Neurotoxicity

Neuroinflammation, a transient neuroprotective state, can when chronic transform into a self-propagating and potentially neurodegenerative state. Recent studies suggest that neuroinflammation is an early, or even a causative, event in the development of Alzheimer's and Parkinson's diseases. The mechanisms underlying the switch from protective/reparative neuroinflammation to a pro-degenerative function is not well understood and its characterization poses one promising venue for better assessment of neurotoxic hazard of compounds.

Neuroinflammation is a brain-specific response to a variety of cues including exposure to environmental pollutants. This research area utilizes in vitro models to identify and characterise key events leading to the switch of neuroinflammatory state from neuroprotective to neurodegenerative. In particular, the focus is on passage through the blood-brain barrier and metabolic reprogramming of brain cells occurring during the inflammatory process. Early changes in proteins and metabolite levels linked to changes in brain cell metabolism will be evaluated for their utility as early markers of toxicity, with the ultimate goal of measuring them in human biological fluids.

The identification of human biomarkers of neuroinflammation that can be measured in biological fluids could aid in assessing onset of potentially neurodegenerative inflammation. Furthermore, the obtained data will be used for the development of Adverse Outcome Pathways (AOPs), that will be useful for risk assessment and finally for regulatory purposes. 

WP1 - Human in vitro blood-brain barrier model

Rationale for the project

The blood-brain barrier (BBB) controls and limits the movement of substances between blood and brain and possesses active properties that enable cells to pump unwanted molecules back into the bloodstream. In addition, the BBB has a metabolic function that can alter the chemical properties of those molecules which do get into the brain. The failure of BBB structural integrity and function can be directly induced by chemicals or indirectly by neuroinflammation, which seems to play a pivotal role in the pathogenesis of many diseases of the central nervous system. The aim of the project is to use in vitro models of the blood-brain barrier and brain-like neural tissues to dissect out the mechanisms which are involved in BBB-related neuroinflammation.

Objectives

The objective of this project is to assess the ability of xenobiotics to cross the BBB, to disrupt it by inducing a neurovascular inflammatory response, and to cause neurotoxicity, using an innovative one step in vitro screening technology based on neural tissue derived from human stem cells. This approach will combine expertise in tissue engineering of human stem cells and expertise in electrophysiological technologies.

Partners: Serge Rudaz (University of Geneva), Jean-Charles Sanchez (University of Geneva)

WP2 - Metabolic perturbations of glial cell during neuroinflammation

Rationale for the project

Several classes of neurotoxic compounds have been shown to induce neuroinflammation, and evidence is accumulating that neuroinflammation is involved in the pathogenesis of neurodegenerative diseases. In response to environmental stressors, inflammatory responses metabolic adaptations have been shown in reactive astrocytes and microglial cells. Astrocytic metabolic and inflammatory changes have recently been reported as a function of age, leading to the hypothesis that mitochondrial metabolism and inflammatory responses are interconnected and promote the functional switch of astrocytes from neurotrophic to neurotoxic.

Objectives

The first objective of this work package is to describe the changes in glial cellular metabolism (metabolic reprogramming of astrocytes) which occur during neuroinflammation triggered by selected chemicals, and the relationship between these changes and neurodegeneration. The second objective is to establish the extracellular vesicle (EV) content signatures of neuroinflammation. It will then be determined whether the early metabolic changes in glial cells and/or EV signatures could be used as early in vitro biomarkers of neurotoxicity.

Partners: Luc Pellerin (University of Lausanne), Serge Rudaz (University of Geneva), Jean-Charles Sanchez (University of Geneva)

WP3 - Metabolomic profile alteration by neuroinflammatory conditions

Rationale for the project

Neurodegenerative disorders such as Parkinson or Alzheimer’s diseases are an increasing public health burden. More information on the possible role of environmental chemical exposure in these diseases would be very valuable for risk managers. OECD Adverse Outcome Pathways (AOPs) for neuroinflammation and neurodegeneration link exposure to various chemicals to the shared key event “neuroinflammation” and then to neurodegeneration. Currently, it is quite difficult to quantify neuroinflammation. More efficient markers are needed.

The aim of the project is to develop improved biomarkers of neuroinflammation using metabolomic data. Metabolomics aim at the identification and the quantification of all metabolites (m/z < 1000 Da) in a biological system; it constitutes a powerful technique to establish profiles and characterise the effects of disease and toxic chemical exposures.

Objectives

Analytical conditions to separate and analyse the neurotransmitters will be optimised. Although reverse-phase Liquid Chromatography (LC) constitutes one of the gold standards in metabolomics, the use of orthogonal separation mechanisms with different selectivity is currently broadening the achievable coverage of small molecules. Among them, HILIC fits remarkably well the separation performance required by polar compounds such as neurotransmitters and similar small polar metabolites. Once the complete analytical platform will be validated, a qualification of the normal metabolome for each model will be established.  Neural cells (donated by core project partners) will be exposed to the neuroinflammatory toxicants at various doses and exposure times. For untargeted metabolomics, cell contents will be analysed using LC-High Resolution Mass Spectrometry (HRMS). Specific metabolic patterns will be characterised by the analysis of individual variable contributions to each effect. Metabolomics patterns will be analysed in combination with proteomics data using system biology integrative software. 

Partners: Luc Pellerin (University of Lausanne), Luc Stoppini (Hepia), Marie-Gabrielle Zurich (University of Lausanne)

WP4 - Proteomics strategies for discovering markers of neuroinflammation and neurotoxicity

Rationale for the project

Proteomics has been a rapidly growing technology-driven science and is now essential in any biomedical research study. It uses specific mass spectrometry instruments to detect, identify and quantify multiple proteins in very complex biological samples in time and space. Proteomics is performed in the context of when and where proteins are expressed and how they operate as single entities, as functional modules, and as part of networks that control and relay biological information. The widespread use of this enabling strategy and its application to the toxicity sciences is set to have a huge impact on the understanding of processes which are activated or repressed in the early stage of neuroinflammation.

Objectives

The objective of this project is to discover proteins and pathways using proteomics strategies, to identify early biomarkers (key event readouts) of neurotoxicity. This will be performed on different types of neurovasculature unit (NVU) and brain parenchyma cell cultures exposed to model molecules (TNFα or IL6) and neurotoxicants (trimethyltin, paraquat and morphine).

Partners: Serge Rudaz (University of Geneva), Luc Stoppini (Hepia), Marie-Gabrielle Zurich (University of Lausanne)

WP5 - Extracellular vesicles as markers of neuroinflammation and neurotoxicity

Rationale for the project

In the field of biomarker research, extracellular vesicles (EVs) such as microparticles and exosomes are potential biomarker holders that avoid the problem of the wide dynamic concentration range of blood molecules. EVs are circulating vesicles released into the blood stream from almost all cell types, and are composed of a large number of biomolecules such as mRNAs, miRNAs, proteins, metabolites and phospholipids. EVs are therefore potential early indicators of a key event, containing precious information for the monitoring of pathologies. Xenobiotic exposure can trigger key events that converge in neuroinflammation and ultimately neurodegenerative disorders, such as Parkinson's and Alzheimer's disease.

Objectives

The objective of this project is to establish the extracellular vesicles (EVs) signature of neuroinflammation in order to identify early biomarkers (KE readouts) of neurotoxicity. This will be performed using a co-culture blood-brain barrier (BBB) model with human endothelial cells, astrocytes and microglia exposed to neurotoxicants (TNFα, trimethyltin, paraquat and morphine).

Partners: Serge Rudaz (University of Geneva), Luc Stoppini (Hepia), Marie-Gabrielle Zurich (University of Lausanne)