found to investigate the associations between home pesticide use and childhood leukemia in an equally large sample, it would appear prudent to limit the use of home pesticides before and during pregnancy, and during childhood.

Histone deacetylase inhibitors (HDACi) are a class of putative chemotherapeutic agents for which the mechanism of toxicity has not been fully identified. To explore the possibility that HDACi are genotoxic, human TK6 lymphoblastoid cells were exposed to trichostatin A (TSA) and genetic damage was measured. TSA caused a dose-dependent increase of G1-arrested cells at 24 h that correlated with increasing levels of p21 and apoptosis. Significantly elevated frequencies of structural chromosomal aberrations in cells exposed to TSA were observed using both the kinetochore-antibody micronucleus assay and nonbanding metaphase chromosome analysis. Increased tail intensities, indicative of elevated levels of DNA damage, were observed using the alkaline comet assay. Elevated levels of phosphorylated histone gammaH2AX protein were observed as early as 3 h following TSA exposure and peaked at 12 h for 200nM TSA. Significant levels of aneuploidy at the 200nM TSA dose were observed using metaphase analysis, but interestingly, kinetochore-positive micronuclei were not detected at any dose using the kinetochore micronucleus assay, suggesting that TSA induces aneuploidy via a nondisjunction event rather than chromosome lagging. Increases in chromosomal loss and breakage were observed using simultaneous FISH metaphase analysis of chromosomes 5, 7, 8, and 21, consistent with data obtained from the micronucleus and metaphase chromosome analyses. We conclude that TSA is both a clastogen and aneugen in the TK6 cell line and propose that the observed cytostatic and apoptotic properties of TSA may partially be due to this genotoxicity.

Environmental planning at different levels has been called for by environmental departments in China during last years to integrate environment consideration to economic development. Based on the accomplished environmental planning practice in Xiamen China, the paper suggests a general environmental planning process with ecological function zoning as its key steps, at the same time an effective method for ecological function zoning was formulated. The case studies of Dongfu suburban town environmental planning and Xiamen eco-city conceptual planning indicate that scientifically sound results of ecological function zoning can contribute to improving not only the adaptability and acceptability of environmental planning, also the environmental management and the decision-making. The experience shows that the ecological function zoning methods should be applied according to the principles of adaptive management, resource-based and community-based, so that it can integrate science into decision-making process, avoiding both narrow-minded viewpoints of planners and natural resource use conflicts among variety of stakeholders. Problems to be resolved in the future are also pointed out in the ending part.

In China, concern is growing with regard to environmental issues related to the growth of peri-urban towns. Rapid urbanization and the backlog in constructing needed environmental infrastructure have resulted in severe environmental issues for many peri-urban towns, affecting their sustainable development. In Xiamen, a coastal city, town-level environmental planning was proposed in 2003 as an important tool to control pollution and conserve ecosystems. But for environmental planning to work, it needs to be more adaptable to local circumstances and decision-making processes. This paper presents a framework for adaptive environmental management and discusses how it was implemented in Dongfu Town, one of the peri-urban towns around Xiamen. It describes the principles of adaptive peri-urban town environmental planning and how this process integrates information from multiple sources and different stakeholders. This experience shows how science can be integrated into the decision-making process, avoiding both the narrow-minded viewpoints of planners and the natural resource use conflicts among a variety of stakeholders. It also shows how the environmental planning process can propose results that are acceptable to local communities and decision makers, even when they suggest directions that were not in accordance with the public's initial priorities. Finally, it shows that sustainable development of targeted areas can be achieved through well-designed environmental planning.

Swine finishing produces a large contribution of manure to nearby surface water and ground water in Jiulong upstream watershed, south Fujian Province, China. It is urgent to find a comprehensive, cost-effective way, not only to solve the manure pollution problem, but also to utilize the nutrients of manure for local farming. Gekou, a small hoggery village in the dense swine-farming core area of the upstream watershed, was selected as the study area. Mixed integer programming models based on uncertain interval numbers were employed to generate optimal whole farm plans for specialized hoggery. Emphasis was placed on discerning the most profitable ways to formulate hog feed rations, manure collection, manure storage, and manure application to achieve the maximum profit and the minimum pollution emission. Analyses were concentrated on ammonia emissions, excess total nitrogen and excess total phosphorus from manure operations, not only as nutrients applied to croplands, but also as pollutants to be minimized in compliance with environmental criteria. Three different hog feed rations, two alternative ways of collecting hog manure, and two alternative storage methods were combined with five alternatives for field application of manure to provide different combinations of hog feeding and manure-handling to be evaluated in the mixed integer programming models. Results indicated tradeoffs between economic goals and environmental goals, the latter being achieved only at some expense to farm net returns. The kaleyard and fishpond are recommended alternatives for field application of manure compared to three other alternatives.

Field surplus nitrogen and farm disposal nitrogen (N) are major sources of environmental pollution in farm systems. The purpose of this study was to use nitrogen balances at the field and farm level approaches to estimate the effects of pig production in the Jiulong River watershed, Fujian province, China, on nitrogen losses to the environment on 67 pig farms. The field surplus nitrogen ranged from 6.19 to 18.19 t nitrogen ha−1, which was caused by high pig density and excessive application of manure. Manure application rates of more than 450 to 510 kg nitrogen ha −1 would increase the potential nitrate concentration to more than 10 to 20 mg l −1. Therefore, 44 to 48 heads of pig ha −1 was suggested to as the environmental capacity for sustaining optimal nitrogen cycling in pig farms in the Jiulong River watershed. The majority of farms have potential environmental risks because of high farm nitrogen imbalances. With more than a 2:1 ratio of N inputs to outputs, N inputs were 50 percent higher than outputs. Feed was the main input source, while pig production and manure exported were the major managed outputs. Based on the farm nitrogen balance calculation, the management options that would contribute to a more favorable nitrogen balance were also identified.

The prevailing systems for decision-making in many countries tend to separate environmental factors at the policy, planning and management levels, a practice that has important implications for sustainable development. Among many methods, at different decision-making levels, environmental planning and environmental assessment are currently the most widely used in China. This article briefly introduces four case studies featuring environmental planning and assessment in Xiamen, a coastal city in southeast China. Differences in scope of application, timing, nature, legal status, methodologies and effectiveness are described. Six principles for the successful integration of environmental protection and development are advanced based on the analytical results. As well, a simplified integration framework is proposed.

This paper presents the experience of institutional capacity building in environmental education, and the achievement of the participatory approach in environmental management for the Canadian International Development Agency funded project ‘Community-Based Conservation Management: China and Vietnam’ in Xiamen. The project has enhanced teaching capacity and applied research skills of the local training institutions with integrated social and physical science lessons in multiple levels of a post-secondary environmental education programme. The focus of the project was to integrate the local secondary and post-secondary schools environmental education plans. Approaches to promote public awareness on environmental conservation are also illustrated. The study highlights the conservation activities of the Community Based Conservation Management project in Xiamen to encourage the involvement of a variety of community groups, including university students, youth and women. In addition, it explores the social responsibility of the faculty as an essential link between the public and decision-makers for effective public involvement in environmental management. We argue that the success of public environmental education relies on institutional capacity building, interdisciplinary studies and community participation.

Background: Blood miRNAs are a new promising area of disease research, but variability in miRNA measurements may limit detection of true-positive findings. Here, we measured sources of miRNA variability and determine whether repeated measures can improve power to detect fold-change differences between comparison groups. Methods: Blood from healthy volunteers (N = 12) was collected at three time points. The miRNAs were extracted by a method predetermined to give the highest miRNA yield. Nine different miRNAs were quantified using different qPCR assays and analyzed using mixed models to identify sources of variability. A larger number of miRNAs from a publicly available blood miRNA microarray dataset with repeated measures were used for a bootstrapping procedure to investigate effects of repeated measures on power to detect fold changes in miRNA expression for a theoretical case–control study. Results: Technical variability in qPCR replicates was identified as a significant source of variability (P < 0.05) for all nine miRNAs tested. Variability was larger in the TaqMan qPCR assays (SD = 0.15–0.61) versus the qScript qPCR assays (SD = 0.08–0.14). Inter- and intraindividual and extraction variability also contributed significantly for two miRNAs. The bootstrapping procedure demonstrated that repeated measures (20%–50% of N) increased detection of a 2-fold change for approximately 10% to 45% more miRNAs. Conclusion: Statistical power to detect small fold changes in blood miRNAs can be improved by accounting for sources of variability using repeated measures and choosing appropriate methods to minimize variability in miRNA quantification. Impact: This study demonstrates the importance of including repeated measures in experimental designs for blood miRNA research. See all the articles in this CEBP Focus section, “Biomarkers, Biospecimens, and New Technologies in Molecular Epidemiology.” Cancer Epidemiol Biomarkers Prev; 23(12); 2658–66. ©2014 AACR.

Although the toxicity of benzene has been linked to its metabolism, the dose-related production of metabolites is not well understood in humans, particularly at low levels of exposure. We investigated unmetabolized benzene in urine (UBz) and all major urinary metabolites [phenol (PH), E,E-muconic acid (MA), hydroquinone (HQ) and catechol (CA)] as well as the minor metabolite, S-phenylmercapturic acid (SPMA), in 250 benzene-exposed workers and 139 control workers in Tianjin, China. Median levels of benzene exposure were approximately 1.2 p.p.m. for exposed workers (interquartile range: 0.53-3.34 p.p.m.) and 0.004 p.p.m. for control workers (interquartile range: 0.002-0.007 p.p.m.). (Exposures of control workers to benzene were predicted from levels of benzene in their urine.) Metabolite production was investigated among groups of 30 workers aggregated by their benzene exposures. We found that the urine concentration of each metabolite was consistently elevated when the group's median benzene exposure was at or above the following air concentrations: 0.2 p.p.m. for MA and SPMA, 0.5 p.p.m. for PH and HQ, and 2 p.p.m. for CA. Dose-related production of the four major metabolites and total metabolites (micromol/l/p.p.m. benzene) declined between 2.5 and 26-fold as group median benzene exposures increased between 0.027 and 15.4 p.p.m. Reductions in metabolite production were most pronounced for CA and PH

Samples of surface sediment and shellfish were collected from Minjiang Estuary-Mazu and Xiamen-Jinmen sea areas and were analyzed for hexachlorocyclohexanes (HCHs), dichlorodiphenyl-trichlorophenyl (DDT) and its degraded derivatives dichloro-diphenyl-dichloroethane (DDT) and dichloro-dichlorophenyl-ethylene (DDE), and polychlorinated biphenyls (PCBs) by gas chromatography. Concentration ranges of HCHs, DDTs, and PCBs in the sediments were 0.03-0.48, 1.10-14.3, and ND to 0.76 ng/g dw, respectively; those for shellfish samples were ND to 1.5, 21.5-892, dw and ND-3.7 ng/g dw, respectively. The results showed that the concentrations of such oganochlorine compounds (OCs) in the sediments were usually low as compared with other sea areas and estuaries, and the organochlorines were accumulated in marine bivalve mollusks, particularly in oyster. With respect to the concentrations of HCHs, DDTs, and PCBs in the samples of either sediment or shellfish, DDTs were higher than HCHs and PCBs. The concentrations of OCs in shellfish samples varied with the sampling locations and organism species but were generally higher than those in the sediments. The concentration distributions, compositions, and residue levels of these OCs are extensively discussed and evaluated.

'Omic' technologies include genomics, transcriptomics (gene expression profiling), proteomics and metabolomics. We are utilizing these new technologies in an effort to develop novel biomarkers of exposure, susceptibility and response to benzene. Advances in genomics allow one to study hundreds to thousands of single nucleotide polymorphisms simultaneously on small quantities of DNA using array-based technologies. We are currently utilizing these technologies to examine genetic variation in pathways relating to biotransformation, DNA repair, folate metabolism and immune response with the goal of finding biomarkers of susceptibility to benzene hematotoxicity. Transcriptomics is used to measure the full complement of activated genes, mRNAs or transcripts in a particular tissue at a particular time typically using microarray technology. We have applied microarrays to the study of global gene expression in the peripheral blood cells of benzene-exposed workers. More than 100 genes were identified as being potentially differentially expressed, with genes related to apoptosis and immune function being the most significantly affected. Initial studies employing proteomics have also shown that several proteins are altered in the serum of exposed compared to control subjects and these proteins are potential biomarkers of benzene exposure. Omic technologies therefore have significant potential in generating novel biomarkers of exposure, susceptibility and response to benzene.

Benzene is an established human leukemogen. The mechanism of benzene-induced leukemogenesis, however, remains unclear, but chromosomal damage is thought to play a critical role. We previously reported that the loss of chromosomes 5 and 7 (monosomy 5 and 7) and the gain of chromosomes 8 and 21 (trisomy 8 and 21) are significantly increased in benzene-exposed workers in comparison to matched controls. To determine if selective effects of benzene can occur, we employed three-color painting on an 8-square slide to screen numerical changes in all 24 human chromosomes (OctoChrome FISH) in a pilot study of 11 subjects (6 exposed to5 ppm benzene and 5 age- and sex-matched controls). The effects of benzene on each chromosome were assessed as the incidence rate ratio (IRR) from a Poisson regression model with the strongest effects being reflected by the highest IRR values. Monosomy of chromosomes 5, 6, 7 and 10 had the highest IRRs and statistical significance in this preliminary study (IRR2.5, p0.01). On the other hand, the monosomy levels of six other chromosomes (1, 4, 9, 11, 22 and Y) were unchanged in the exposed workers with IRRs close to 1.0. Similarly, selective effects were also observed on trisomy induction with chromosomes 8, 9, 17, 21 and 22 (IRR2.5, p0.01). These results suggest that benzene has the capability of producing selective effects on certain chromosomes, which is supported by our in vitro findings showing that chromosomes 5 and 7 are more sensitive to loss than other chromosomes following exposure to benzene metabolites. We are currently investigating potential mechanisms for this induction of selective aneuploidy.

We used cysteinyl adducts of serum albumin (Alb) to investigate the production of two reactive benzene metabolites, namely, benzene oxide (BO) and 1,4-benzoquinone (1,4-BQ) in workers exposed to benzene. Adducts were measured in 160 benzene-exposed workers who did not use respiratory protection (based upon individual geometric mean benzene exposure levels: median = 5.27 ppm, interquartile range = 2.14–13.4 ppm, range = 0.074–328 ppm) and 101 local controls, from populations in Shanghai and Tianjin, China. After isolation of Alb, these adducts (designated as BO-Alb and 1,4-BQ-Alb) were cleaved from the protein with methanesulfonic acid and trifluoroacetic anhydride and measured by gas chromatography-mass spectrometry. Although BO-Alb and 1,4-BQ-Alb were measured in all subjects, levels of both adducts were 2.4-fold greater (median value) in exposed subjects than in controls (interquartile-fold range = 1.63–4.05 for BO-Alb and 1.64–3.69 for 1,4-BQ-Alb). Log–log plots of the individual adduct levels versus exposure were quasi-linear with straight-line slopes of about 0.3 for both BO-Alb and 1,4-BQ-Alb. Since these log-space slopes were significantly less than one, we infer that adduct production was nonlinear, i.e., less-than proportional to benzene exposure, over the indicated range. This behavior points to saturation of CYP2E1 as a critical metabolic consequence of high exposure to benzene in humans. Thus, the biologically effective dose of BO and 1,4-BQ should be proportionally greater in persons exposed to low rather than high levels of benzene.

The human FHIT (fragile histidine triad) gene is a putative tumor suppressor gene located at chromosome region 3p14.2. Previous studies have shown that loss of heterozygosity, homozygous deletions, and abnormal expression of the FHIT gene are involved in several types of human malignancies. A CpG island is present in the 5′ promoter region of the FHIT gene, and methylation in this region correlates with loss of FHIT expression. To test whether aberrant methylation of the FHIT gene may play a role in pediatric leukemia, we assessed the FHIT methylation status of 10 leukemia cell lines and 190 incident population-based cases of childhood acute lymphocytic and myeloid leukemias using methylation-specific PCR. Conventional and fluorescence in situ hybridization cytogenetic data were also collected to examine aneuploidy, t(12, 21), and other chromosomal rearrangements. Four of 10 leukemia cell lines (40%) and 52 of 190 (27.4%) bone marrows from childhood leukemia patients demonstrated hypermethylation of the promoter region of FHIT. Gene expression analyses and 5-aza-2′-deoxycytidine treatment showed that promoter hypermethylation correlated with FHIT inactivation. Among primary leukemias, hypermethylation of FHIT was strongly correlated with acute lymphoblastic leukemia (ALL) histology (P = 0.008), high hyperdiploid (P < 0.0001), and translocation-negative (P < 0.0001) categories. Hyperdiploid B-cell ALLs were 23-fold more likely to be FHIT methylated compared with B-cell ALL harboring TEL-AML translocations. FHIT methylation was associated with high WBC counts at diagnosis, a known prognostic indicator. These results suggest that hypermethylation of the promoter region CpG island of the FHIT gene is a common event and may play an important role in the etiology and pathophysiology of specific cytogenetic subtypes of childhood ALL.

1,3-Butadiene (BD) is an important industrial chemical and pollutant. Its ability to induce genetic damage and cause hematological malignancies in humans is controversial. We have examined chromosome damage by fluorescence in situ hybridization (FISH) and mutations in the HPRT gene in the blood of Chinese workers exposed to BD. Peripheral blood samples were collected and cultured from 39 workers exposed to BD (median level 2 ppm, 6 h time-weighted average) and 38 matched controls in Yanshan, China. No difference in the level of aneuploidy or structural changes in chromosomes 1, 7, 8, and 12 was detected in metaphase cells from exposed subjects in comparison with matched controls, nor was there an increase in the frequency of HPRT mutations in the BD-exposed workers. Because genetic polymorphisms in glutathione S-transferase (GST) enzymes and microsomal epoxide hydrolase (EPHX1) may affect the genotoxic effects of BD and its metabolites, we also related chromosome alterations and gene mutations to GSTT1, GSTM1 and EPHX1 genotypes. Overall, there was no effect of variants in these genotypes on numerical or structural changes in chromosomes 1, 7, 8 and 12 or on HPRT mutant frequency in relation to BD exposure, but the GST genotypes did influence background levels of both hyperdiploidy and HPRT mutant frequency. In conclusion, our data show no increase in chromosomal aberrations or HPRT mutations among workers exposed to BD, even in potentially susceptible genetic subgroups. The study is, however, quite small and the levels of BD exposure are not extremely high, but our findings in China do support those from a similar study conducted in the Czech Republic. Together, these studies suggest that low levels of occupational BD exposure do not pose a significant risk of genetic damage.

This paper uses the travel cost method to evaluate the recreational benefits of a beach along the eastern coast of Xiamen Island in China. Our results indicate that the total value for the beach and its associated recreation is in excess of US$53 million. This paper also discusses the protection of this significant tourism resource and considers the use of a suitable entrance fee.

Acute lymphoblastic leukemia (ALL) is the most common form of childhood cancer. The peak incidence of ALL between ages 2 and 5 is accounted for by one subtype, referred to as common acute lymphoblastic leukemia (cALL). About 25% of cALL patients have the TEL-AML1 gene fusion derived from the t(12;21) chromosomal translocation. Recent evidence from retrospective analysis of neonatal blood spots (Guthrie cards) in Europe has demonstrated that this chromosome translocation may arise prenatally. The aim of our study was to determine whether TEL-AML1 fusions arise prenatally in a U.S. population of cALL patients. TEL-AML1-positive cALL cases (n = 14) were identified by fluorescence in situ hybridization, and the genomic breakpoints were identified by a streamlined long-distance PCR approach and sequenced. Clonotypic primers were designed for each patient breakpoint, and a nested PCR assay was used to determine the presence of the TEL-AML1 fusion sequence in neonatal Guthrie cards. Seven of 14 cases demonstrated clonotypic sequences on the archival Guthrie cards. The oldest patient that was positive was 6.7 years old at the time of diagnosis of leukemia. These results confirm previously published findings of a prenatal origin of TEL-AML1 in Europe by demonstrating its occurrence in a California-born population. Secondary changes were also similar to those described previously, with deletion of the second TEL allele being the most common. Other secondary changes included duplication of the fusion gene, trisomy 21, and monosomy X.

Recent reports have established the prenatal origin of leukemia translocations and resultant fusion genes in some patients, including MLL-AF4 translocations in infants and TEL-AML1 translocations in children. We now report evidence for the prenatal origin of a translocation in childhood acute myeloid leukemia (AML). The t(8;21) AML1-ETO translocations were sequenced at the genomic level in 10 diagnostic leukemia samples from children with available neonatal Guthrie blood spots. Clonotypic genomic AML1-ETO sequences were detected in the Guthrie spots for 5 individuals, providing unambiguous evidence of prenatal origin in these cases. Two of these patients were older than 10 years of age at diagnosis, indicative of a protracted postnatal latency. Three of the patients were assessed for the persistence of genomic fusion sequences in complete clinical remission samples and were found to be positive. These data indicate that t(8;21) in childhood AML can arise in utero, possibly as an initiating event in childhood AML, and may establish a long-lived or stable parental clone that requires additional secondary genetic alterations to cause leukemia.

This paper describes the results of the environmental risk assessment of the pesticides used in agriculture in Xiamen, China. The goal was to assess the impact on water resources, particularly on fisheries and mariculture. Data on ecotoxicological properties of the pesticides and their physico-chemical profile were collected. The simulation of the environmental behaviour of the pesticides in relation to the load applied onto the agricultural areas was done using the SoilFug model. Risk assessment was performed, pesticides approximate concentrations have been calculated, chemicals at highest risk were identified, and risk management measures were indicated. This study could represent a cost-effective method that may be used before engaging in expensive monitoring programs for pesticide use in developing countries, where analytical facilities are lacking.

Benzene is an established cause of human leukemia that is thought to act by producing chromosomal aberrations and altered in cell differentiation. In several recent studies increased levels of chromosomal aberrations in peripheral blood lymphocytes were correlated with a heightened risk of cancer, especially hematological malignancies. Thus, chromosomal aberrations may be a predictor of future leukemia risk. Previous studies exploring whether benzene exposure induces chromosomal aberrations have yielded mostly positive results. However, it remains unclear whether the chromosomal aberrations induced by benzene occur in a distinct pattern. Here, we thoroughly review the major chromosome studies published to date in benzene-exposed workers, benzene-poisoned and preleukemia patients, and leukemia cases associated with benzene expose. Although three cytogenetic markers (chromosomal aberrations, sister chromatid exchanges, and micronuclei) are commonly examined, our primary focus is on studies of chromosomal aberrations, because only this marker has so far been correlated with increased cancer risk. This review surveys the published literature, analyzes the study results, and discusses the characteristics of effects reported. In most studies of currently exposed workers, increases in chromosomal aberrations were observed. However, due to the relatively small number of affected individuals and variability in the reported aberrations, firm conclusions cannot be made about the involvement of specific chromosomes or chromosome regions. Further, in leukemia cases associated with benzene exposure, there is no evidence of a unique pattern of benzene-induced chromosomal aberrations in humans. Leukemia cases associated with benzene exposure are, however, more likely to contain clonal chromosome aberrations then those arising de novo in the general population.

It has been suggested that home paint exposure increases the risk of childhood acute lymphoblastic leukemia (ALL). We obtained individual level data from eight case–control studies participating in the Childhood Leukemia International Consortium. All studies had home paint exposure data (sometimes including lacquers and varnishes) for the pregnancy period with additional data for the 1–3-month period before conception in five, the year before conception in two, and the period after birth in four studies, respectively. Cytogenetic subtype data were available for some studies. Data were harmonized to a compatible format. Pooled analyses of individual data were undertaken using unconditional logistic regression. Based on 3,002 cases and 3,836 controls, the pooled odds ratio (OR) for home paint exposure in the 1–3 months before conception and risk of ALL was 1.54 [95 % confidence interval (CI) 1.28, 1.85], while based on 1,160 cases and 1,641 controls for exposure in the year before conception, it was 1.00 (95 % CI 0.86, 1.17). For exposure during pregnancy, using 4,382 cases and 5,747 controls, the pooled OR was 1.14 (95 % CI 1.04, 1.25), and for exposure after birth, the OR was 1.22 (95 % CI 1.07, 1.39), based on data from 1,962 cases and 2,973 controls. The risk was greater for certain cytogenetic subtypes and if someone other than the parents did the painting. Home paint exposure shortly before conception, during pregnancy, and/or after birth appeared to increase the risk of childhood ALL. It may be prudent to limit exposure during these periods.

Exposure to benzene, a known leukemogen and probable lymphomagen, has been demonstrated to result in oxidative stress, which has previously been associated with altered telomere length (TL). TL specifically has been associated with several health outcomes in epidemiologic studies, including cancer risk, and has been demonstrated to be altered following exposure to a variety of chemical agents. To evaluate the association between benzene exposure and TL, we measured TL by monochrome multiplex quantitative PCR in 43 workers exposed to high levels of benzene and 43 age and sex-matched unexposed workers in Shanghai, China. Benzene exposure levels were monitored using organic vapor passive dosimetry badges before phlebotomy. The median benzene exposure level in exposed workers was 31 ppm. The mean TL in controls, workers exposed to levels of benzene below the median (≤31 ppm), and above the median (>31 ppm) was 1.26 ± 0.17, 1.25 ± 0.16, and 1.37 ± 0.23, respectively. Mean TL was significantly elevated in workers exposed to >31 ppm of benzene compared with controls (P = 0.03). Our findings provide evidence that high levels of occupational benzene exposure are associated with TL. Environ. Mol. Mutagen. 55:673–678, 2014.

Trace organic pollutants (PAHs, DDTs, HCHs, PCBs) in sediments, suspended particulate matter and bivalves were studied during 1994 and 1997 in three estuaries of China: the Jiulong River, the Minjiang River and the Pearl River estuaries. Results indicated that there were some differences between the composition characteristics of the trace organic pollutants in the suspended particulate matter and the sediments. Concentrations of trace organic pollutants in the bivalves were 2–300 times higher than those in the sediments. In the three estuaries, the concentrations of PAHs and PCBs were relatively higher in Pearl River estuary, but the concentrations of DDTs were higher in Jiulong River and Minjiang River estuaries. Relatively high values of PAHs appeared at the outlet of river, waterway and harbor. It is suspected that these PAHs mainly came from sewage, shipping discharge and local pollutant sources. Relatively high concentrations of PCBs appeared at the Pearl River estuary that may be related t...

We conducted genome-wide association studies of non-Hodgkin lymphoma using Illumina HumanHap550 BeadChips to identify subtype-specific associations in follicular, diffuse large B-cell and chronic lymphocytic leukemia/small lymphocytic lymphomas. We found that rs6457327 on 6p21.33 was associated with susceptibility to follicular lymphoma (FL; N = 189 cases, 592 controls) with validation in another 456 FL cases and 2,785 controls (combined allelic P = 4.7 x 10(-11)). The region of strongest association overlapped C6orf15 (STG), located near psoriasis susceptibility region 1 (PSORS1).

Samples of surface sediment and suspended particulate matter (SPM) were collected from the Pearl River estuary, China, and the distribution and concentration of hexachlorocyclohexanes (HCHs), DDTs and polychlorinated biphenyls (PCBs) were extensively studied. The concentration ranges of HCHs, DDTs and PCBs in the sediments were 0.28–1.23 ng g−1, 1.36–8.99 ng g−1 and 0.18–1.82 ng g−1, respectively. The concentrations of HCHs, DDTs and PCBs in the SPM varied both with the sampling locations and the season of collection. Higher concentrations were recorded in the SPM as compared with sediments. The distribution pattern of such organochlorine compounds (OCs), in the Pearl River estuary, showed that sources were some major river mouths and input from Shenzhen Bay. The concentrations of OCs were, however, low as compared with other estuaries and seas. The environmental fate of the OCs during estuarine mixing was determined, in part, by physicochemical and biochemical properties. The absorption and sedimentation of SPM were also considered important factors. Marine sediments may, therefore, be regarded as an important reservoir of hydrophobic and persistent OCs.

Two of the most common cytogenetic changes in therapy- and chemically-related leukemia are the loss and long (q) arm deletions of chromosomes 5 and 7 (i.e. -5, -7, del(5q) and del(7q)). We have used a novel fluorescence in situ hybridization (FISH) procedure to determine if the benzene metabolites hydroquinone (HQ) and 1,2,4-benzenetriol (BT) can induce these specific changes in human lymphocytes cultured as whole blood. Metaphase spreads were prepared and hybridized with centromeric probes for chromosomes 1, 5 and 7 and sequence specific probes for 5q31 and 7q36-qter. HQ and BT significantly increased monosomy 5 and 7 by 3-5 fold (p < 0.0001). Both HQ and BT also significantly increased the rate of del(5q) and del(7q) by 8-12 fold (p < 0.0001). Chromosome 7 was especially susceptible to aneusomy induction by HQ and BT at low doses. These results show that metabolites of benzene are highly effective in inducing changes in chromosomes 5 and 7 that are involved in the development of myeloid leukemia.

Molybdenum is an essential element in plants and animals as a cofactor for enzymes. Molybdenum trioxide is used in metallurgical processes, in cosmetics as a pigment, and in contact lens solution, yet limited information is available on molybdenum genotoxicity. In the present study the micronucleus (MN) assay in human lymphocytes and mouse bone marrow and the dominant lethal assay in mice were used to assess the genotoxic effects of molybdenum salts in vitro and in vivo. Two salts of molybdenum were tested in whole blood cultures. Ammonium molybdate was more potent than sodium molybdate in causing a dose-dependent decrease in viability and replicative index and an increase in MN formation in binucleated lymphocytes (P < 0.001). A dose-response in both kinetochore-positive MN (caused by chromosome lagging) and kinetochore-negative MN (associated with chromosome breakage) was observed. Based on the results of a toxicity study of sodium molybdate, two doses, 200 and 400 mg/kg, were assessed in the bone marrow MN assay in mice (two i.p. injections 24 and 48 hr prior to euthanasia). A modest but statistically significant increase in MN frequency in polychromatic erythrocytes was observed (P < 0.05). The same treatment protocol was used to analyze dominant lethality. A dose-dependent increase in postimplantation loss represented mostly by early resorptions was observed the first week after treatment (P = 0.003). These preliminary data suggest that sodium molybdate induces dominant lethality at the postmeiotic stage of spermatogenesis. Overall, molybdenum salts produced moderately positive results both in vitro in human cells and in vivo in mice.

1,3-Butadiene is a carcinogen in rodents, but its potential carcinogenicity to humans remains controversial. Numerous studies have shown that butadiene and its metabolites cause sister chromatid exchanges in vitro and in vivo. To test for other types of genotoxicity, the micronucleus assay and fluorescence in situ hybridization (FISH) have been used to detect chromosome damage in human lymphocytes caused by two reactive metabolites of butadiene, diepoxybutane (DEB) and monoepoxybutene (MEB). DEB (0.5-5.0 microM) significantly increased micronucleus formation 4- to 6-fold (P

Predictive toxicology plays an important role in the assessment of toxicity of chemicals and the drug development process. While there are several well-established in vitro and in vivo assays that are suitable for predictive toxicology, recent advances in high-throughput analytical technologies and model systems are expected to have a major impact on the field of predictive toxicology. This commentary provides an overview of the state of the current science and a brief discussion on future perspectives for the field of predictive toxicology for human toxicity. Computational models for predictive toxicology, needs for further refinement and obstacles to expand computational models to include additional classes of chemical compounds are highlighted. Functional and comparative genomics approaches in predictive toxicology are discussed with an emphasis on successful utilization of recently developed model systems for high-throughput analysis. The advantages of three-dimensional model systems and stem cells and their use in predictive toxicology testing are also described.

Cancers are heterogeneous and genetically unstable. New methods are needed that provide the sensitivity and specificity to query single cells at the genetic loci that drive cancer progression, thereby enabling researchers to study the progression of individual tumors. Here, we report the development and application of a bead-based hemi-nested microfluidic droplet digital PCR (dPCR) technology to achieve 'quantitative' measurement and single-molecule sequencing of somatically acquired carcinogenic translocations at extremely low levels (

The human transcriptome is complex, comprising multiple transcript types, mostly in the form of non-coding RNA (ncRNA). The majority of ncRNA is of the long form (lncRNA, ≥ 200 bp), which plays an important role in gene regulation through multiple mechanisms including epigenetics, chromatin modification, control of transcription factor binding, and regulation of alternative splicing. Both mRNA and ncRNA exhibit additional variability in the form of alternative splicing and RNA editing. All aspects of the human transcriptome can potentially be dysregulated by environmental exposures. Next-generation RNA sequencing (RNA-Seq) is the best available methodology to measure this although it has limitations, including experimental bias. The third phase of the MicroArray Quality Control Consortium project (MAQC-III), also called Sequencing Quality Control (SeQC), aims to address these limitations through standardization of experimental and bioinformatic methodologies. A limited number of toxicogenomic studies have been conducted to date using RNA-Seq. This review describes the complexity of the human transcriptome, the application of transcriptomics by RNA-Seq or microarray in molecular epidemiology studies, and limitations of these approaches including the type of cell or tissue analyzed, experimental variation, and confounding. By using good study designs with precise, individual exposure measurements, sufficient power and incorporation of phenotypic anchors, studies in human populations can identify biomarkers of exposure and/or early effect and elucidate mechanisms of action underlying associated diseases, even at low doses. Analysis of datasets at the pathway level can compensate for some of the limitations of RNA-Seq and, as more datasets become available, will increasingly elucidate the exposure-disease continuum.

Benzene is a well-established hematotoxin. However, reports of its effects on specific blood cells have been somewhat inconsistent and the relative toxicity of benzene metabolites on peripheral blood cells in humans has not been evaluated. We compared hematologic outcomes in a cross-sectional study of 44 workers heavily exposed to benzene (median: 31 parts permillion [ppm] as an 8-hr time-weighted average [TWA] and 44 age and gender-matched unexposed controls from Shanghai, China. All hematologic parameters (total white blood cells [WBC], absolute lymphocyte count, platelets, red blood cells, and hematocrit) were decreased among exposed workers compared to controls, with the exception of the red blood cell mean corpuscular volume (MCV), which was higher among exposed subjects. In a subgroup of workers who were not exposed to more than 31 ppm benzene on any of 5 sampling days (n = 11, median 8 hr TWA = 7.6 ppm, range = 1-20 ppm), only the absolute lymphocyte count was significantly different between exposed workers (mean [sd]1.6 [0.4] x 10(3) mu L) and controls (1.9 [0.4] x l0(3) uL, p = 0.03). Among exposed subjects, a dose response relationship with various measures of current benzene exposure (i.e., personal air monitoring, benzene metabolites in urine) was present only for the total WBC count, the absolute lymphocyte count, and the MCV. Correlations between benzene metabolites and hematologic parameters were generally similar, although hydroquinone was somewhat more strongly associated with a decrease in the absolute lymphocyte count, and catechol was more strongly associated with an increase in MCV. Morphologic review of peripheral blood slides demonstrated an excess of red blood cell abnormalities (i.e., stomatocytes and target cells) only in the most heavily exposed workers, with no differences in granulocyte, lymphocyte, or platelet morphology noted. Although benzene can affect all the major peripheral blood elements, our results support the use of the absolute lymphocyte count as the most sensitive indicator of benzene-induced hematotoxicity.

1,2,4-Benzenetriol is an active metabolite of the human leukemogen benzene that reacts rapidly with molecular oxygen (O2). The mechanism of autoxidation of benzenetriol is scantily characterized, and little is known of the effects of metals, metal chelators, radical scavengers, and antioxidants on the rate of reduction of O2. Here, we report that catalytic amounts of Cu2+ and Fe3+ accelerated the oxidation of benzenetriol (250 mu M) in a dose-dependent manner. Fe3+ (50 mu M) increased the rate of autoxidation by 91%, and Cu2+ (10 mu M) increased it 11-fold. In the absence of added metals, superoxide dismutase inhibited and desferrioxamine stimulated the autoxidation. In the Cu2+ -catalyzed reaction, superoxide dismutase neither inhibited nor stimulated, while desferrioxamine abolished the catalysis by Cu2+. In the presence of Fe3+, superoxide dismutase slowed the reaction, but desferrioxamine, surprisingly, did not. The presence of both superoxide dismutase and desferrioxamine blocked the autoxidation, either in the presence or absence of metals. We conclude: (1) superoxide is a propagator of sequential one-electron transfer reactions in the absence of added metals; (2) addition of Cu2+, unlike Fe3+, removes the dependence of the reaction on propagation by superoxide, presumably changing the radical-propagated chain reaction to a concerted two-electron transfer; (3) the further addition of desferrioxamine restores superoxide-dependent propagation. Taken with our previous data on the genotoxicity of benzenetriol, these findings have implications regarding a role for transition metals in the carcinogenicity of benzene.

Benzene is known to have toxic effects on the blood and bone marrow, but its impact at levels below the U.S. occupational standard of 1 part per million (ppm) remains uncertain. In a study of 250 workers exposed to benzene, white blood cell and platelet counts were significantly lower than in 140 controls, even for exposure below 1 ppm in air. Progenitor cell colony formation significantly declined with increasing benzene exposure and was more sensitive to the effects of benzene than was the number of mature blood cells. Two genetic variants in key metabolizing enzymes, myeloperoxidase and NAD(P)H:quinone oxidoreductase, influenced susceptibility to benzene hematotoxicity. Thus, hematotoxicity from exposure to benzene occurred at air levels of 1 ppm or less and may be particularly evident among genetically susceptible subpopulations.

To evaluate the immunotoxicity of trichloroethylene (TCE), we conducted a cross-sectional molecular epidemiology study in China of workers exposed to TCE. We measured serum levels of IL-6, IL-10, and TNF-α, which play a critical role in regulating various components of the immune system, in 71 exposed workers and 78 unexposed control workers. Repeated personal exposure measurements were taken in workers before blood collection using 3 M organic vapor monitoring badges. Compared to unexposed workers, the serum concentration of IL-10 in workers exposed to TCE was decreased by 70% (P = 0.001) after adjusting for potential confounders. Further, the magnitude of decline in IL-10 was >60% and statistically significant in workers exposed to

Though current functional genomic screening systems are useful for investigating human susceptibility to chemical toxicity, they have limitations. Well-established, high-throughput yeast mutant screens identify only evolutionarily conserved processes. RNA interference can be applied in human cells but is limited by incomplete gene knockout and off-target effects. Human haploid cell screening is advantageous as it requires knockdown of only a single copy of each gene. A human haploid cell mutant library (KBM7-Mu), derived from a chronic myeloid leukemia (CML) patient, was recently developed and has been used to identify genes that modulate sensitivity to infectious agents and pharmaceutical drugs. Here, we sought to improve the KBM7-Mu screening process to enable efficient screening of environmental chemicals. We developed a semi-solid medium based screening approach that cultures individual mutant colonies from chemically resistant cells, faster (by 2–3 weeks) and with less labor than the original liquid medium-based approach. As proof of principle, we identified genetic mutants that confer resistance to the carcinogen formaldehyde (FA, 12 genes, 18 hits) and the CML chemotherapeutic agent imatinib (6 genes, 13 hits). Validation experiments conducted on KBM7 mutants lacking each of the 18 genes confirmed resistance of 6 FA mutants (CTC1, FCRLA, GOT1, LPR5, M1AP, and MAP2K5) and 1 imatinib-resistant mutant (LYRM9). Despite the improvements to the method, it remains technically challenging to limit false positive findings. Nonetheless, our findings demonstrate the broad applicability of this optimized haploid approach to screen toxic chemicals to identify novel susceptibility genes and gain insight into potential mechanisms of toxicity.

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Background Formaldehyde is used in many occupational settings, most notably in manufacturing, health care, and embalming. Formaldehyde has been classified as a human carcinogen, but its mechanism of action remains uncertain. Methods We carried out a cross-sectional study of 43 formaldehyde-exposed workers and 51 unexposed age and sex-matched controls in Guangdong, China to study formaldehyde's early biologic effects. To follow up our previous report that the total lymphocyte count was decreased in formaldehyde-exposed workers compared with controls, we evaluated each major lymphocyte subset (i.e., CD4+ T cells, CD8+ T cells, natural killer [NK] cells, and B cells) and T cell lymphocyte subset (CD4+ naive and memory T cells, CD8+ naive and memory T cells, and regulatory T cells). Linear regression of each subset was used to test for differences between exposed workers and controls, adjusting for potential confounders. Results Total NK cell and T cell counts were about 24% (P = 0.037) and 16% (P = 0.0042) lower, respectively, among exposed workers. Among certain T cell subsets, decreased counts among exposed workers were observed for CD8+ T cells (P = 0.026), CD8+ effector memory T cells (P = 0.018), and regulatory T cells (CD4+FoxP3+: P = 0.04; CD25+FoxP3+: P = 0.008). Conclusions Formaldehyde-exposed workers experienced decreased counts of NK cells, regulatory T cells, and CD8+ effector memory T cells; however, due to the small sample size; these findings need to be confirmed in larger studies. Am. J. Ind. Med. 56:252–257, 2013. © 2011 Wiley Periodicals, Inc.

Characterization of toxicogenomic signatures of carcinogen exposure holds significant promise for mechanistic and predictive toxicology. In vitro transcriptomic studies allow the comparison of the response to chemicals with diverse mode of actions under controlled experimental conditions. We conducted an in vitro study in TK6 cells to characterize gene expression signatures of exposure to 15 genotoxic carcinogens frequently used in European industries. We also examined the dose-responsive changes in gene expression, and perturbation of biochemical pathways in response to these carcinogens. TK6 cells were exposed at 3 dose levels for 24 h with and without S9 human metabolic mix. Since S9 had an impact on gene expression (885 genes), we analyzed the gene expression data from cells cultures incubated with S9 and without S9 independently. The ribosome pathway was affected by all chemical-dose combinations. However in general, no similar gene expression was observed among carcinogens. Further, pathways, i.e. cell cycle, DNA repair mechanisms, RNA degradation, that were common within sets of chemical-dose combination were suggested by clustergram. Linear trends in dose-response of gene expression were observed for Trichloroethylene, Benz[a]anthracene, Epichlorohydrin, Benzene, and Hydroquinone. The significantly altered genes were involved in the regulation of (anti-) apoptosis, maintenance of cell survival, tumor necrosis factor-related pathways and immune response, in agreement with several other studies. Similarly in S9+ cultures, Benz[a]pyrene, Styrene and Trichloroethylene each modified over 1000 genes at high concentrations. Our findings expand our understanding of the transcriptomic response to genotoxic carcinogens, revealing the alteration of diverse sets of genes and pathways involved in cellular homeostasis and cell cycle control. ispartof: PLoS One vol:7 issue:6 ispartof: location:United States status: published

The mechanism of toxicity of the leukemogen benzene is not entirely known. This pilot study used RNA-sequencing (RNA-seq) technology to examine the effect of benzene exposure on gene expression in peripheral blood mononuclear cells obtained from 10 workers occupationally exposed to high levels of benzene (≥5 ppm) in air and 10 matched unexposed control workers, from a large study (n = 125) in which gene expression was previously measured by microarray. RNA-seq is more sensitive and has a wider dynamic range for the quantification of gene expression. Further, it has the ability to detect novel transcripts and alternative splice variants. The main conclusions from our analysis of the 20 workers by RNA-seq are as follows: The Pearson correlation between the two technical replicates for the RNA-seq experiments was 0.98 and the correlation between RNA-seq and microarray signals for the 20 subjects was around 0.6. 60% of the transcripts with detected reads from the RNA-seq experiments did not have corresponding probes on the microarrays. Fifty-three percent of the transcripts detected by RNA-seq and 99% of those with probes on the microarray were protein-coding. There was a significant overlap (P < 0.05) in transcripts declared differentially expressed due to benzene exposure using the two technologies. About 20% of the transcripts declared differentially expressed using the RNA-seq data were non-coding transcripts. Six transcripts were determined (false-discovery rate < 0.05) to be alternatively spliced as a result of benzene exposure. Overall, this pilot study shows that RNA-seq can complement the information obtained by microarray in the analysis of changes in transcript expression from chemical exposures.

Fluorescence in situ hybridization (FISH) is a technique that allows specific DNA sequences to be detected on metaphase or interphase chromosomes in cell nuclei(1). The technique uses DNA probes with unique sequences that hybridize to whole chromosomes or specific chromosomal regions, and serves as a powerful adjunct to classic cytogenetics. For instance, many earlier studies reported the frequent detection of increased chromosome aberrations in leukemia patients related with benzene exposure, benzene-poisoning patients, and healthy workers exposed to benzene, using classic cytogenetic analysis(2). Using FISH, leukemia-specific chromosomal alterations have been observed to be elevated in apparently healthy workers exposed to benzene(3-6), indicating the critical roles of cytogentic changes in benzene-induced leukemogenesis. Generally, a single FISH assay examines only one or a few whole chromosomes or specific loci per slide, so multiple hybridizations need to be conducted on multiple slides to cover all of the human chromosomes. Spectral karyotyping (SKY) allows visualization of the whole genome simultaneously, but the requirement for special software and equipment limits its application(7). Here, we describe a novel FISH assay, OctoChrome-FISH, which can be applied for Chromosomics, which we define here as the simultaneous analysis of all 24 human chromosomes on one slide in human studies, such as chromosome-wide aneuploidy study (CWAS)(8). The basis of the method, marketed by Cytocell as the Chromoprobe Multiprobe System, is an OctoChrome device that is divided into 8 squares, each of which carries three different whole chromosome painting probes (Figure 1). Each of the three probes is directly labeled with a different colored fluorophore, green (FITC), red (Texas Red), and blue (Coumarin). The arrangement of chromosome combinations on the OctoChrome device has been designed to facilitate the identification of the non-random structural chromosome alterations (translocations) found in the most common leukemias and lymphomas, for instance t(9;22), t(15;17), t(8;21), t(14;18)(9). Moreover, numerical changes (aneuploidy) in chromosomes can be detected concurrently. The corresponding template slide is also divided into 8 squares onto which metaphase spreads are bound (Figure 2), and is positioned over the OctoChrome device. The probes and target DNA are denatured at high-temperature and hybridized in a humid chamber, and then all 24 human chromosomes can be visualized simultaneously. OctoChrome FISH is a promising technique for the clinical diagnosis of leukemia and lymphoma and for detection of aneuploidies in all chromosomes. We have applied this new Chromosomic approach in a CWAS study of benzene-exposed Chinese workers(8,10).