Abstract
Toxicity testing is a crucial step in the development and approval of chemical compounds for human contact and consumption. However, existing model systems often fall short in their prediction of human toxicity in vivo because they may not sufficiently recapitulate human physiology. The complexity of three-dimensional (3D) human organ-like cell culture systems (“organoids”) can generate potentially more relevant models of human physiology and disease, including toxicity predictions. However, so far, the inherent biological heterogeneity and cumbersome generation and analysis of organoids has rendered efficient, unbiased, high throughput evaluation of toxic effects in these systems challenging. Recent advances in both standardization and quantitative fluorescent imaging enabled us to dissect the toxicities of compound exposure to separate cellular subpopulations within human organoids at the single-cell level in a framework that is compatible with high throughput approaches. Screening a library of 84 compounds in standardized human automated midbrain organoids (AMOs) generated from two independent cell lines correctly recognized known nigrostriatal toxicants. This approach further identified the flame retardant 3,3′,5,5′-tetrabromobisphenol A (TBBPA) as a selective toxicant for dopaminergic neurons in the context of human midbrain-like tissues for the first time. Results were verified with high reproducibility in more detailed dose-response experiments. Further, we demonstrate higher sensitivity in 3D AMOs than in 2D cultures to the known neurotoxic effects of the pesticide lindane. Overall, the automated nature of our workflow is freely scalable and demonstrates the feasibility of quantitatively assessing cell-type-specific toxicity in human organoids in vitro.
Highlights
All living things, humans included, are exposed to a plethora of natural and artificial compounds on a daily basis
Compounds can have distinct effects on specific cell subpopulations of complex tissues in vivo, which may not be detectable in standard cellular assays that rely on monocultures of single cell lines or cell types (Kasurinen et al, 2018)
We have recently developed a highly homogeneous and reproducible 3D model system of the human midbrain (‘‘automated midbrain organoids’’, AMOs), which is designed for high throughput screening applications (Renner et al, 2020)
Summary
Humans included, are exposed to a plethora of natural and artificial compounds on a daily basis. Well-established protocols based on induced pluripotent stem cells (hiPSCs) (Takahashi et al, 2007) have provided virtually unlimited access to a multitude of human cell types, including those that reflect genotypes and phenotypes from patients with specific diseases (Nirmalanandhan and Sittampalam, 2009; Krewski et al, 2011; Friese et al, 2019; Fritsche et al, 2020) These typically two-dimensional (2D) cell culture models may not recreate the specific and diverse cell populations of human tissues in vivo with their intricate architectures and interdependent molecular interactions, potentially limiting their predictive value for human toxicity (Bus and Becker, 2009; Hartung and Daston, 2009; Horvath et al, 2016). 3D models may better recapitulate the native tissue niche of cells (Duval et al, 2017; Ho et al, 2018)
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