Integration of Genetics, Environment, and Time (G x E x T) for Neurodevelopmental Toxicology and Risk Assessment
Park, Ju Young
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Processes of neurodevelopment are complex and highly orchestrated, making the brain development one of the most sensitive time frames for perturbations. Because of this complexity and sophistication, in vivo studies have been conducted traditionally for neurodevelopmental toxicity. With a movement to save the number of experimental animals used, neurodevelopmental toxicology is an area where there is a pressing need for in vitro alternative methods development. In this research, two in vitro neurodevelopmental models were evaluated: a primary fetal mouse midbrain micromass system and a human neural progenitor cell culture. Dynamics of C57BL/6 and A/J micromass cultures over 22 days in vitro (DIV) were characterized. Changes in morphology, protein content, the extent of differentiation, and stage-specific proteins were assessed over time, which revealed similar dynamics to in vivo. The second in vitro model used differentiating human neural progenitor cells (hNPCs). Without any additional external morphogens, hNPCs are thought to differentiate towards forebrain identity over 21 days in culture. When gene ontology (GO) processes and relative gene expression intensity of genes associated with GO terms of interest were compared using gene expressions obtained from microarray, our hNPCs showed high concordance with in vivo human neocortical brain tissue. Together, these observations defined the potential of two in vitro neurodevelopmental models as a screening tool for prioritizing compounds. Using characterized in vitro neurodevelopmental models, factors that may contribute to susceptibility were evaluated. Exposures to silver nanoparticles (AgNPs) with different sizes and coating on C57BL/6 or A/J micromass cultures at three different developmental stages illustrated differential yet significant adverse effects of particle coatings, sizes, and time of exposures on cytotoxicity. Genetic backgrounds also contributed to differential responses to AgNPs. Exposures to chlorpyrifos, arsenic, and cadmium on proliferating and differentiating hNPCs in vitro revealed compound- and stage-specific impacts on cell viability and epigenetics. AgNPs-exposed hNPCs showed that particle coatings and sizes significantly contributed to cell viability. We found that fetal mouse midbrain cells and hNPCs were generally most sensitive to various types of compounds during early differentiation when differentiation is initiated, but proliferation signaling was still active. Moreover, in vitro hNPCs were differentially sensitive to AgNPs from embryonic mouse midbrain cells, suggesting potential regional-specific responses to AgNPs. Altogether, the results suggest that factors including developmental stages, genetics, and region of the brain contribute to susceptibility; therefore, the impacts of genetics/epigenetics and time of exposures on neurodevelopment also need to be assessed to better define response-disease pathway that can later be applied in risk assessments and regulatory decisions.
- Environmental health