Use of serum-free media for peripheral blood mononuclear cell culture and the impact on T and B cell readouts.

Introduction: As part of a wider programme of work developing next-generation risk assessment approaches (NGRA) using non-animal methods (NAMs) for safety assessment of materials, Unilever SEAC is exploring the use of a peripheral blood mononuclear cell (PBMC) system to investigate how cells from different arms of the human immune system are impacted by different treatments. To maximise human relevance, the cell cultures are supported by human serum, but this came with some challenges, including an inability to measure induced levels of immunoglobulins due to high background levels. Therefore, a study comparing use of human sera containing media with three different chemically defined serum-free media was undertaken.

Impact of a varied set of stimuli on a suite of immunological parameters within peripheral blood mononuclear cells: toward a non-animal approach for assessing immune modulation by materials intended for human use.

In toxicology, steps are being taken towards more mechanism-focused and human relevant approaches to risk assessment, requiring new approaches and methods. Additionally, there is increasing emphasis by regulators on risk assessment of immunotoxicity. Here we present data from a peripheral blood mononuclear cell (PBMC) system whereby a varied set of stimuli, including those against the TCR and Toll-like receptors, enable readouts of cytokine and prostaglandin E2 (PGE2) production with monocyte, T cell and B cell viability, proliferation, and associated activation markers.

Searching for LINCS to Stress: Using Text Mining to Automate Reference Chemical Curation.

Adaptive stress response pathways (SRPs) restore cellular homeostasis following perturbation but may activate terminal outcomes like apoptosis, autophagy, or cellular senescence if disruption exceeds critical thresholds. Because SRPs hold the key to vital cellular tipping points, they are targeted for therapeutic interventions and assessed as biomarkers of toxicity. Hence, we are developing a public database of chemicals that perturb SRPs to enable new data-driven tools to improve public health.

60 Years of the 3Rs symposium: Lessons learned and the road ahead.

When The Principles of Humane Experimental Technique was published in 1959, authors William Russell and Rex Burch had a modest goal: to make researchers think about what they were doing in the laboratory – and to do it more humanely. Sixty years later, their groundbreaking book was celebrated for inspiring a revolution in science and launching a new field: The 3Rs of alternatives to animal experimentation. On November 22, 2019, some pioneering and leading scientists and researchers in the field gathered at the Johns Hopkins Bloomberg School of Public Health in Bal­timore for the 60 Years of the 3Rs Symposium: Lessons Learned and the Road Ahead.

Brazilian National Network of Alternative Methods (RENAMA) 10th Anniversary: Meeting of the Associated Laboratories, May 2022.

The Brazilian National Network of Alternative Methods (RENAMA), which is linked to the Ministry of Science, Technology and Innovation, is currently comprised of 51 laboratories from CROs, academia, industry and government. RENAMA's aim is to develop and validate new approach methodologies (NAMs), as well as train researchers and disseminate information on their use - thus reducing Brazilian, and consequently Latin American, dependence on external technology. Moreover, it promotes the adoption of NAMs by educators and trained researchers, as well as the implementation of good laboratory practice (GLP) and the use of certified products.

Exploring the effects of experimental parameters and data modeling approaches on in vitro transcriptomic point-of-departure estimates.

Multiple new approach methods (NAMs) are being developed to rapidly screen large numbers of chemicals to aid in hazard evaluation and risk assessments. High-throughput transcriptomics (HTTr) in human cell lines has been proposed as a first-tier screening approach for determining the types of bioactivity a chemical can cause (activation of specific targets vs. generalized cell stress) and for calculating transcriptional points of departure (tPODs) based on changes in gene expression.

Invariant NKT cells are more abundant in peanut-allergic adults and a subset of CD8 iNKT cells are depleted after peanut oil exposure.

Introduction: Peanut allergy is one of the most prevalent food allergies globally. Currently, most research into the mechanisms involved in protein allergy focuses on the protein allergens under investigation, and information on the function of accompanying compounds, such as lipids, is scarce. Thus, this research investigates the role of peanut-associated lipids and invariant natural killer T (iNKT) cells in peanut allergy using a novel, human, assay.

Repeat-exposure in vitro protocol to assess the risk of antimicrobial resistance (AMR) development from use of personal care products: Case study using an antibacterial liquid handwash.

The global crisis we are facing with regard to antibiotic resistance has been largely attributed to the overuse and misuse of antibiotics in healthcare and agriculture. However, there is also growing global concern about cross-resistance between biocides and antibiotics. This has made clear the need for more research in this area along with easy-to-perform, but realistic, methods to characterise the potential risk associated with cross-resistance to antibiotics due to biocide use.

Non-animal approaches for photoallergenicity safety assessment: Needs and perspectives for the toxicology for the 21st century.

Certain chemicals and/or their byproducts are photoactivated by UV/VIS and trigger a dermal allergenic response, clinically recognized as photoallergic contact dermatitis (PACD). It is important to identify the chemicals which are potentially photoallergenic, not only for establishing the correct differential diagnosis between PACD and other photodermatoses, but also as causative agents which should be avoided as a preventative measure. Moreover, materials with photoallergenic properties need to be correctly identified to allow thorough safety assessments for their use in finished products (e.

Land use diversification may mitigate on-site land use impacts on mammal populations and assemblages.

Land use is a major cause of biodiversity decline worldwide. Agricultural and forestry diversification measures, such as the inclusion of natural elements or diversified crop types, may reduce impacts on biodiversity. However, the extent to which such measures may compensate for the negative impacts of land use remains unknown.

Longitudinal characterization of TK6 cells sequentially adapted to animal product-free, chemically defined culture medium: considerations for genotoxicity studies.

approaches are an essential tool in screening for toxicity of new chemicals, products and therapeutics. To increase the reproducibility and human relevance of these assessments, it is advocated to remove animal-derived products such as foetal bovine serum (FBS) from the cell culture system. Currently, FBS is routinely used as a supplement in cell culture medium, but batch-to-batch variability may introduce inconsistency in inter- and intra-lab assessments.

Data-driven learning of narcosis mode of action identifies a CNS transcriptional signature shared between whole organism Caenorhabditis elegans and a fish gill cell line.

With the large numbers of man-made chemicals produced and released in the environment, there is a need to provide assessments on their potential effects on environmental safety and human health. Current regulatory frameworks rely on a mix of both hazard and risk-based approaches to make safety decisions, but the large number of chemicals in commerce combined with an increased need to conduct assessments in the absence of animal testing makes this increasingly challenging. This challenge is catalysing the use of more mechanistic knowledge in safety assessment from both in silico and in vitro approaches in the hope that this will increase confidence in being able to identify modes of action (MoA) for the chemicals in question.

Latent Variables Capture Pathway-Level Points of Departure in High-Throughput Toxicogenomic Data.

Estimation of points of departure (PoDs) from high-throughput transcriptomic data (HTTr) represents a key step in the development of next-generation risk assessment (NGRA). Current approaches mainly rely on single key gene targets, which are constrained by the information currently available in the knowledge base and make interpretation challenging as scientists need to interpret PoDs for thousands of genes or hundreds of pathways. In this work, we aimed to address these issues by developing a computational workflow to investigate the pathway concentration-response relationships in a way that is not fully constrained by known biology and also facilitates interpretation.

Fate of household and personal care chemicals in typical urban wastewater treatment plants indicate different seasonal patterns and removal mechanisms.

Studies on the presence and fate of household and personal care chemicals (HPCCs) in wastewater treatment plants (WWTPs) are important due to their increasing consumption worldwide. The seasonal patterns and removal mechanisms of HPCCs are not well understood for WWTPs that apply different treatment technologies. To answer these questions, the sewage and sludge samples were taken from 10 typical WWTPs in Northeast China.

Identifying key priorities for research to protect the consumer with food hypersensitivity: A UK Food Standards Agency Priority Setting Exercise.

Introduction: Food hypersensitivity (FHS), including food allergy, coeliac disease and food intolerance, is a major public health issue. The Food Standards Agency (FSA), an independent UK Government department working to protect public health and consumers' wider interests in food, sought to identify research priorities in the area of FHS.

Methods: A priority setting exercise was undertaken, using a methodology adapted from the James Lind Alliance-the first such exercise with respect to food hypersensitivity.

Validation of sampling techniques and SPE-UPLC/MS/MS for home and personal care chemicals in the Songhua Catchment, Northeast China.

A method for the simultaneous determination of 18 home and personal care chemicals (HPCCs) in river water and wastewater was developed using solid-phase extraction and ultra-high-performance liquid chromatography with tandem mass spectrometry (SPE and UPLC/MS/MS). A series of tests were designed to find the potential background interference and loss of HPCCs during the sample preservation and pretreatment process. Our results suggested that a considerable amount of some target compounds were lost with increasing days of storage even at 4 °C.

Towards a surrogate system to express human lipid binding TCRs.

Background: Previously we reported that natural nut lipids were necessary for sensitization and that natural killer T cells (NKTs) must play a critical role in the development of food allergic responses. A major bottleneck in further understanding the interaction of nut lipids with the cells of the human immune system is the lack of well-characterized lipid responsive human cell lines.

Objective: In the present study, we engineered human T cell receptor (TCR) sequences TRAV10 and TRBV25 responsive to α-GalCer into a stable murine iNKT hybridoma and surrogate human T cell lines.

Characterizing the coverage of critical effects relevant in the safety evaluation of food additives by AOPs.

There is considerable interest in adverse outcome pathways (AOPs) as a means of organizing biological and toxicological information to assist in data interpretation and method development. While several chemical sectors have shown considerable progress in applying this approach, this has not been the case in the food sector. In the present study, safety evaluation reports of food additives listed in Annex II of Regulation (EC) No 1333/2008 of the European Union were screened to qualitatively and quantitatively characterize toxicity induced in laboratory animals.

IL7 receptor signaling in T cells: A mathematical modeling perspective.

Interleukin-7 (IL7) plays a nonredundant role in T cell survival and homeostasis, which is illustrated in the severe T cell lymphopenia of IL7-deficient mice, or demonstrated in animals or humans that lack expression of either the IL7Rα or γ chain, the two subunits that constitute the functional IL7 receptor. Remarkably, IL7 is not expressed by T cells themselves, but produced in limited amounts by radio-resistant stromal cells. Thus, T cells need to constantly compete for IL7 to survive.

How to bring absolute sustainability into decision-making: An industry case study using a Planetary Boundary-based methodology.

The Planetary Boundaries concept has emerged as a framework for articulating environmental limits, gaining traction as a basis for considering sustainability in business settings, government policy and international guidelines. There is emerging interest in using the Planetary Boundaries concept as part of life cycle assessment (LCA) for gauging absolute environmental sustainability. We tested the applicability of a novel Planetary Boundaries-based life cycle impact assessment methodology on a hypothetical laundry washing case study at the EU level.

Application of a spatially resolved model to contextualise monitoring data for risk assessment of down-the-drain chemicals over large scales.

Many regulatory screening level exposure assessments are based on simple large scale conceptual scenarios. However, exposure, and therefore risks associated with chemicals, are characterised by high spatial variability. The Scenario assembly tool (ScenAT) is a global screening level model to enable spatially resolved local predictions of environmental concentrations of home and personal care chemicals.

Assessing food allergy risks from residual peanut protein in highly refined vegetable oil.

Refined vegetable oils including refined peanut oil are widely used in foods. Due to shared production processes, refined non-peanut vegetable oils can contain residual peanut proteins. We estimated the predicted number of allergic reactions to residual peanut proteins using probabilistic risk assessment applied to several scenarios involving food products made with vegetable oils.

Genotoxicity testing of a Hoodia gordonii extract.

Hoodia gordonii extract consists of a mixture of steroid glycosides, fatty acids, plant sterols and alcohols. As part of the overall safety assessment H. gordonii extract was assessed for genotoxicity in two assays in vitro: a bacterial mutation assay; and a gene mutation assay using mouse lymphoma cells.

Modelling concentrations of decamethylcyclopentasiloxane in two UK rivers using LF2000-WQX.

Current regulatory environmental exposure assessments for decamethylcyclopentasiloxane (D(5)), used in a range of personal care products, are based on a number of erroneous assumptions. Using an estimated D(5) flux to waste water of 11.6 mg cap(-1) d(-1), a 95.