High throughput toxicity screening and intracellular detection of nanomaterials

EC FP7 NANoREG (Grant Agreement NMP4-LA-2013-310584)Free PMC Article: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5215403/With the growing numbers of nanomaterials (NMs), there is a great demand for rapid and reliable ways of testing NM safety—preferably using in vitro approaches, to avoid the ethical dilemmas associated with animal research. Data are needed for developing intelligent testing strategies for risk assessment of NMs, based on grouping and read-across approaches. The adoption of high throughput screening (HTS) and high content analysis (HCA) for NM toxicity testing allows the testing of numerous materials at different concentrations and on different types of cells, reduces the effect of inter-experimental variation, and makes substantial savings in time and cost.info:eu-repo/semantics/publishedVersio

Determination of peracetic acid exposure concentration without effect on Atlantic salmon parr health, welfare or growth cultured in RAS.

Recirculating aquaculture systems (RAS) are replacing land-based flow through systems in Norway and other producing countries. RAS has many advances but also bring some biological challenges as water disinfection that may compromise biofilters performance and fish health and welfare. For example, Denmark is using peracetic acid (PAA) as a water disinfection strategy to control pathogens for rainbow trout production. PAA is a highly reactive peroxygen compound; 100 times more effective than hydrogen peroxide and has the potential to control bacterial, and parasitic infections and has a wide range of antimicrobial effects. PAA can be applied continuously or as a pulse to the water and degrades into oxygen and water. In Norway, PAA has been used to disinfect surfaces and different types of equipment in RAS facilities for salmonids, but not for water treatment. The effects of PAA on Atlantic salmon (Salmo salar) smolt and rainbow trout (Oncorhynchus mykiss) have already been studied, but no studies on Atlantic salmon parr have been performed to date. In this study, two trials (9 identical RAS in each trial) have been performed to identify the PAA concentration threshold that bears no effect on welfare and growth of Atlantic salmon parr. In trial 1, which included looking at fish external welfare, histology of gills and skin, swimming behaviour, and hand feeding was used to identify threshold. In the trial, the parr were exposed to PAA concentrations of 0, 0.05, 0.1,0.2 0.4, 0.8, 1.6, 3.2, and 6.4 mg/L for 1h and re-exposed two days later with the same treatment for 1h. The two highest treatments resulted in significant histological effects at skin and gills’ level. The feeding and swimming behaviour was significantly reduced with treatments of 1.6 - 6.4 mg/L PAA, respectively. Overall welfare indexes indicated that a treatment over 0.8 mg/L had a negative impact on fish wellbeing. In trial 2, DNA standard brakes in gills was analysed, growth and external fish welfare was used to identify threshold to Atlantic salmon. Three PAA treatments in triplicates, control (no PAA), low (0.1 mg/L) and high (1.0 mg/L) were performed. The PAA was added continuously for 4 weeks after the first sampling. There was no significant difference in fish growth, external welfare score, overall welfare index, or cellular DNA damage in the gills as measured by comet assay. The latter showed that 95% of the gill cells were intact and not affected by PAA. On the conditions tested in the present work, PAA exposure equal or below 1 mg/L seems to be safe for Atlantic salmon parr welfare and it can be considered a potential disinfectant in salmon production

ANALISIS KERUSAKAN DNA PADA SEL LIMFOSIT PASIEN PASCA-RADIOTERAPI

Analyses of DNA Damage in the Patient’s Lymphocyte Cells Post-Radiotherapy Radiotherapy given in high doses to kill cancer cells can also induce DNA damage in surrounding normal cells. The radiation dose is divided into smaller doses called fractionation to decrease the effect of radiation on normal tissue. For this reason, it is necessary to monitor the peripheral blood lymphocytes to evaluate the patient's DNA damage. The alkaline comet test is a simple and sensitive technique for detecting DNA instability. This study involved 11 patients who underwent radiotherapy up to 20 Gy, and 11 healthy subjects as controls. This study aims to see how much DNA damage is caused by a 20 Gy fractionated radiation dose in patients with various cancers. The results showed that the mean frequency of damaged cells in patients was 80.54 ± 12.52% with a mean comet tail length of 49.98 ± 12.93 µm. There was a significant difference in both the frequency of damaged cells and the mean value of the comet tail length against the control group (p < 0.001). It was concluded that high doses of radiation can cause DNA damage to peripheral blood lymphocytes. Radioterapi yang diberikan dalam dosis tinggi untuk mematikan sel kanker juga dapat menginduksi kerusakan DNA pada sel normal di sekitarnya. Dosis radiasi dibagi menjadi dosis yang lebih kecil yang disebut fraksinasi untuk menurunkan efek radiasi pada jaringan normal. Untuk itu perlu pemantauan pada limfosit darah tepi untuk mengevaluasi kerusakan DNA pasien. Uji komet alkali merupakan teknik yang sederhana dan sensitif untuk mendeteksi ketidakstabilan DNA. Penelitian ini melibatkan 11 pasien yang menjalani radioterapi hingga 20 Gy, dan 11 subyek sehat sebagai kontrol. Penelitian ini bertujuan untuk melihat seberapa besar kerusakan DNA akibat dosis radiasi fraksinasi 20 Gy pada pasien dengan variasi kanker. Hasil penelitian menunjukkan bahwa rerata frekuensi sel yang rusak pada pasien 80,54 ± 12,52% dengan rerata panjang ekor komet 49,98 ± 12,93 µm terdapat perbedaan nyata baik pada frekuensi sel yang rusak maupun nilai rerata panjang ekor komet terhadap kelompok kontrol (p < 0,001). Penelitian ini menyimpulkan bahwa radiasi dosis tinggi dapat menyebabkan kerusakan DNA sel limfosit darah tepi

Assessment of DNA structure and integrity in the human spermatozoon

Male fertility is routinely assessed by basic semen analysis, but this is poorly predictive of fertility outcome. One promising advanced diagnostic is assessing sperm DNA damage, but there is a lack of method standardisation, clinical thresholds and comparison data for different assays. The objective of this study was to validate standardised methods for the assessment of sperm DNA quality using TUNEL, Acridine orange (AO) and Chromomycin A3 (CMA3) in slide-based assays; and assess prognostic value for fertility and miscarriage. The data obtained reveals that using differing optimal mounting solution for different assays is key to reliable results. The use of DNA fragmentation inductors such as DNase and hydrogen peroxide in donor samples confirmed and validated the use of AO and TUNEL for detecting DNA damaged cells. A novel semi-automated computer-based scoring system for fluorescence microscopy images was devised and compared with visual operator results for intra-assay variability of AO and TUNEL assays. This system allowed objective and consistent results free of operator subjectivity. The assessment of TUNEL, AO and CMA3 values in a subset of patients from the HABSelect trial showed no correlation between the assays corroborating that different assays measure different aspects of DNA quality. The number of patient samples assessed were insufficiently powered to draw firm conclusions related to clinical outcome, but we believe they are useful in making a case for further investigations in the field

Analytical procedures and interpretation basis for biomarkers used in environmental effect monitoring for the water column at the Norwegian continental shelf

Prosjektleder Jonny BeyerHerein, method description documents for the biomarker parameters that are required or recommended by the current edition of the guidelines for offshore water column monitoring at the Norwegian shelf (M-300, revised version 2020) are systemised, examined and discussed. The objective of this study is to assess the quality status of open and available method and analysis descriptions, identify any significant shortages on biomarker method descriptions and method validation issues and to discuss/suggest possible correcting and improving measures.Offshore Norge (formerly NOROG, Norwegian Oil and gas)publishedVersio

Genotoxicity testing of nanomaterials

Nanomaterials have outstanding and unprecedented advantageous material properties but may also cause adverse effects in humans upon exposure. Testing nanomaterials for genotoxic properties is challenging because traditional testing methods were designed for small, soluble molecules and may not be easily applicable without modifications. This review critically examines available genotoxicity tests for use with nanomaterials, including DNA damage tests such as the comet assay, gene mutation tests such as the mouse lymphoma and hprt assay, and chromosome mutation tests such as the micronucleus test and the chromosome aberration test. It presents arguments for the relative usefulness of various tests, such as preferring the micronucleus test over the chromosome aberration test for scoring chromosome mutations and preferring mammalian cell gene mutation tests because the Ames test has limited utility. Finally, it points out the open questions and further needs in adapting genotoxicity tests for nanomaterials, such as validation, reference nanomaterials, and the selection of top test concentrations, as well as the relevance and applicability of test systems and the need to define testing strategies. This article is categorized under: Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicin

Semi-automated Ontology Generation for Biocuration and Semantic Search

Background: In the life sciences, the amount of literature and experimental data grows at a tremendous rate. In order to effectively access and integrate these data, biomedical ontologies – controlled, hierarchical vocabularies – are being developed. Creating and maintaining such ontologies is a difficult, labour-intensive, manual process. Many computational methods which can support ontology construction have been proposed in the past. However, good, validated systems are largely missing. Motivation: The biocuration community plays a central role in the development of ontologies. Any method that can support their efforts has the potential to have a huge impact in the life sciences. Recently, a number of semantic search engines were created that make use of biomedical ontologies for document retrieval. To transfer the technology to other knowledge domains, suitable ontologies need to be created. One area where ontologies may prove particularly useful is the search for alternative methods to animal testing, an area where comprehensive search is of special interest to determine the availability or unavailability of alternative methods. Results: The Dresden Ontology Generator for Directed Acyclic Graphs (DOG4DAG) developed in this thesis is a system which supports the creation and extension of ontologies by semi-automatically generating terms, definitions, and parent-child relations from text in PubMed, the web, and PDF repositories. The system is seamlessly integrated into OBO-Edit and Protégé, two widely used ontology editors in the life sciences. DOG4DAG generates terms by identifying statistically significant noun-phrases in text. For definitions and parent-child relations it employs pattern-based web searches. Each generation step has been systematically evaluated using manually validated benchmarks. The term generation leads to high quality terms also found in manually created ontologies. Definitions can be retrieved for up to 78% of terms, child ancestor relations for up to 54%. No other validated system exists that achieves comparable results. To improve the search for information on alternative methods to animal testing an ontology has been developed that contains 17,151 terms of which 10% were newly created and 90% were re-used from existing resources. This ontology is the core of Go3R, the first semantic search engine in this field. When a user performs a search query with Go3R, the search engine expands this request using the structure and terminology of the ontology. The machine classification employed in Go3R is capable of distinguishing documents related to alternative methods from those which are not with an F-measure of 90% on a manual benchmark. Approximately 200,000 of the 19 million documents listed in PubMed were identified as relevant, either because a specific term was contained or due to the automatic classification. The Go3R search engine is available on-line under www.Go3R.org

Semi-automated Ontology Generation for Biocuration and Semantic Search

Background: In the life sciences, the amount of literature and experimental data grows at a tremendous rate. In order to effectively access and integrate these data, biomedical ontologies – controlled, hierarchical vocabularies – are being developed. Creating and maintaining such ontologies is a difficult, labour-intensive, manual process. Many computational methods which can support ontology construction have been proposed in the past. However, good, validated systems are largely missing. Motivation: The biocuration community plays a central role in the development of ontologies. Any method that can support their efforts has the potential to have a huge impact in the life sciences. Recently, a number of semantic search engines were created that make use of biomedical ontologies for document retrieval. To transfer the technology to other knowledge domains, suitable ontologies need to be created. One area where ontologies may prove particularly useful is the search for alternative methods to animal testing, an area where comprehensive search is of special interest to determine the availability or unavailability of alternative methods. Results: The Dresden Ontology Generator for Directed Acyclic Graphs (DOG4DAG) developed in this thesis is a system which supports the creation and extension of ontologies by semi-automatically generating terms, definitions, and parent-child relations from text in PubMed, the web, and PDF repositories. The system is seamlessly integrated into OBO-Edit and Protégé, two widely used ontology editors in the life sciences. DOG4DAG generates terms by identifying statistically significant noun-phrases in text. For definitions and parent-child relations it employs pattern-based web searches. Each generation step has been systematically evaluated using manually validated benchmarks. The term generation leads to high quality terms also found in manually created ontologies. Definitions can be retrieved for up to 78% of terms, child ancestor relations for up to 54%. No other validated system exists that achieves comparable results. To improve the search for information on alternative methods to animal testing an ontology has been developed that contains 17,151 terms of which 10% were newly created and 90% were re-used from existing resources. This ontology is the core of Go3R, the first semantic search engine in this field. When a user performs a search query with Go3R, the search engine expands this request using the structure and terminology of the ontology. The machine classification employed in Go3R is capable of distinguishing documents related to alternative methods from those which are not with an F-measure of 90% on a manual benchmark. Approximately 200,000 of the 19 million documents listed in PubMed were identified as relevant, either because a specific term was contained or due to the automatic classification. The Go3R search engine is available on-line under www.Go3R.org