Wednesday, May 14, 2025
5/14/2025
An Engineered Bioprinting Platform to Study Neural Migration in Assembloids - Neural organoids represent a powerful tool for modeling the brain and neurodevelopmental disorders (NDDs), but they are unable to capture the interactions between distinct brain regions, which are critical for later brain development. To capture these important interactions, our team established a novel modular brain organoid system, termed brain assembloids, in which organoids of separate brain regions are combined in a single culture. Using these assembloids, we demonstrated that Timothy syndrome, a rare genetic condition affecting cognitive function, is associated with defects in GABAergic interneuron migration. A key limitation of this new technology is that current assembloid protocols require manual positioning of organoids within liquid media, resulting in poor reproducibility and lack of physiologically-relevant brain geometries and connectivities, while making more complex assembloids (i.e., of 3 or more organoid types) prohibitively laborious. Additionally, individual organoid building blocks are commonly cultured in either Matrigel, a poorly defined, highly variable derivative of mouse sarcoma cells, or as free-floating suspensions, which commonly result in unwanted organoid fusion. This results in high variability of organoid size and quality within and between batches. Here, we propose a novel bioengineering platform to enhance the quality, reproducibility, and complexity of neural assembloids to strengthen their use as physiologically- and spatially-relevant in vitro models of interneuron migration in neurodevelopment and NDDs. We leverage a novel magnetic 3D-bioprinting technology (termed SPOT) to generate assembloids of precisely-controlled spatial orientation, constructing physiologically relevant neural connections, and using well-defined biocompatible polymers to enhance reproducibility in organoid and assembloid size and quality. In Aim 1, we begin by evaluating the hypothesis that increasing the viscosity of the surrounding medium will improve the size and organizational reproducibility of brain organoids from several distinct regions that resemble a mid-gestation developmental stage: cerebral cortex (hCO), medial ganglionic eminence (MGE)-like subpallium (hSO), and lateral ganglionic eminence (LGE)-like striatum (hStrO). We will use SPOT to print two-part assembloids of various combinations and evaluate interneuron migration and cortical neuron projections. In Aim 2, we will use SPOT to print three-part assembloids with precise spatial positioning between the MGE-, LGE-, and cortex-like organoids. We hypothesize that reproducible control over geometric positioning of the individual organoids will significantly improve the reliability of interneuron path-finding, allowing this human microphysiological system to be used for NDD studies. In Aim 3, we leverage our three-part assembloids to study the effects of several NDD-related genes on interneuron migration and validate the utility of our bioengineered platform. This transformational work will provide an advanced platform for the controlled construction of complex assembloids, enabling unprecedented molecular and functional studies into human brain development and diseases.
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