Employing a Toxic Aging Coin approach to assess hexavalent chromium (Cr[VI])-induced neurotoxic effects on behavior: Heads for age differences.

We are facing a rapidly growing geriatric population (65+) that will live for multiple decades and are challenged with environmental pollution far exceeding that of previous generations. Consequently, we currently have a poor understanding of how environmental pollution will impact geriatric health distinctly from younger populations. Few toxicology studies have considered age differences with geriatric individuals. Critically, all top ten most prevalent age-related diseases are linked to metal exposures. Hexavalent chromium [Cr(VI)] is a metal of major environmental health concern that can induce aging phenotypes and neurotoxicity. However, there are many knowledge gaps for Cr(VI) neurotoxicity, including how Cr(VI) impacts behavior. To address this, we exposed male rats across three ages (3-, 7-, and 18-months old) to Cr(VI) in drinking water (0, 0.05, 0.1 mg/L) for 90 days. These levels reflect the maximum contaminant levels determined by the World Health Organization (WHO) and the U.S. Environmental Protection Agency (US EPA). Here, we report how these Cr(VI) drinking water levels impacted rat behaviors using a battery of behavior tests, including grip strength, open field assay, elevated plus maze, Y-maze, and 3-chamber assay. We observed adult rats were the most affected age group and memory assays (spatial and social) exhibited the most significant effects. Critically, the significant effects were surprising as rats should be particularly resistant to these Cr(VI) drinking water levels due to the adjustments applied in risk assessment from rodent studies to human safety, and because rats endogenously synthesize vitamin C in their livers (vitamin C is a primary reducer of Cr[VI] to Cr[III]). Our results emphasize the need to broaden the scope of toxicology research to consider multiple life stages and suggest the current regulations for Cr(VI) in drinking water need to be revisited.

Clonal variability in chromosomal instability as a potential driver in the acquisition of tumorigenic phenotype in chronic arsenic-exposed and hsa-miR-186 overexpressing human keratinocytes.

Chronic arsenic exposure through drinking water is a global health issue, affecting >200 million people. Arsenic is a group I human carcinogen and causes chromosomal instability (CIN). Arsenic exposure is the second most common cause of skin cancer after UV radiation. hsa-miR-186 is overexpressed in arsenic-induced squamous cell carcinoma relative to premalignant hyperkeratosis. Among predicted targets of hsa-miR-186 are cell cycle regulators including regulators of mitotic progression. Disruption of mitotic progression can contribute to CIN. Thus, we hypothesized that hsa-miR-186 overexpression contributes to malignant transformation of arsenic exposed HaCaT cells by induction of CIN. Stable clones of HaCaT cells transfected with pEP-hsa-miR-186 expression vector or empty vector were maintained under puromycin selection and exposed to 0 or 100 nM NaAsO2 and cultured for 29 weeks. HaCaT clones overexpressing hsa-miR-186 and exposed to NaAsO2 showed increased CIN and anchorage independent growth at 29 weeks in a stochastic manner, in contrast to unexposed empty vector transfected clones. These results suggest that clonal variability mediates arsenic-induced carcinogenesis in hsa-miR-186 overexpressing human keratinocytes.

Prolonged particulate hexavalent chromium exposure induces RAD51 foci inhibition and cytoplasmic accumulation in immortalized and primary human lung bronchial epithelial cells.

Hexavalent chromium [Cr(VI)] is a human lung carcinogen with widespread exposure risks. Cr(VI) causes DNA double strand breaks that if unrepaired, progress into chromosomal instability (CIN), a key driving outcome in Cr(VI)-induced tumors. The ability of Cr(VI) to cause DNA breaks and inhibit repair is poorly understood in human lung epithelial cells, which are extremely relevant since pathology data show Cr(VI)-induced tumors originate from bronchial epithelial cells. In the present study, we considered immortalized and primary human bronchial epithelial cells. Cells were treated with zinc chromate at concentrations ranging 0.05 to 0.4µg/cm2 for acute (24 h) and prolonged (120 h) exposures. DNA double strand breaks (DSBs) were measured by neutral comet assay and the status of homologous recombination repair, the main pathway to fix Cr(VI)-induced DSBs, was measured by RAD51 foci formation with immunofluorescence, RAD51 localization with confocal microscopy and sister chromatid exchanges. We found acute and prolonged Cr(VI) exposure induced DSBs. Acute exposure induced homologous recombination repair, but prolonged exposure inhibited it resulting in chromosome instability in immortalized and primary human bronchial epithelial cells.

miR-186 induces tetraploidy in arsenic exposed human keratinocytes.

Chronic inorganic arsenic (iAs) exposure in drinking water is a global issue affecting >225 million people. Skin is a major target organ for iAs. miRNA dysregulation and chromosomal instability (CIN) are proposed mechanisms of iAs-induced carcinogenesis. CIN is a cancer hallmark and tetraploid cells can better tolerate increase in chromosome number and aberration, contributing to the evolution of CIN. miR-186 is overexpressed in iAs-induced squamous cell carcinoma relative to iAs-induced hyperkeratosis. Bioinformatic analysis indicated that miR-186 targets mRNAs of important cell cycle regulators including mitotic checkpoint serine/threonine kinase B (BUB1) and cell division cycle 27 (CDC27). We hypothesized that miR-186 overexpression contributes to iAs-induced transformation of keratinocytes by targeting mitotic regulators leading to induction of CIN. Ker-CT cells, a near diploid human keratinocyte cell line, were transduced with miR-186 overexpressing or scrambled control lentivirus. Stable clones were isolated after puromycin selection. Clones transduced with lentivirus expressing either a scrambled control miRNA or miR-186 were maintained with 0 or 100 nM iAs for 4 weeks. Unexposed scrambled control clones were considered as passage matched controls. Chronic iAs exposure increased miR-186 expression in miR-186 clones. miR-186 overexpression significantly reduced CDC27 levels irrespective of iAs exposure. The percentage of tetraploid or aneuploid cells was increased in iAs exposed miR-186 clones. Aneuploidy can arise from a tetraploid intermediate. Suppression of CDC27 by miR-186 may lead to impairment of mitotic checkpoint complex formation and its ability to maintain cell cycle arrest leading to chromosome misalignment. As a result, cells overexpressing miR-186 and chronically exposed to iAs may have incorrect chromosome segregation and CIN. These data suggest that dysregulation of miRNA by iAs mediates tetraploidy, aneuploidy and chromosomal instability contributing to iAs-induced carcinogenesis.

Chromium distribution in an oropharyngeal aspiration model for hexavalent chromium in rats.

Hexavalent chromium [Cr(VI)] is a well-known and widespread environmental contaminant associated with a variety of adverse health effects, in particular lung cancer. The primary route of exposure in humans is through inhalation. Particulate forms of Cr(VI) are the most potent but in vivo studies are difficult. Intratracheal instillation requires highly trained surgical procedures which also limits the number of repeated exposures possible and thus requires high doses. Inhalation studies can deliver lower more chronic doses but are expensive and generate dangerous aerosols. We evaluated an oropharyngeal aspiration exposure route for zinc chromate particles in Wistar rats. Animals were treated once per week for 90 days. We found chromium accumulated in the lungs, blood, and reproductive tissues of all treated animals. Additionally, we found inflammatory indicators in the lung were elevated and circulating lymphocytes had increased chromosomal damage. These results show oropharyngeal aspiration provides a practicable exposure route for chronic and sub-chronic exposures of Cr(VI) particles.

Overexpression of hsa-miR-186 induces chromosomal instability in arsenic-exposed human keratinocytes.

The mechanism of arsenic-induced skin carcinogenesis is not yet fully understood. Chromosomal instability contributes to aneuploidy and is a driving force in carcinogenesis. Arsenic causes mitotic arrest and induces aneuploidy. hsa-miR-186 overexpression is associated with metastatic cancers as well as arsenic-induced squamous cell carcinoma and is reported to target several mitotic regulators. Decreased levels of these proteins can dysregulate chromatid segregation contributing to aneuploidy. This work investigates the potential aneuploidogenic role of hsa-miR-186 in arsenic carcinogenesis. Clones of immortalized human keratinocytes (HaCaT) stably transfected with a hsa-miR-186 expression or empty vector were isolated. Three clones with high and low hsa-miR-186 expression determined by RT-qPCR were selected for further analysis and cultured with 0 or 100 nM NaAsO2 for 8 weeks. Analysis of mitoses revealed that chromosome number and structural abnormalities increased in cells overexpressing hsa-miR-186 and were further increased by arsenite exposure. Double minutes were the dominant structural aberrations. The peak number of chromosomes also increased. Cells with >220 to >270 chromosomes appeared after 2 months in hsa-miR-186 overexpressing cells, indicating multiple rounds of endomitosis had occurred. The fraction of cells with increased chromosome number or structural abnormalities did not increase in passage matched control cells. Levels of selected target proteins were determined by western blot. Expression of BUB1, a predicted hsa-miR-186 target was suppressed in hsa-miR-186 overexpressing clones, but increased with arsenite exposure. CDC27 remained constant under all conditions. These results suggest that overexpression of miR-186 in arsenic exposed tissues likely induces aneuploidy contributing to arsenic-induced carcinogenesis.

A comparison of particulate hexavalent chromium cytotoxicity and genotoxicity in human and leatherback sea turtle lung cells from a one environmental health perspective.

Evaluating health risks of environmental contaminants can be better achieved by considering toxic impacts across species. Hexavalent chromium [Cr(VI)] is a marine pollutant and global environmental contaminant. While Cr(VI) has been identified as a human lung carcinogen, health effects in marine species are poorly understood. Little is known about how Cr(VI) might impact humans and marine species differently. This study used a One Environmental Health Approach to compare the cytotoxicity and genotoxicity of particulate Cr(VI) in human and leatherback sea turtle (Dermochelys coriacea) lung fibroblasts. Leatherbacks may experience prolonged exposures to environmental contaminants and provide insight to how environmental exposures affect health across species. Since humans and leatherbacks may experience prolonged exposure to Cr(VI), and prolonged Cr(VI) exposure leads to carcinogenesis in humans, in this study we considered both acute and prolonged exposures. We found particulate Cr(VI) induced cytotoxicity in leatherback cells comparable to human cell data supporting current research that shows Cr(VI) impacts health across species. To better understand mechanisms of Cr(VI) toxicity we assessed the genotoxic effects of particulate Cr(VI) in human and leatherback cells. Particulate Cr(VI) induced similar genotoxicity in both cell lines, however, human cells arrested at lower concentrations than leatherback cells. We also measured intracellular Cr ion concentrations and found after prolonged exposure human cells accumulated more Cr than leatherback cells. These data indicate Cr(VI) is a health concern for humans and leatherbacks. The data also suggest humans and leatherbacks respond to chemical exposure differently, possibly leading to the discovery of species-specific protective mechanisms.

Hexavalent chromium induces chromosome instability in human urothelial cells.

Numerous metals are well-known human bladder carcinogens. Despite the significant occupational and public health concern of metals and bladder cancer, the carcinogenic mechanisms remain largely unknown. Chromium, in particular, is a metal of concern as incidences of bladder cancer have been found elevated in chromate workers, and there is an increasing concern for patients with metal hip implants. However, the impact of hexavalent chromium (Cr(VI)) on bladder cells has not been studied. We compared chromate toxicity in two bladder cell lines; primary human urothelial cells and hTERT-immortalized human urothelial cells. Cr(VI) induced a concentration- and time-dependent increase in chromosome damage in both cell lines, with the hTERT-immortalized cells exhibiting more chromosome damage than the primary cells. Chronic exposure to Cr(VI) also induced a concentration-dependent increase in aneuploid metaphases in both cell lines which was not observed after a 24h exposure. Aneuploidy induction was higher in the hTERT-immortalized cells. When we correct for uptake, Cr(VI) induces a similar amount of chromosome damage and aneuploidy suggesting that the differences in Cr(VI) sensitivity between the two cells lines were due to differences in uptake. The increase in chromosome instability after chronic chromate treatment suggests this may be a mechanism for chromate-induced bladder cancer, specifically, and may be a mechanism for metal-induced bladder cancer, in general.

Disruptive chemicals, senescence and immortality.

Carcinogenesis is thought to be a multistep process, with clonal evolution playing a central role in the process. Clonal evolution involves the repeated 'selection and succession' of rare variant cells that acquire a growth advantage over the remaining cell population through the acquisition of 'driver mutations' enabling a selective advantage in a particular micro-environment. Clonal selection is the driving force behind tumorigenesis and possesses three basic requirements: (i) effective competitive proliferation of the variant clone when compared with its neighboring cells, (ii) acquisition of an indefinite capacity for self-renewal, and (iii) establishment of sufficiently high levels of genetic and epigenetic variability to permit the emergence of rare variants. However, several questions regarding the process of clonal evolution remain. Which cellular processes initiate carcinogenesis in the first place? To what extent are environmental carcinogens responsible for the initiation of clonal evolution? What are the roles of genotoxic and non-genotoxic carcinogens in carcinogenesis? What are the underlying mechanisms responsible for chemical carcinogen-induced cellular immortality? Here, we explore the possible mechanisms of cellular immortalization, the contribution of immortalization to tumorigenesis and the mechanisms by which chemical carcinogens may contribute to these processes.

Hexavalent chromium is cytotoxic and genotoxic to hawksbill sea turtle cells.

Sea turtles are a charismatic and ancient ocean species and can serve as key indicators for ocean ecosystems, including coral reefs and sea grass beds as well as coastal beaches. Genotoxicity studies in the species are absent, limiting our understanding of the impact of environmental toxicants on sea turtles. Hexavalent chromium (Cr(VI)) is a ubiquitous environmental problem worldwide, and recent studies show it is a global marine pollutant of concern. Thus, we evaluated the cytotoxicity and genotoxicity of soluble and particulate Cr(VI) in hawksbill sea turtle cells. Particulate Cr(VI) was both cytotoxic and genotoxic to sea turtle cells. Concentrations of 0.1, 0.5, 1, and 5µg/cm(2) lead chromate induced 108, 79, 54, and 7% relative survival, respectively. Additionally, concentrations of 0, 0.1, 0.5, 1, and 5µg/cm(2) lead chromate induced damage in 4, 10, 15, 26, and 36% of cells and caused 4, 11, 17, 30, and 56 chromosome aberrations in 100 metaphases, respectively. For soluble Cr, concentrations of 0.25, 0.5, 1, 2.5, and 5µM sodium chromate induced 84, 69, 46, 25, and 3% relative survival, respectively. Sodium chromate induced 3, 9, 9, 14, 21, and 29% of metaphases with damage, and caused 3, 10, 10, 16, 26, and 39 damaged chromosomes in 100 metaphases at concentrations of 0, 0.25, 0.5, 1, 2.5, and 5µM sodium chromate, respectively. These data suggest that Cr(VI) may be a concern for hawksbill sea turtles and sea turtles in general.

Concentrations of the genotoxic metals, chromium and nickel, in whales, tar balls, oil slicks, and released oil from the gulf of Mexico in the immediate aftermath of the deepwater horizon oil crisis: is genotoxic metal exposure part of the deepwater horizon legacy?

Concern regarding the Deepwater Horizon oil crisis has largely focused on oil and dispersants while the threat of genotoxic metals in the oil has gone largely overlooked. Genotoxic metals, such as chromium and nickel, damage DNA and bioaccumulate in organisms, resulting in persistent exposures. We found chromium and nickel concentrations ranged from 0.24 to 8.46 ppm in crude oil from the riser, oil from slicks on surface waters and tar balls from Gulf of Mexico beaches. We found nickel concentrations ranged from 1.7 to 94.6 ppm wet weight with a mean of 15.9 ± 3.5 ppm and chromium concentrations ranged from 2.0 to 73.6 ppm wet weight with a mean of 12.8 ± 2.6 ppm in tissue collected from Gulf of Mexico whales in the wake of the crisis. Mean tissue concentrations were significantly higher than those found in whales collected around the world prior to the spill. Given the capacity of these metals to damage DNA, their presence in the oil, and their elevated concentrations in whales, we suggest that metal exposure is an important understudied concern for the Deepwater Horizon oil disaster.

Prolonged Particulate Hexavalent Chromium Exposure Induces DNA Double-Strand Breaks and Inhibits Homologous Recombination Repair in Primary Rodent Lung Cells.

Hexavalent chromium [Cr(VI)] is a known lung carcinogen and a driving mechanism in human lung cells for Cr(VI)-induced lung cancer is chromosome instability, caused by prolonged Cr(VI) exposure inducing DNA double-strand breaks, while simultaneously inhibiting the repair of these breaks. In North Atlantic right whales, Cr(VI) induces breaks but does not inhibit repair. It is unclear if this repair inhibition is specific to human lung cells or occurs in other species, as it has only been considered in humans and North Atlantic right whales. We evaluated these outcomes in rodent cells, as rodents are an experimental model for metal-induced lung carcinogenesis. We used a guinea pig lung fibroblast cell line, JH4 Clone 1, and rat lung fibroblasts. Cells were exposed to two different particulate Cr(VI) compounds, ranging from 0 to 0.5 ug/cm2, for 24 or 120 h and assessed for cytotoxicity, DNA double-strand breaks, and DNA double-strand break repair. Both particulate Cr(VI) compounds induced a concentration-dependent increase in cytotoxicity and DNA double-strand breaks after acute and prolonged exposures. Notably, while the repair of Cr(VI)-induced DNA double-strand breaks increased after acute exposure, the repair of these breaks was inhibited after prolonged exposure. These results are consistent with outcomes in human lung cells indicating rodent cells respond like human cells, while whale cells have a markedly different response.

A whale of a tale: whale cells evade the driving mechanism for hexavalent chromium-induced chromosome instability.

Chromosome instability, a hallmark of lung cancer, is a driving mechanism for hexavalent chromium [Cr(VI)] carcinogenesis in humans. Cr(VI) induces structural and numerical chromosome instability in human lung cells by inducing DNA double-strand breaks and inhibiting homologous recombination repair and causing spindle assembly checkpoint (SAC) bypass and centrosome amplification. Great whales are long-lived species with long-term exposures to Cr(VI) and accumulate Cr in their tissue, but exhibit a low incidence of cancer. Data show Cr(VI) induces fewer chromosome aberrations in whale cells after acute Cr(VI) exposure suggesting whale cells can evade Cr(VI)-induced chromosome instability. However, it is unknown if whales can evade Cr(VI)-induced chromosome instability. Thus, we tested the hypothesis that whale cells resist Cr(VI)-induced loss of homologous recombination repair activity and increased SAC bypass and centrosome amplification. We found Cr(VI) induces similar amounts of DNA double-strand breaks after acute (24 h) and prolonged (120 h) exposures in whale lung cells, but does not inhibit homologous recombination repair, SAC bypass, or centrosome amplification, and does not induce chromosome instability. These data indicate whale lung cells resist Cr(VI)-induced chromosome instability, the major driver for Cr(VI) carcinogenesis at a cellular level, consistent with observations that whales are resistant to cancer.

Acute particulate hexavalent chromium exposure induces DNA double strand breaks and activates homologous recombination repair in rat lung tissue.

Hexavalent chromium [Cr(VI)] is an established human lung carcinogen, but the carcinogenesis mechanism is poorly understood. Chromosome instability, a hallmark of lung cancer, is considered a major driver of Cr(VI)-induced lung cancer. Unrepaired DNA double strand breaks are the underlying cause, and homologous recombination repair is the primary mechanism preventing Cr(VI)-induced DNA breaks from causing chromosome instability. Cell culture studies show acute Cr(VI) exposure causes DNA double strand breaks and increases homologous recombination repair activity. However, the ability of Cr(VI)-induced DNA breaks and repair impact has only been reported in cell culture studies. Therefore, we investigated whether acute Cr(VI) exposure could induce breaks and homologous recombination repair in rat lungs. Male and female Wistar rats were acutely exposed to either zinc chromate particles in a saline solution or saline alone by oropharyngeal aspiration. This exposure route resulted in increased Cr levels in each lobe of the lung. We found Cr(VI) induced DNA double-strand breaks in a concentration-dependent manner, with females being more susceptible than males, and induced homologous recombination repair at similar levels in both sexes. Thus, these data show this driving mechanism discovered in cell culture indeed translates to lung tissue in vivo.

Chromium and genomic stability.

Many metals serve as micronutrients which protect against genomic instability. Chromium is most abundant in its trivalent and hexavalent forms. Trivalent chromium has historically been considered an essential element, though recent data indicate that while it can have pharmacological effects and value, it is not essential. There is no data indicating that trivalent chromium promotes genomic stability and, instead may promote genomic instability. Hexavalent chromium is widely accepted as highly toxic and carcinogenic with no nutritional value. Recent data indicate that it causes genomic instability and also has no role in promoting genomic stability.

Protein tyrosine phosphatase (PTP) inhibition enhances chromosomal stability after genotoxic stress: decreased chromosomal instability (CIN) at the expense of enhanced genomic instability (GIN)?

Inappropriate survival signaling after DNA damage may facilitate clonal expansion of genetically compromised cells, and it is known that protein tyrosine phosphatase (PTP) inhibitors activate key survival pathways. In this study we employed the genotoxicant, hexavalent chromium [Cr(VI)], which is a well-documented carcinogen of occupational and environmental concern. Cr(VI) induces a complex array of DNA damage, including DNA double strand breaks (DSBs). We recently reported that PTP inhibition bypassed cell cycle arrest and abrogated Cr(VI)-induced clonogenic lethality. Notably, PTP inhibition resulted in an increase in forward mutations at the HPRT locus, supporting the hypothesis that PTP inhibition in the presence of DNA damage may lead to genomic instability (GIN), via cell cycle checkpoint bypass. The aim of the present study was to determine the effect of PTP inhibition on DNA DSB formation and chromosomal integrity after Cr(VI) exposure. Diploid human lung fibroblasts were treated with Cr(VI) in the presence or absence of the PTP inhibitor, sodium orthovanadate, for up to 24h, and cells were analyzed for DNA DSBs and chromosomal damage. Cr(VI) treatment induced a rapid increase in DNA DSBs, and a significant increase in total chromosomal damage (chromatid breaks and gaps) after 24h. In sharp contrast, PTP inhibition abrogated both DNA DSBs and chromosomal damage after Cr(VI) treatment. In summary, PTP inhibition in the face of Cr(VI) genotoxic stress decreases chromosomal instability (CIN) but increases mutagenesis, which we postulate to be a result of error-prone DNA repair.

Characterization of a castrate-resistant prostate cancer xenograft derived from a patient of West African ancestry.

BACKGROUND: Prostate cancer is a clinically and molecularly heterogeneous disease, with highest incidence and mortality among men of African ancestry. To date, prostate cancer patient-derived xenograft (PCPDX) models to study this disease have been difficult to establish because of limited specimen availability and poor uptake rates in immunodeficient mice. Ancestrally diverse PCPDXs are even more rare, and only six PCPDXs from self-identified African American patients from one institution were recently made available. METHODS: In the present study, we established a PCPDX from prostate cancer tissue from a patient of estimated 90% West African ancestry with metastatic castration resistant disease, and characterized this model's pathology, karyotype, hotspot mutations, copy number, gene fusions, gene expression, growth rate in normal and castrated mice, therapeutic response, and experimental metastasis. RESULTS: This PCPDX has a mutation in TP53 and loss of PTEN and RB1. We have documented a 100% take rate in mice after thawing the PCPDX tumor from frozen stock. The PCPDX is castrate- and docetaxel-resistant and cisplatin-sensitive, and has gene expression patterns associated with such drug responses. After tail vein injection, the PCPDX tumor cells can colonize the lungs of mice. CONCLUSION: This PCPDX, along with others that are established and characterized, will be useful pre-clinically for studying the heterogeneity of prostate cancer biology and testing new therapeutics in models expected to be reflective of the clinical setting.

Polycyclic Aromatic Hydrocarbon-DNA Adducts in Gulf of Mexico Sperm Whale Skin Biopsies Collected in 2012.

The northern Gulf of Mexico has a long history of polycyclic aromatic hydrocarbon (PAH) contamination from anthropogenic activities, natural oil seepages, and the 2010 Deepwater Horizon explosion and oil spill. The continental shelf of the same area is a known breeding ground for sperm whales (Physeter macrocephalus). To evaluate PAH-DNA damage, a biomarker for potential cancer risk, we compared skin biopsies collected from Gulf of Mexico sperm whales in 2012 with skin biopsies collected from sperm whales in areas of the Pacific Ocean in 1999-2001. All samples were obtained by crossbow and comprised both epidermis and subcutaneous blubber. To evaluate exposure, 7 carcinogenic PAHs were analyzed in lipids extracted from Pacific Ocean sperm whale blubber, pooled by sex, and location. To evaluate PAH-DNA damage, portions of all tissue samples were formalin-fixed, paraffin-embedded, sectioned, and examined for PAH-DNA adducts by immunohistochemistry (IHC) using an antiserum elicited against benzo[a]pyrene-modified DNA, which crossreacts with several high molecular weight carcinogenic PAHs bound to DNA. The IHC showed widespread epidermal nuclear localization of PAH-DNA adducts in the Gulf of Mexico whales (n = 15) but not in the Pacific Ocean whales (n = 4). A standard semiquantitative scoring system revealed significantly higher PAH-DNA adducts in the Gulf of Mexico whales compared to the whales from the Pacific Ocean study (p = .0002).

Aneuploidy as an early mechanistic event in metal carcinogenesis.

Aneuploidy has recently been proposed as an initiating event for carcinogenesis. There is significant evidence that carcinogenic metals induce aneuploidy. Here we review the mechanisms for how carcinogenic metals may induce aneuploidy and the evidence that carcinogenic metals induce an aneugenic effect which can destabilize the genome leading to genomic instability and cancer.

Chronic exposure to zinc chromate induces centrosome amplification and spindle assembly checkpoint bypass in human lung fibroblasts.

Hexavalent chromium (Cr(VI)) compounds are known human lung carcinogens. Solubility plays an important role in its carcinogenicity with the particulate or insoluble form being the most potent. Of the particulate Cr(VI) compounds, zinc chromate appears to be the most potent carcinogen; however, very few studies have investigated its carcinogenic mechanism. In this study, we investigated the ability of chronic exposure to zinc chromate to induce numerical chromosome instability. We found no increase in aneuploidy after a 24 h exposure to zinc chromate, but with more chronic exposures, zinc chromate induced concentration- and time-dependent increases in aneuploidy in the form of hypodiploidy, hyperdiploidy, and tetraploidy. Zinc chromate also induced centrosome amplification in a concentration- and time-dependent manner in both interphase and mitotic cells after chronic exposure, producing cells with centriolar defects. Furthermore, chronic exposure to zinc chromate induced concentration- and time-dependent increases in spindle assembly checkpoint bypass with increases in centromere spreading, premature centromere division, and premature anaphase. Last, we found that chronic exposure to zinc chromate induced a G2 arrest. All together, these data indicate that zinc chromate can induce chromosome instability after prolonged exposures.