Toward Spatial Identities in Human Brain Organoids-on-Chip Induced by Morphogen-Soaked Beads

Abstract

Recent advances in stem-cell technologies include the differentiation of human embryonic stem cells (hESCs) into organ-like structures (organoids). These organoids exhibit remarkable self-organization that resembles key aspects of in vivo organ development. However, organoids have an unpredictable anatomy, and poorly reflect the topography of the dorsoventral, mediolateral, and anteroposterior axes. In vivo the temporal and the spatial patterning of the developing tissue is orchestrated by signaling molecules called morphogens. Here, we used morphogen-soaked beads to influence the spatial identities within hESC-derived brain organoids. The morphogen- and synthetic molecules-soaked beads were interpreted as local organizers, and key transcription factor expression levels within the organoids were affected as a function of the distance from the bead. We used an on-chip imaging device that we have developed, that allows live imaging of the developing hESC-derived organoids. This platform enabled studying the effect of changes in WNT/BMP gradients on the expression of key landmark genes in the on-chip human brain organoids. Titration of CHIR99201 (WNT agonist) and BMP4 directed the expression of telencephalon and medial pallium genes; dorsal and ventral midbrain markers; and isthmus-related genes. Overall, our protocol provides an opportunity to study phenotypes of altered regional specification and defected connectivity, which are found in neurodevelopmental diseases.

Atypical Teratoid Rhabdoid Tumours Are Susceptible to Panobinostat-Mediated Differentiation Therapy

“…Detailed inspection of these functional groups further revealed enrichment of genes that are essential in nervous system development (Figure 4a,b). Several key promoters and regulators in central nervous system development (Nitric Oxide Synthase 1 (NOS1) [33][34][35][36]), neuron differentiation (T-box Transcription Factor 3 (TBX3) [37], Desmoplakin 3 (DPF3) [38,39], Bone Morphogenetic Protein 4 (BMP4) [37,40]) and neuronal development (Neurexin 2 (NRXN2) [41,42], Ataxin 1 (ATXN1) [43,44]) were upregulated, supporting neuronal differentiation [45] (Figure 4a,b). Additionally, there was also a prevalence of functional groups associated with cell division regulation including the mitotic cell cycle, nuclear division, regulation of metaphase/anaphase transition of the cell cycle, regulation of chromosome separation and spindle organization (Figure 3c,d).…”

Engineering Brain Organoids: Toward Mature Neural Circuitry with an Intact Cytoarchitecture

“…The degree of signaling may differ based on the concentration of the signaling molecules absorbed in the beads or the distance from the beads, through which a signaling gradient is established with diffused morphogens. Indeed, a recent study utilized a WNT agonist- and BMP4-soaked beads to successfully direct dorsoventral and anteroposterior topography in brain organoids ( 41 ). The spatial topography was determined by the concentration and position of the beads, in which stronger exposure to WNT and BMP yields more anterior/rostral molecular signatures.…”

Modeling Neurological Disorders in 3D Organoids Using Human-Derived Pluripotent Stem Cells

“…While glial progenitors are generated by the end of the 3rd month, another 3 months are required to generate functional astrocytes (Krencik and Zhang, 2011). Recently, some researchers recommend using protocols that differentiate astrocytes from hiPSCs with the expression of GFAP and S100β more quickly (Palm et al, 2015;Ben-Reuven and Reiner, 2020;Soubannier et al, 2020) (Table 2). Like neurons, astrocytes are also classified into many subtypes because of their location, morphology, and molecular /physiological functions.…”

“…While different protocols (Bagley et al, 2017 ; Wang et al, 2018 ) were established to develop the D-V and A-P axis in human brain organoids, morphogen-loaded beads were found to be more effective in establishing spatial identities in human brain organoids (Ben-Reuven and Reiner, 2020 ). In this method, organoids with morphogen-soaked beads were co-cultured to diffuse morphogens from the beads to the tissue, which create a concentration gradient of morphogen secretions in the developing organoids.…”