The Future of 3D Brain Cultures in Developmental Neurotoxicity Testing

Abstract

Human brain is undoubtedly the most complex organ in the body. Thus, it is difficult to develop adequate and at the same time human relevant test systems and models to cover the aspects of brain homeostasis and even more challenging to address brain development. Animal tests for Developmental Neurotoxicity (DNT) have been devised, but because of complex underlying mechanisms of neural development, and interspecies differences, there are many limitations of animal-based approaches. The high costs, high number of animals used per test and technical difficulties of these tests are prohibitive for routine DNT chemical screening. Therefore, many potential DNT chemicals remain unidentified. New approach methodologies (NAMs) are needed to change this. Experts in the field have recommended the use of a battery of human in vitro tests to be used for the initial prioritization of high-risk environmental chemicals for DNT testing. Microphysiological systems (MPS) of the brain mimic the in vivo counterpart in terms of cellular composition, recapitulation of regional architecture and functionality. These systems amendable to use in a DNT test battery with promising features such as (i) complexity, (ii) closer recapitulation of in vivo response and (iii) possibility to multiplex many assays in one test system, which can increase throughput and predictivity for human health. The resent progress in 3D brain MPS research, advantages, limitations and future perspectives are discussed in this review.

Oligodendrogenesis and myelination tracing in a CRISPR/Cas9-engineered brain microphysiological system

“…Current in vivo approaches are prohibitively expensive, time-consuming, and have low predictivity for humans ( Smirnova et al, 2014 ; Meigs et al, 2018 ). OLs migration, differentiation, and axonal myelination are key events of neural development but are very difficult to recapitulate in vitro using traditional monolayer cultures ( Marangon et al, 2021 ; Hogberg and Smirnova, 2022 ). Therefore, the PLP1 gene, an OL marker, was selected to be tagged with sfGFP to trace OL differentiation and migration during brain organoid maturation.…”

“…As access to human CNS samples is limited, the development of human in vitro models that recapitulate key events of OL biology in their native microenvironment is essential. Advances in stem cell research have paved the road for the development of human microphysiological systems (MPS) such as 3D organoids and organs-on-chip, which more closely resemble human cell physiology compared to traditional monocultures or animal models ( Alépée et al, 2014 ; Jensen and Teng, 2020 ; Marx et al, 2020 ; Hogberg and Smirnova, 2022 ). To this end, our lab and others have developed hiPSC-derived bMPS models with efficient and reproducible differentiation and maturation of neurons and supporting CNS cells (to name a few: Pamies et al, 2017a ; Sloan et al, 2018 ; Albanese et al, 2020 ; Ahn et al, 2021 ).…”

Recent advances and current challenges of new approach methodologies in developmental and adult neurotoxicity testing

“…Therefore, considering this definition, an organoid has three characteristics: (i) it is spatially organized in a way that resembles a human organ, not only at the cellular level, but also in terms of tissue structure and developmental trajectory (ii) it contains several organ-specific cell types, and (iii) it recapitulates a specific function. The main difference between spheroids and organoids is that the former typically lack distinctive cytoarchitecture (Hogberg and Smirnova 2022 ). The main pros and cons of 2D versus 3D cell culture methods are reported in Table 4 .…”

High-Throughput Screening Assay for Detecting Drug-Induced Changes in Synchronized Neuronal Oscillations and Potential Seizure Risk Based on Ca2+ Fluorescence Measurements in Human Induced Pluripotent Stem Cell (hiPSC)-Derived Neuronal 2D and 3D Cultures

“…Recently, human induced pluripotent stem cell-derived neurons (hiPSC-neurons) have emerged as a promising human-derived neuronal in vitro platform that might be useful for the early de-risking phase of preclinical safety evaluations [ 13 , 14 , 15 ]. hiPSC-derived neurons co-cultured in 3D spheroids have been suggested to be amenable to phenotypic in vitro profiling use as well [ 16 , 17 , 18 ]. Applying such approach could increase throughput and reduce concerns about species differences between preclinical assays and outcomes in humans and, thus, be consistent with the 3Rs principles.…”