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Innovation
The 3Rs Centre Utrecht: connecting the 3Rs and the NAMs
This animation of the 3Rs Centre Utrecht shows the differences, but also the similarities, between the 3Rs (Replacement, Reduction, Refinement of animal testing) approach and the NAMs (New Approach Methodologies) approach when trying to replace or reduce experimental animal use.
Innovation examples
HealthInnovation
Whole blood assessment of thrombosis tendency
Transgenic animals are often subjected to short and long term experimental models of thrombosis and atherosclerosis with considerable discomfort to the animal. This project aims to: 1) replace (human blood instead of animal blood), 2) reduce (a few drops of blood per test), and 3) refine (replace in vivo by in vitro testing with isolated blood) the use of laboratory animals with two new small blood volume function tests—the perfusion chamber and the thrombin generation test. Both tests will be equipped with a simple detection capability, which is affordable for laboratories. Their application is not only in the field of thrombosis and haemostasis but also for the investigation of other blood-related diseases, such as arteriosclerosis, diabetes and cancer. By Sanne Brouns (Department of Biochemistry CARIM, Maastricht University, the Netherlands) and Linda Herfs (Flowchamber B.V.).
Conferences abstracts
Biotransformation of two proteratogenic anti-epileptics in the zebrafish (Danio rerio) embryo
The zebrafish (Danio rerio) embryo has gained interest as an alternative model for developmental toxicity testing, which still mainly relies on in vivo mammalian models (e.g., rat, rabbit). However, cytochrome P450 (CYP)-mediated drug metabolism, which is critical for the bioactivation of several proteratogens, is still under debate for this model. Therefore, we investigated the potential capacity of zebrafish embryos/larvae to bioactivate two known mammalian proteratogens, carbamazepine (CBZ) and phenytoin (PHE) into their mammalian active metabolites, carbamazepine-10,11-epoxide (E-CBZ) and 5-(4-hydroxyphenyl)-5-phenylhydantoin (HPPH), respectively. Zebrafish embryos were exposed to three concentrations (31.25, 85, and 250 μM) of CBZ and PHE from 51⁄4 to 120 hours post fertilization (hpf) at 28.5°C under a 14/10 hour light/dark cycle. For species comparison, also adult zebrafish, rat, rabbit and human liver microsomes (200 μg/ml) were exposed to 100 μM of CBZ or PHE for 240 minutes at 28.5°C. Potential formation of the mammalian metabolites was assessed in the embryo medium (48, 96, and 120 hpf); pooled (n=20) whole embryos/larvae extracts (24 and 120 hpf); and in the microsomal reaction mixtures (at 5 and 240 minutes) by targeted investigation using a UPLC–Triple Quadrupole MS system with lamotrigine (0.39 μM) as internal standard. Our study showed that zebrafish embryos metabolize CBZ to E-CBZ, but only at the end of organogenesis (from 96 hpf onwards), and no biotransformation of PHE to HPPH occurred. In contrast, our in vitro drug metabolism assay showed that adult zebrafish metabolize both compounds into their active mammalian metabolites. However, significant differences in metabolic rate were observed among the investigated species. These results highlight the importance of including the zebrafish in the in vitro drug metabolism testing battery
for accurate species selection in toxicity studies.
Projects and initiatives
HealthToxicologyInnovationIn vitro
Cells4Thought: using iPSCs for neurodevelopmental health
The prevalence of neurodevelopmental disorders (NDDs), including cognitive impairments, is increasing worldwide with great impact on daily life quality. There is evidence that exposure to chemicals may contribute to the incidence of NDD. However, a causal link is lacking. Towards this goal, a human-relevant in vitro model system mimicking parts of brain development, such as neuronal network functioning, could be used for mechanistic research on how gene-environment interactions contribute to the development of NDD. This is going to be studied in the project Cells4Thought, using induced pluripotent stem cells form different individuals to study the effect of chemicals on neuronal differentiation.
Various subjects
InnovationPolicy
TPI.tv: improving science through animal-free innovations and research
Introducing TPI.tv : a video platform by experts striving to improve science through animal-free innovations and research.
Innovation examples
ToxicologyInnovationIn vitro
Human neuronal cell models for in vitro neurotoxicity screening and seizure liability assessment
Anke Tukker was a PhD candidate in the Neurotoxicology Research group of Dr. Remco Westerink at the Institute for Risk Assessment Sciences at Utrecht University. Dr Westerink’s research group investigates the mechanisms of action of toxic substances on a cellular and molecular level using in vitro systems. Anke's project aimed to develop a human induced pluripotent stem cell (hiPSC)-derived neuronal model for in vitro neurotoxicity screening and seizure liability assessment. Using micro-electrode arrays (MEAs), she showed that these models mimic in vivo neuronal network activity. When these hiPSC-derived neurons are mixed with hiPSC-derived astrocytes, they can be used for in vitro seizure liability assessment. Comparing these data with data obtained from the current used model of ex vivo rodent cortical cultures, she found that these human cells outperform the rodent model. Here research thus contributes towards animal-free neurotoxicity testing.
Anke Tukker has won the public vote of the Hugo van Poelgeest prize 2020 for her research on human neuronal cell models for in vitro neurotoxicity screening and seizure liability assessment.
Neurotoxicology Research Group, IRAS, Utrecht University: https://ntx.iras.uu.nl/NTXatIras
Conferences abstracts
Liquid marbles: a cost-effective platform to generate cardiospheres from co-cultured cardiomyocytes and cardiac fibroblast for disease modelling
Advances in three-dimensional (3D) culture techniques have shown several advantages over 2D cultures, especially by more accurately mimicking the in vivo environment. This has led to improved reproducibility and reliability of experimental results, which are important criteria in disease modelling and toxicity testing. Induced pluripotent stem cells (iPSC) provide an unlimited source for the derivation of all cell types of the adult body, including cardiomyocytes. To improve the current culture methods for multicellular cardiac spheroids, such as the hanging drop method, we explored the use of hydrophobic powders. Fumed silica nanoparticles can be used to encapsulate liquid drops, which could serve as a microenvironment for cell cultures. This microbioreactor stimulates cell coalescence and 3D aggregation while providing optimal gas exchange between the interior and the surrounding environment. Moreover, the properties of liquid marble microbioreactors render them ideal for co-culture experiments. This liquid marble technique has been previously explored and optimized for other cell types. Here we describe a protocol that allows for the derivation of functional cardiac mini organoids, consisting of co-cultured cardiomyocytes and cardiac fibroblasts. These cardiospheres can be valuable for modelling cardiac diseases in vitro and assessing cell interactions to decipher disease mechanisms.
Lab website: https://www.medicalcellbiologylab.com/
Contact: https://www.researchgate.net/profile/Jeffrey-Aalders
RE-place database: https://www.re-place.be/method/liquid-marbles-cost-effective-platform-generate-cardiospheres-co-cultured-cardiomyocytes-and
Innovation examples
HealthInnovationIn vitro
Tumor-on-chips to study delivery of protein therapeutics
Valentina is a PhD candidate at the Department of Biochemistry at Radboudumc. Her research focuses on developing and applying organ-on-chip technologies, such as tumor-on-a-chip systems, to study the tissue-specific and cytosolic delivery of protein therapeutics. Valentina's research has also aimed at bridging the gap between engineers and biologists, promoting the use of microfluidic organ-on-chip technologies to answer more relevant biological questions. One example of this is the development of a mathematical model that could be applied to study drug delivery and diffusion in a tumor-on-a-chip system and to extrapolate possible outcomes of the delivery of therapeutic proteins to tumors in the human body. Another collaboration led to the development of a tumor-on-a-chip where hypoxic conditions can be replicated and investigated, and where the targeting of specific hypoxia markers in tumor cells can be investigated.
Various subjects
Five simple tricks for making your own video for TPI.tv
This video shows you how to make a video yourself. It's really not that difficult! See also https://tpi.tv/how-to-submit for additional information.
Innovation examples
New approaches for cancer hazard assessment
Chemical substances are subjected to assessment of genotoxic and carcinogenic effects before being marketed to protect man and the environment from health risks. For cancer hazard assessment, the long-term rodent carcinogenicity study is the current mainstay for the detection of nongenotoxic carcinogens. However, carcinogenicity studies are shown to have prominent weaknesses and are subject to ethical and scientific debate. A transition toward a mechanism-based weight of evidence approach is considered a requirement to enhance the prediction of carcinogenic potential for chemicals. At RIVM, we are working on this alternative approach for cancer hazard assessment, which makes optimal use of innovative (computational) tools and be less animal demanding.
For more information, click on the link in the video or read on here (https://doi.org/10.1080/10408444.2020.1841732) and here (https://doi.org/10.1080/10408444.2018.1458818).
Contact the expert (https://nl.linkedin.com/in/mirjamluijten)
Innovation examples
HealthToxicologyInnovationIn vitro
Platform for in vitro airborne inhalation testing
The air-liquid interface (ALI) technique uses lung cells cultured on a tiny polymer membrane in a cup. On one side of the membrane is a liquid containing the medium necessary for the cells to survive, while the other side is in contact with air. This is similar to the situation in the human lung. The compound to be tested is administered via an aerosol, vapor, or gas to mimic the situation in human lungs. By monitoring different parameters in the cell model before and after the compound is added, it is possible to measure the effects on lung cells. Depending on the test to be carried out, the lung cells can come from different regions in the respiratory tract and even from a variety of people, including individuals who smoke a lot or have specific diseases such as chronic obstructive pulmonary disease or asthma.
In vitro ALI inhalation testing (https://doi.org/10.1021/acs.est.7b00493) adds value for e.g. pre-clinical trials and research in the pharmaceutical industry and testing (new) compounds for the chemical sector and beyond. The advantages of ALI inhalation testing are that it is a non-animal method, it reduces the use of in vivo experiments, pre-clinical testing with human-derived cell models is more realistic and limits clinical trial failures and it provides faster and more efficient testing of compound
Innovation examples
ToxicologyInnovationIn vitro
Cartilage-on-a-chip for studying joint degenerative diseases
Carlo Alberto Paggi is currently a PhD candidate at the University of Twente in the research group of Prof. Marcel Karperien and Prof. Séverine Le Gac. Karperien’s lab focus on the biological aspects of osteoarthritic research while Le Gac’s specialize in organ-on-chip development. The project of Carlo Alberto is developing a joint-on-chip platform to create a reliable in vitro model to study disease progression in osteo- or rheumatoid arthritis. The model combines different organ-on-chips aimed at replicating each a tissue around the joint such as cartilage, bone and ligaments. This new technology focuses on better reproducing human models and at substituting the use of animal models for drug research. If you want to know something more about the project and the groups, you can follow the link in the video.
Carlo Paggi was nominated for the Hugo van Poelgeest prize for his research on a cartilage-on-a-chip model to study joint degenerative diseases
Karperien’s lab of Developmental Bioengineering: https://www.utwente.nl/en/tnw/dbe/
Le Gac’s lab of Applied Microfluidics for BioEngineering Research: http://www.severinelegac.com/
Linkedin: https://www.linkedin.com/in/carlo-alberto-paggi-76500b135/