Engineering human cerebral organoids to explore mechanisms of arsenic-induced developmental neurotoxicity.

Abstract

Modeling brain development and function is challenging due to complexity of the organ. Human pluripotent stem cell (PSC)-derived brain-like organoids provide new tools to study the human brain. Compared with traditional in vivo toxicological studies, these 3D models, together with 2D cellular assays, enhance our understanding of the mechanisms of developmental neurotoxicity (DNT) during the early stages of neurogenesis and offer numerous advantages including a rapid, cost-effective approach for understanding compound mechanisms and assessing chemical safety. Arsenic (As) exposure is associated with DNT, although the mechanisms involved are not well-defined. Here, we used 3D PSC-derived embryoid bodies (EBs) to recapitulate events involved in embryogenesis and neurogenesis before neural induction, and EB-derived cerebral organoids to mimic neural development in vivo. As (0.5 μM; 35 ppb) increased ectoderm differentiation within the EBs by upregulating genes (PAX6, SOX1) critical for embryonic development. Histological staining of EBs showed As disrupted neural rosette structures. qPCR and RNA-seq showed As inhibited expression of markers of mature neural cells (MAP2+/vGLUT2+) and astrocytes (GFAP+). In organoids, Ingenuity Pathway Analysis was used to identify the top 5 pathways affected by As exposure, and Gene Ontology enrichment analysis found several key signaling pathways to be inhibited by As exposure. These data provide insights into pathways contributing to As-induced inhibition of neurite outgrowth and disrupted neural rosette structures in the 2D neurite outgrowth assay and in organoids, respectively. Results herein show this multipronged 2D/3D approach can provide valuable insights into cellular events and molecular mechanisms of As-induced DNT.