Mutations In Germ Cells Research Articles

Germline mutations and blood malignancy (Review).

Germline mutations are congenital genetic mutations in germ cells that originate from sperm or ovum and are generally incorporated into every cell of the offspring's body. Somatic mutations are acquired genetic mutations that form under the influence of environmental factors during embryo formation and epigenetic development. Generally, only a portion of the cells in the human body have the same somatic mutations. Clinical detection of germline mutations is intended to determine inherited malignancies and identify high‑risk families, and detection of somatic mutation is proposed to find targeted drugs, monitor tumor loading for guided therapy, and evaluate prognosis. Large‑scale population cohort studies have shown that germline mutations are closely related to the occurrence, development, and prognosis of diseases. Patients with cancer‑predisposition germline mutations can be used as sentinels in high‑risk families. Traditional histopathology is no longer enough to identify types of cancers. Even within a particular type of tumor, there is great heterogeneity between internal molecules. The Pan‑Cancer Research Program as well as other projects seek to use large quantities of data from different types of tumor research databases to carry out integrated analysis in order to establish potential non‑tumor‑specific tumor markers and targets by increasing the sample size to identify more molecular mechanisms. This review intends to summarize some of the relevant mechanisms underlying germline mutations in blood disorders.

Measuring mutagenicity in ecotoxicology: A case study of Cd exposure in Chironomus riparius.

Existing mutagenicity tests for metazoans lack the direct observation of enhanced germline mutation rates after exposure to anthropogenic substances, therefore being inefficient. Cadmium (Cd) is a metal described as a mutagen in mammalian cells and listed as a group 1 carcinogenic and mutagenic substance. But Cd mutagenesis mechanism is not yet clear. Therefore, in the present study, we propose a method coupling short-term mutation accumulation (MA) lines with subsequent whole genome sequencing (WGS) and a dedicated data analysis pipeline to investigate if chronic Cd exposure on Chironomus riparius can alter the rate at which de novo point mutations appear. Results show that Cd exposure did not affect the basal germline mutation rate nor the mutational spectrum in C.riparius, thereby arguing that exposed organisms might experience a range of other toxic effects before any mutagenic effect may occur. We show that it is possible to establish a practical and easily implemented pipeline to rapidly detect germ cell mutagens in a metazoan test organism. Furthermore, our data implicate that it is questionable to transfer mutagenicity assessments based on invitro methods to complex metazoans.

Heritable hazards of smoking: Applying the "clean sheet" framework to further science and policy.

All the cells in our bodies are derived from the germ cells of our parents, just as our own germ cells become the bodies of our children. The integrity of the genetic information inherited from these germ cells is of paramount importance in establishing the health of each generation and perpetuating our species into the future. There is a large and growing body of evidence strongly suggesting the existence of substances that may threaten this integrity by acting as human germ cell mutagens. However, there generally are no absolute regulatory requirements to test agents for germ cell effects. In addition, the current regulatory testing paradigms do not evaluate the impacts of epigenetically mediated intergenerational effects, and there is no regulatory framework to apply new and emerging tests in regulatory decision making. At the 50th annual meeting of the Environmental Mutagenesis and Genomics Society held in Washington, DC, in September 2019, a workshop took place that examined the heritable effects of hazardous exposures to germ cells, using tobacco smoke as the example hazard. This synopsis provides a summary of areas of concern regarding heritable hazards from tobacco smoke exposures identified at the workshop and the value of the Clean Sheet framework in organizing information to address knowledge and testing gaps.

Wild black rats (Rattus rattus Linnaeus, 1758) as zoomonitor of genotoxicity and systemic toxicity induced by hazardous emissions from Abule Egba unsanitary landfill, Lagos, Nigeria.

Wild black rats (Rattus rattus) inhabiting Abule Egba landfill (AEL) were used as zoomonitor to assess health risk associated with exposure to hazardous chemicals from landfills. Twenty five R. rattus (16♂ and 9♀) captured within AEL and 15 (9♂ and 6♀) (control) caught from Iyano Ipaja (10 km away from AEL) were examined for bone marrow micronucleated polychromatic erythrocytes (MNPCE) and polychromatic erythrocytes/normochromatic erythrocytes (PCE/NCE) ratio, abnormal sperm morphology, alterations in hematological indices and erythrocyte morphology, and histopathology of the viscera using standard protocols. There was significant (p < 0.05) increase in MNPCE but decrease PCE/NCE ratio in bone marrow cells of exposed rats than the reference site. MNPCE was insignificantly higher in male than females. Cauda epididymal sperms from exposed rats showed significant high frequency of teratozoospermia. Erythrocyte count, hemoglobin concentrations, percentage hematocrits, mean corpuscular hemoglobin concentrations, leucocyte count, and lymphocytes decreased while mean corpuscular volume, neutrophils, and mean corpuscular hemoglobin increased in the exposed rats compared to the control. Also, abnormal erythrocyte morphology: acanthocytes, codocytes (target cells), schizocytes, and tear drops significantly increased in the exposed rats. Marginal sexual dimorphism was observed between males and females in the incidence of hematological indices. Histopathological lesions including interstitial edema, hemorrhage, lymphoid depletion, cellular infiltrations, proliferation of the alveolar pneumocytes, necrosis, tissue degeneration, and reduced germinal epithelium were observed in the testes, liver, lungs, heart, kidney, and spleen from the exposed rats compared to the control. Some physicochemicals and metals analyzed in leachates from the landfill are capable of inducing genome instability and systemic toxicity in the exposed rats. Rattus rattus exposed to hazardous chemicals from AEL harbored somatic and germ cell mutations, and tissue damage compared to the control rats. We suggest that R. rattus are useful sentinel for genotoxicity and system toxicity assessment of landfill-polluted sites.Graphical abstract.

A germ cell-specific ageing pattern in otherwise healthy men.

Life‐long sperm production leads to the assumption that male fecundity remains unchanged throughout life. However, recently it was shown that paternal age has profound consequences for male fertility and offspring health. Paternal age effects are caused by an accumulation of germ cell mutations over time, causing severe congenital diseases. Apart from these well‐described cases, molecular patterns of ageing in germ cells and their impact on DNA integrity have not been studied in detail. In this study, we aimed to assess the effects of ‘pure’ ageing on male reproductive health and germ cell quality. We assembled a cohort of 198 healthy men (18–84 years) for which end points such as semen and hormone profiles, sexual health and well‐being, and sperm DNA parameters were evaluated. Sperm production and hormonal profiles were maintained at physiological levels over a period of six decades. In contrast, we identified a germ cell‐specific ageing pattern characterized by a steady increase of telomere length in sperm and a sharp increase in sperm DNA instability, particularly after the sixth decade. Importantly, we found sperm DNA methylation changes in 236 regions, mostly nearby genes associated with neuronal development. By in silico analysis, we found that 10 of these regions are located in loci which can potentially escape the first wave of genome‐wide demethylation after fertilization. In conclusion, human male germ cells present a unique germline‐specific ageing process, which likely results in diminished fecundity in elderly men and poorer health prognosis for their offspring.

Doxorubicine and doxorubicine hydrochloride – inhalable fraction. Documentation of proposed values of occupational exposure limits (OELs)

Doxorubicin (CAS: 23214-92-8) and its hydrochloride (CAS: 25136 40 9) are organic chemicals soluble in water. It is a cytostatic drug from the group of anthracycline antibiotics, used in antimitotic antitumor chemotherapy, primarily by intravenous, intravesical, and also in the case of lung cancer in the form of an aerosol for inhalation. In Poland, according to data from the Central Data Register on Exposure to Chemicals, Mixtures Thereof, Factors or Technological Processes with Carcinogenic or Mutagenic Effect, conducted at the Institute of Occupational Medicine in Łódź, the number of people exposed to doxorubicin and its hydrochloride in 2016 totaled 587. Administration of doxorubicin or its hydrochloride to patients at therapeutic doses may lead to myelosuppression, cardiomyopathy and myocardial fibrosis as well as neurotoxicity. Adverse effects of doxorubicin administration included cough, shortness of breath, chest pain, wheezing, hoarseness, hemoptysis, and bronchospasm. Systemic toxicity was defined as mild and transient and included sore throat, anorexia, dysgeusia, fatigue, nausea, tongue pain, tachycardia. Doxorubicin manufacturers state in their safety data sheets that inhalation of dust or aerosol is hazardous to health, may cause discomfort and nuisance, nausea, vomiting, bone marrow suppression, stomatitis, hair loss, and cardiotoxicity. Animal carcinogenicity studies have shown that doxorubicin was carcinogenic to rats after intravenous and subcutaneous administration, mainly causing mammary gland tumors. Doxorubicin has been shown to have genotoxic effects on somatic and embryonic mouse cells. Doxorubicin is toxic for reproduction. It may damage fertility and the unborn child. In Poland and in other countries, the highest permissible concentrations of doxorubicin and its hydrochloride in the work environment have not yet been determined. Occupational exposure limits are recommended by its manufacturers: FormuMax Scientific, Inc. and Pfizer at 0.0005 mg/m3. It was proposed to set up the MAC value for doxorubicin and its hydrochloride at the equivalent concentration level up to 0.1% of the lowest inhalational therapeutic dose found in the literature Dw = 0.04 mg/kg, i.e., 0.0003 mg/m3 – inhalable fraction. There are no substantive grounds to determine the STEL value. It is recommended to label the substance with the notation “skin” – the absorption of the substance through the skin may be just as important as when inhaled. The letters “Ft” should also be used – toxic for reproduction, Carc. 1B – carcinogen category 1B and Muta. 1B – germ cell mutagen category 1B. This article discusses the problems of occupational safety and health, which are covered by health sciences and environmental engineering.

Classified Chemicals in Accordance with the Globally Harmonized System of Classification and Labeling of Chemicals: Comparison of Lists of the European Union, Japan, Malaysia and New Zealand.

BackgroundThe Globally Harmonized System of Classification and Labeling of Chemicals (GHS) was developed to enhance chemical classification and hazard communication systems worldwide. However, some of the elements such as building blocks and data sources have the potential to cause “disharmony” to the GHS, particularly in its classification results. It is known that some countries have developed their own lists of classified chemicals in accordance with the GHS to “standardize” the classification results within their respective countries. However, the lists of classified chemicals may not be consistent among these countries.MethodIn this study, the lists of classified chemicals developed by the European Union, Japan, Malaysia, and New Zealand were selected for comparison of classification results for carcinogenicity, germ cell mutagenicity, and reproductive toxicity.ResultsThe findings show that only 54%, 66%, and 37% of the classification results for each Carcinogen, Mutagen and Reproductive toxicants hazard classes, respectively are the same among the selected countries. This indicates a “moderate” level of consistency among the classified chemicals lists.ConclusionBy using classification results for the carcinogenicity, germ cell mutagenicity, and reproductive toxicity hazard classes, this study demonstrates the “disharmony” in the classification results among the selected countries. We believe that the findings of this study deserve the attention of the relevant international bodies.

Assessment of anilofos-induced mutagenicity in bone marrow and germ cells of Swiss albino mice.

Anilofos is an organophosphate compound and is used extensively as a preemergence and early postemergence herbicide for the management of sedges, annual grasses, and some broad-leaved weeds in rice fields. The present study was aimed to assess the mutagenic potential of anilofos after sub-chronic exposure in Swiss albino mice. For this, a combined approach employing micronucleus (MN), chromosomal aberration (CA) studies and sperm-head abnormalities (SHAs) was used. Three dose levels of 1%, 2%, and 4% of maximum tolerated dose (MTD) (235 mg/kg b.wt.), that is, 2.35, 4.7 and 9.4 mg/kg b.wt., respectively, were administered orally daily for 90 days. A higher incidence of micronucleated erythrocytes (polychromatic erythrocytes + normochromatic erythrocytes), significant increase in CA frequency, and significant decrease in the ratio of polychromatic/normochromatic erythrocytes (P/N) ratio were observed at the 4.7 and 9.4 mg/kg b.wt. dose levels. A significant increase in SHA was observed in all treatment groups (2.35, 4.7, and 9.4 mg/kg b.wt.) from the control group. In conclusion, anilofos exposure of 2% and 4% of MTD caused a higher rate of micronucleated erythrocytes, increased frequency of CA, increase in SHA, and lower P/N ratio, and pesticide exposure of 1% of MTD only resulted in higher SHAs. Thus, anilofos was found to have mutagenic potential in mice when administered daily orally at dose rate of 4.7 and 9.4 mg/kg b.wt. for 90 days.

50 years existence and active participation of EEMS (now EEMGS) in the scientific community: A driver of European and international scientific collaborations for the protection of the environment and human health from genome stressors

EEMS and its successor Society EEMGS have provided a dynamic and successful platform to stimulate research and exchanges among the different actors involved in the protection of the environment and of human health from exposure to genome stressors. It includes basic, translational and applied research projects. This was possible due to the enthusiasm, creativity and support of scientists convinced of the importance of these issues.In the future young scientists will take over with new questions, new challenges, new technologies, new discoveries and new applications. A major challenge is the ethical questions emerging from the impressive potential of present genetic technologies capable of impacting the evolution of nature and humankind. The EEMGS, where academics, regulators and industries meet, should play a central role in these aspects, in particular in support of primary prevention and the establishment of internationally recognized guidelines. Collaboration with colleagues and other teams are of great importance to establish a stimulating open dialogue on scientific questions. However the key issues remain to do careful and rigorous research; to use logic and background knowledge; to define adequate experimental designs; to provide transparency in the protocols; to check repeatability of the results and to combine several statistical approaches in the quest to get to the truth.Among the many challenges ahead, re-evaluation of some key fundamental questions is necessary, such as the interplay between genetics and epigenetics, the existence of specific germ cell mutagens or the identification of the mechanisms leading to mutagen induced diseases. Translational and applied research will further include the development of systemic biomonitoring protocols, if possible in a single biological sample, the redaction of internationally harmonized guidelines but also the organization of platforms between geneticists and physicians open to all actors in the field. The creation of an independent European center to assess risk from exposure to mutagens, in particular in the light of the problematic of global warming might be very helpful.

Celebrating 50 Years of EMGS: A Visionary Idea Continues.

Celebrating 50 Years of EMGS: A Visionary Idea Continues.

A Return to the Origin of the EMGS: Rejuvenating the Quest for Human Germ Cell Mutagens and Determining the Risk to Future Generations.

Fifty years ago, the Environmental Mutagen Society (now Environmental Mutagenesis and Genomics Society) was founded with a laser-focus on germ cell mutagenesis and the protection of "our most vital assets"-the sperm and egg genomes. Yet, five decades on, despite the fact that many agents have been demonstrated to induce inherited changes in the offspring of exposed laboratory rodents, there is no consensus on whether human germ cell mutagens exist. We argue that it is time to reevaluate the available data and conclude that we already have evidence for the existence of environmental exposures that impact human germ cells. What is missing are definite data to demonstrate a significant increase in de novo mutations in the offspring of exposed parents. We believe that with over two decades of research advancing knowledge and technologies in genomics, we are at the cusp of generating data to conclusively show that environmental exposures cause heritable de novo changes in the human offspring. We call on the research community to harness our technologies, synergize our efforts, and return to our Founders' original focus. The next 50 years must involve collaborative work between clinicians, epidemiologists, genetic toxicologists, genomics experts and bioinformaticians to precisely define how environmental exposures impact germ cell genomes. It is time for the research and regulatory communities to prepare to interpret the coming outpouring of data and develop a framework for managing, communicating and mitigating the risk of exposure to human germ cell mutagens. Environ. Mol. Mutagen. 61:42-54, 2020. © 2019 Her Majesty the Queen in Right of Canada.

The mutagenesis moonshot: The propitious beginnings of the environmental mutagenesis and genomics society.

A mutagenesis moonshot addressing the influence of the environment on our genetic wellbeing was launched just 2 months before astronauts landed on the moon. Its impetus included the discovery that X-rays (Muller HJ. [1927]: Science 64:84-87) and chemicals (Auerbach and Robson. [1946]: Nature 157:302) were germ-cell mutagens, the introduction of a growing number of untested chemicals into the environment after World War II, and an increasing awareness of the role of environmental pollution on human health. Due to mounting concern from influential scientists that germ-cell mutagens might be ubiquitous in the environment, Alexander Hollaender and colleagues founded in 1969 the Environmental Mutagen Society (EMS), now the Environmental Mutagenesis and Genomics Society (EMGS); Frits Sobels founded the European EMS in 1970. As Fred de Serres noted, such societies were necessary because protecting populations from environmental mutagens could not be addressed by existing scientific societies, and new multidisciplinary alliances were required to spearhead this movement. The nascent EMS gathered policy makers and scientists from government, industry, and academia who became advocates for laws requiring genetic toxicity testing of pesticides and drugs and helped implement those laws. They created an electronic database of the mutagenesis literature; established a peer-reviewed journal; promoted basic and applied research in DNA repair and mutagenesis; and established training programs that expanded the science worldwide. Despite these successes, one objective remains unfulfilled: identification of human germ-cell mutagens. After 50 years, the voyage continues, and a vibrant EMGS is needed to bring the mission to its intended target of protecting populations from genetic hazards. Environ. Mol. Mutagen. 61:8-24, 2020. © 2019 Wiley Periodicals, Inc.

Paternal exposure to benzo(a)pyrene induces genome-wide mutations in mouse offspring

Understanding the effects of environmental exposures on germline mutation rates has been a decades-long pursuit in genetics. We used next-generation sequencing and comparative genomic hybridization arrays to investigate genome-wide mutations in the offspring of male mice exposed to benzo(a)pyrene (BaP), a common environmental pollutant. We demonstrate that offspring developing from sperm exposed during the mitotic or post-mitotic phases of spermatogenesis have significantly more de novo single nucleotide variants (1.8-fold; P < 0.01) than controls. Both phases of spermatogenesis are susceptible to the induction of heritable mutations, although mutations arising from post-fertilization events are more common after post-mitotic exposure. In addition, the mutation spectra in sperm and offspring of BaP-exposed males are consistent. Finally, we report a significant increase in transmitted copy number duplications (P = 0.001) in BaP-exposed sires. Our study demonstrates that germ cell mutagen exposures induce genome-wide mutations in the offspring that may be associated with adverse health outcomes.

Carbendazim induces death in alveolar epithelial cells: A comparison between submerged and at the air-liquid interface cell culture

The fungicide Carbendazim is widely used in agriculture and preservation of films and fibers. In mammals, it can promote germ cell mutagenicity, carcinogenicity, and reproductive toxicity. However, few data about the effects of this toxicant upon the respiratory system are available. In this work, we evaluated Carbendazim toxicity upon A549 alveolar cells both in monolayer and upon air-liquid interface cell system. Monolayer cell exposed to non-cytotoxic concentrations of this fungicide showed cell arrest at G2/M phase, and did not show additional alterations. On the other hand, alveolar 3D reconstructed epithelial model (air-liquid interface cell system) was characterized and exposed to IC25 of Carbendazim using the Vitrocell® Cloud 12 chamber. Expression of Active Caspase-3, α-tubulin and ROS was significantly increased after such exposure. Mitochondrial activity was also reduced after exposed to Carbendazim. The obtained results indicate that besides the environmental and reproductive toxicity concerns regarding Carbendazim exposure, pulmonary toxicity must be considered for this fungicide. In addition, we observed that the way of exposure impacts considerably on the cell response for in vitro assessment of chemicals inhalation toxicity profile.

Significant increase in the dominant lethal mutations: benzo(a)pyrene

Background: Use of biomarkers particularly under in vivo condition is of importance in risk assessment. This study is proposed to identify the in vivo antigenotoxic potential using dominant lethal mutation test in B(a)P exposed, turmeric fed animals and evaluate the role of turmeric in counteracting the germ cell mutations. Methods: Twenty four male mice were taken and divided into 4 groups each group containing 6 mice/group. The first and second group received stock diet (control) while the third and fourth groups received 5% turmeric diet for a period of one month. At the end of feeding period, second and fourth group received a single dose of benzo(a)pyrene 1mg/mouse intraperitoneally. After 1 week each male mouse was mated to 3 female mice at 1,4, 8 and 12 week intervals. On 13th day from mid point of mating, the females were sacrificed and live and dead embryos were counted to study the pre and post implantation loss of embryos. The differences in the frequencies of pre and post implant deaths were done by comparing treated with control group at 4-time periods by analysis of variance (ANOVA) and chi-square test which is used to compare treatment group and positive control with negative control. Results: Feeding of 5% turmeric and treatment with a single dose of B(a)P did not show any significant effect on the mutagenic index or the frequency of pregnancy in treated groups compared with control. There was no apparent induction of dominant lethal mutations in B(a)P or B(a)P+Turmeric treated groups. No post implantation dominant lethal effects produced by B(a)P could be detected in this study. The induction of B(a)P and treatment with turmeric did not influence the dominant lethal test. Conclusion: B(a)P did not produce a significant increase in the dominant lethal mutations. Turmeric also did not give any positive response and may be considered as non-mutagenic. The negative result suggests that under the conditions of the test the test substance may not be genotoxic in the germ cell of the treated sex of the test species.

Simulation of mouse and rat spermatogenesis to inform genotoxicity testing using OECD test guideline 488

The Organisation for Economic Co-operation and Development Test Guideline (TG) 488 for the transgenic rodent (TGR) mutation assay recommends two sampling times for assessing germ cell mutagenicity following the required 28-day exposure period: 28 + > 49 days for mouse sperm and 28 + >70 days for rat sperm from the cauda epididymis, or three days (i.e., 28 + 3d) for germ cells from seminiferous tubules (hereafter, tubule germ cells) plus caudal sperm for mouse and rat. Although the latter protocol is commonly used for mutagenicity testing in somatic tissues, it has several shortcomings for germ cell testing because it provides limited exposure of the proliferating phase of spermatogenesis when mutations are fixed in the transgene. Indeed, analysis of sperm at 28 + 3d has generated negative results with established germ cell mutagens, while the analysis of tubule germ cells has generated both positive and either negative or equivocal results. The Germ Cell workgroup of the Genetic Toxicology Technical Committee of the Health and Environmental Sciences Institute modelled mouse and rat spermatogenesis to better define the exposure history of the cell population collected from seminiferous tubules. The modelling showed that mouse tubule germ cells at 28 + 3d receive, as a whole, 42% of the total exposure during the proliferating phase. This percentage increases to 99% at 28 + 28d and reaches 100% at 28 + 30d. In the rat, these percentages are 22% and 80% at 28 + 3d and 28 + 28d, reaching 100% at 28 + 44d. These results show that analysis of tubule germ cells at 28 + 28d may be an effective protocol for assessing germ cell mutagenicity in mice and rats using TG 488. Since TG 488 recommends the 28 + 28d protocol for slow dividing somatic tissues, this appears to be a better compromise than 28 + 3d when slow dividing somatic tissues or germ cells are the critical tissues of interest.

Identifying germ cell mutagens using OECD test guideline 488 (transgenic rodent somatic and germ cell gene mutation assays) and integration with somatic cell testing

The Organisation for Economic Co-operation and Development Test Guideline 488 (TG 488) provides recommendations for assessing germ cell and somatic cell mutagenicity using transgenic rodent (TGR) models. However, important data gaps exist for selecting an optimal approach for simultaneously evaluating mutagenicity in both cell types. It is uncertain whether analysis of germ cells from seminiferous tubules (hereafter, tubule germ cells) or caudal sperm within the recommended design for somatic tissues (i.e., 28 days of exposure plus three days of fixation time, 28 + 3d) has enough sensitivity to detect an effect as compared with the analysis of sperm within the recommended design for germ cells (i.e., 28 + 49d and 28 + 70d for mouse and rat, respectively). To address these data gaps, the Germ Cell workgroup of the Genetic Toxicology Technical Committee of the Health and Environmental Sciences Institute reviewed the available TGR mutagenicity data in male germ cells, and, characterized the exposure history of tubule germ cells for different sampling times to evaluate its impact on germ cell mutagenicity testing using TG 488. Our analyses suggest that evaluating mutant frequencies in: i) sperm from the cauda epididymis at 28 + 3d does not provide meaningful mutagenicity data; ii), tubule germ cells at 28 + 3d provides reliable mutagenicity data only if the results are positive; and iii) tubule germ cells at 28 + 28d produces reliable positive and negative results in both mice and rats. Thus, the 28 + 28d regimen may provide an approach for simultaneously assessing mutagenicity in somatic tissues and germ cells from the same animals. Further work is required to support the 28 + 28d protocol for tissues other than slowly proliferating tissues as per current TG 488. Finally, recommendations are provided to guide the experimental design for germ cell mutagenicity data for regulatory submission, as well as other possible approaches to increase the reliability of the TGR assay.

Prioritization of reproductive toxicants in unconventional oil and gas operations using a multi-country regulatory data-driven hazard assessment

BackgroundRecent trends have witnessed the global growth of unconventional oil and gas (UOG) production. Epidemiologic studies have suggested associations between proximity to UOG operations with increased adverse birth outcomes and cancer, though specific potential etiologic agents have not yet been identified. To perform effective risk assessment of chemicals used in UOG production, the first step of hazard identification followed by prioritization specifically for reproductive toxicity, carcinogenicity and mutagenicity is crucial in an evidence-based risk assessment approach. To date, there is no single hazard classification list based on the United Nations Globally Harmonized System (GHS), with countries applying the GHS standards to generate their own chemical hazard classification lists. A current challenge for chemical prioritization, particularly for a multi-national industry, is inconsistent hazard classification which may result in misjudgment of the potential public health risks. We present a novel approach for hazard identification followed by prioritization of reproductive toxicants found in UOG operations using publicly available regulatory databases. MethodsGHS classification for reproductive toxicity of 157 UOG-related chemicals identified as potential reproductive or developmental toxicants in a previous publication was assessed using eleven governmental regulatory agency databases. If there was discordance in classifications across agencies, the most stringent classification was assigned. Chemicals in the category of known or presumed human reproductive toxicants were further evaluated for carcinogenicity and germ cell mutagenicity based on government classifications. A scoring system was utilized to assign numerical values for reproductive health, cancer and germ cell mutation hazard endpoints. Using a Cytoscape analysis, both qualitative and quantitative results were presented visually to readily identify high priority UOG chemicals with evidence of multiple adverse effects. ResultsWe observed substantial inconsistencies in classification among the 11 databases. By adopting the most stringent classification within and across countries, 43 chemicals were classified as known or presumed human reproductive toxicants (GHS Category 1), while 31 chemicals were classified as suspected human reproductive toxicants (GHS Category 2). The 43 reproductive toxicants were further subjected to analysis for carcinogenic and mutagenic properties. Calculated hazard scores and Cytoscape visualization yielded several high priority chemicals including potassium dichromate, cadmium, benzene and ethylene oxide. ConclusionsOur findings reveal diverging GHS classification outcomes for UOG chemicals across regulatory agencies. Adoption of the most stringent classification with application of hazard scores provides a useful approach to prioritize reproductive toxicants in UOG and other industries for exposure assessments and selection of safer alternatives.

Discovering human germ cell mutagens with whole genome sequencing: Insights from power calculations reveal the importance of controlling for between-family variability

Mutations in germ cells pose potential genetic risks to offspring. However, de novo mutations are rare events that are spread across the genome and are difficult to detect. Thus, studies in this area have generally been under-powered, and no human germ cell mutagen has been identified. Whole Genome Sequencing (WGS) of human pedigrees has been proposed as an approach to overcome these technical and statistical challenges. WGS enables analysis of a much wider breadth of the genome than traditional approaches. Here, we performed power analyses to determine the feasibility of using WGS in human families to identify germ cell mutagens. Different statistical models were compared in the power analyses (ANOVA and multiple regression for one-child families, and mixed effect model sampling between two to four siblings per family). Assumptions were made based on parameters from the existing literature, such as the mutation-by-paternal age effect. We explored two scenarios: a constant effect due to an exposure that occurred in the past, and an accumulating effect where the exposure is continuing. Our analysis revealed the importance of modeling inter-family variability of the mutation-by-paternal age effect. Statistical power was improved by models accounting for the family-to-family variability. Our power analyses suggest that sufficient statistical power can be attained with 4–28 four-sibling families per treatment group, when the increase in mutations ranges from 40 to 10% respectively. Modeling family variability using mixed effect models provided a reduction in sample size compared to a multiple regression approach. Much larger sample sizes were required to detect an interaction effect between environmental exposures and paternal age. These findings inform study design and statistical modeling approaches to improve power and reduce sequencing costs for future studies in this area.

27Al NMR Study of the pH Dependent Hydrolysis Products of Al₂(SO₄)₃ in Different Physiological Media.

Soluble inorganic aluminium compounds like aluminium sulfate or aluminium chloride have been challenged by the European Chemical Agency to induce germ cell mutagenicity. Before conducting mutagenicity tests, the hydrolysis products in water and in physiological solutions should be determined as a function of the concentration and pH. We used different 27Al NMR spectroscopic techniques (heteronuclear Overhauser effect spectroscopy (HOESY), exchange spectroscopy (EXSY), diffusion ordered (DOSY)) in this work to gain the information to study the aluminium species in solutions with Al2(SO4)3 concentrations of 50.0, 5.0, and 0.5 g/L and their pH and time dependent transformation. At low pH, three different species were present in all physiological solutions and water: [Al(OH)n(H2O)6 − n](3 − n)+ (n = 0–2), [Al(H2O)5SO4]+, and [Al2(OH)2(H2O)8]4+. Increasing pH reduced the amounts of the two monomer species, with a complete loss at pH 5 for solutions with a concentration of 50.0 g/L and at pH 4 for solutions with a concentration of 5.0 g/L. The dimer species [Al2(OH)2(H2O)8]4+ is present in a pH range between 3 and 6. Less symmetric oligomeric and probably asymmetric aluminium species are formed at pH of 5 and 6. The pH value is the driving force for the formation of aluminium species in all media, whereas the specific medium had only minor effect. No conclusive information could be obtained at pH 7 due to signal loss related to fast quadrupole relaxation of asymmetric aluminium species. A slight reduction of the content of the symmetric aluminium species due to the formation of oligomeric species was observed over a period of 6 weeks. Reference 27Al NMR experiments conducted on saturated water solutions of AlCl3 and those with a concentration of 50 g/L show that the type of salt/counter ion at the same concentration and pH influences the hydrolysis products formed.