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bioRxiv · 10.1101/2025.04.23.650161

Cerebral Organoids with Integrated Endothelial Networks Emulate the Neurovascular Unit and Mitigate Core Necrosis

Fumado Navarro, J.·Crilly, S.·Chan, W. K.·Browne, S.·Mason, J.·Giraldo, C. V.·Pandit, A.·Lomora, M.
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Abstract

Cerebral organoids (COs) are multicellular, self-organized, in vitro, 3D brain-like tissues used for developmental biology, disease modelling and drug screening. However, their lack of vascularity renders them less physiologically accurate. Vascularization of COs remains challenging due to the different requirements between COs and vascular cells, limited vascular network penetration within the organoid, and the absence of luminal perfusion. Here, we devised an encapsulation approach in which human brain microvascular endothelial cells (HBMVECs) were delivered to developing COs from progressively degrading extracellular matrix (ECM)-based hydrogel droplets. By tuning this hydrogel concentration and media composition, we observed enhanced vascular-like network formation that expanded within the organoid tissue. Using pathway inhibitors, we showed that a subset of the endothelial cells (ECs) originated from the CO itself, promoting network integration. Endothelial networks displayed blood-brain barrier (BBB) features, including astrocytic end-footlike interactions, pericyte wrapping, and collagen-laminin basal lamina. Vascularized COs exhibited greater media internalization and up to three-fold lower apoptosis than non-vascularized COs. This comprehensive 3D neurovascular model is a promising platform for cerebrovascular research and drug testing applications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/650161v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@175edb7org.highwire.dtl.DTLVardef@1499b91org.highwire.dtl.DTLVardef@184567borg.highwire.dtl.DTLVardef@149c8e4_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG HighlightsO_LIAddition and angiogenic stimulation of human brain microvascular endothelial cells (HBMVECs) co-cultured with cerebral organoids (COs) generate multicellular vascular-like networks C_LIO_LIVascularization induces changes in organoid morphology but not on tissue stiffness C_LIO_LIEndothelial networks morphological features are correlated with the concentration of the supporting matrix C_LIO_LIA number of the endothelial cells (ECs) in the networks originate from the organoid itself C_LIO_LIEndothelial networks integrate within the organoid tissue interacting with astrocytes and pericyte-like cells, and are surrounded by basement membrane-like depositions C_LIO_LIVascularized COs exhibit higher media diffusion, and a reduced necrotic core compared to non-vascularized COs C_LI

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Fumado Navarro, J., Crilly, S., Chan, W. K., Browne, S., Mason, J., Giraldo, C. V., Pandit, A., Lomora, M.. 2025-04-26. Cerebral Organoids with Integrated Endothelial Networks Emulate the Neurovascular Unit and Mitigate Core Necrosis. https://doi.org/10.1101/2025.04.23.650161

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