Towards a reliable assessment of nanomaterial health effects using advanced biological models and assays

A sound scientific basis is needed to assess the risks to workers and consumers, to inform regulatory bodies and to ensure a responsible development of nanotechnology. Most of the existing laboratory (in vitro) biological models, exposure systems and doses, as well data (in silico) models do not reflect the real-life exposure to nanomaterials (NMs). A significant source for unreliable results is represented by possible interactions of NMs with the reagents and detection systems for toxicity evaluation. The fast pace at which NMs enter the market requires a shift from expensive and ethically doubtful animal testing to innovative, reliable and socially acceptable in vitro and in silico test systems. The aim of NanoBioReal is to establish methods and biological models that reflect real-life exposure and which provide a reliable, robust and efficient platform to evaluate the effects of NMs on human health. Our testing system covers a wide area of biological models, from single cells to three-dimensional (3D) models that simulate tissues and organs. These include air-liquid interface (ALI) models for lung exposure, blood and «organ-on-a-chip» systems (lung and microvasculature-on-a-chip) that measure effects in real-time and can detect relevant effects after both short and longtime exposure. To avoid interferences caused by NMs, we established in this project label-free impedance-based methods to evaluate cytotoxicity and cyclic voltammetry to assess oxidative stress. A set of representative NMs has been produced and physico-chemically characterized. Their in vitro cyto- and geno- toxicity was assessed in 2D cellular models and the results were compared to those obtained using the advanced 3D models and an animal model. Advanced models for lung, vasculature and whole blood exposure have been established and used for toxicity, inflammation, oxidative stress, and barrier integrity testing. A microfluidic setup for label-free live monitoring of cells and 3D biological models was established, optimized and its throughput has been increased. The NanoBioReal project has delivered reliable, robust and relevant biological and in silico-models to support a “safe(r)-by-design” approach to the development of NMs and to address the needs of various stakeholders and regulators. National partners: Dept. of Clinical Dentistry (IKO), Fac. of Medicine, Univ. of Bergen (UiB), Norwegian Inst. for Air Research (NILU), National Inst. of Occupational Health (STAMI), and Norwegian Univ. of Science and Technology (NTNU). Subcontractors: NorGenotech. International partners: Catalan Inst. of Nanoscience and Nanotechnology (ICN2), Univ. of Gdansk. Collaborators: Dept. of Physics and Technology (UiB), Dept. of Electrical Engineering (HVL), and NIOM.

The NanoBioReal project contributes to a safer, more responsible and sustainable development of nanotechnology-based industry and use of its products by the society. This is based on addressing the urgent need to generate reliable science-based knowledge regarding the effects of nanomaterials (NMs) on human health and their safety in the working environment, as well as for consumers, and patients. The NanoBioReal project also addresses the RRI aspects outlined in the Nano2021 program. The project has lead to significant improvements of existing methods and biological models by getting closer to real-life exposure and thus enhancing their relevance, as well as by improving their efficiency through increased high throughput testing in order to keep up with the rapid pace of NM development. Furthermore, by maintaining current national nanosafety networks in Norway and establishing new networks with the European NanoSafety Cluster and the international network initiative on safe and sustainable nanotechnology (INISS), the consortium has had access to the best knowledge and expertise in Norway and in Europe. It consolidates the basis for the establishment of new networks, i.e., of a national “NanoSafety platform”. Close cross-cutting interaction of the partners within and between WPs, and their complementary skills has allowed the integration of their research and technology into a common innovative NanoBioReal approach. We have recruited and trained skilled professionals, such as the post-doc and young researchers to train competent staff with broad networks to the industry in the future. NanoBioReal also contributes with nanosafety aspects (UiB) in the Research School for Training the Next Generation of Micro- and Nanotechnology Researchers in Norway (TNNN). Moreover, NanoGenoTox (NGT) (SME participant in the project) will benefit from positioning the company as a reliable nanosafety assessment player. NGT has established new services suitable for NM safety assessment. The NanoBioReal results will contribute to helping other SME start-up companies to resolve HSE issues necessary for their sustainable growth. Overall, the industry will benefit as a result of the time- and cost-efficient, more relevant for the real situation and reliable hazard assessment platform developed in this project. This project contributes also to the improvement of environmental consequences, since similar biological effects could be relevant for organisms in the environment.

This proposal answers to the objective of the 5th thematic priority to expand the insight into the impacts of nanomaterials (NMs) on human health by addressing two of the main sources of uncertainty that hamper the assessment of real-life impact of NMs on human health: i) most of the existing in vitro and in silico models do not reflect real life exposure to NMs and ii) potential interferences of NMs with assays or detection systems. The aim of NanoBioReal is to provide solutions to these challenges by delivering i) beyond state of the art advanced biological models, from single-cells to organ-on-a-chip, representing target tissues and organs and ii) innovative realistic, reliable assessment tools integrated into an in vitro and in silico testing toolbox. The NMs selected for testing are relevant for Norwegian conditions and industry and include TiO2, ZnO, SiO2, nano-silver, nano-gold, and widely used polymer-based dental materials containing embedded nanoparticles. The key biological processes and mechanisms that will be investigated are: i) cellular uptake; ii) inflammation, iii) oxidative stress, iv) genotoxicity, DNA damage and DNA repair and v) cell death. At the end of the project a toolbox will be generated integrating reliable, robust, and efficient in-vivo relevant biological models and methods to support a safe-by-design approach to nanomaterial development and to answer the needs for nanomaterial hazard assessment of various end-users, stakeholders and regulators. The outcome of the project will contribute not only to bringing new mechanistic insights into the health impact of NMs, but also to reducing the uncertainty in health hazard and consequently in risk assessment, as well as to replacing animal use, in conformity with the 3R principles (reduce, refine, replace animal use).