Generation of Blood Vascular Endothelial-Neural 3D Organoids by Serial Induction of Differentiation on Human iPSC-Derived Embryoid Bodies.

Overview

1. Translational Neuroscience Center, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, (J.P.S.)
2. Cancer Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA
3. Bioinformatics Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA
4. Cell Biology and Gene Expression Section, National Institute on Aging, National Institutes of Health, Bethesda, MD 20892, USA
5. Section of Infections of the Nervous System, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA

Abstract

Highlights: What are the main findings?

We reconstitute a 3D organoid model by serially inducing endothelial and neural differentiation to mimic the in vivo development of neural and endothelial cells.

The resulting 3D organoid consists of a variety of neurons, glial and endothelial cells with vascular-like structures, comparable to the human brain.

What is the implication of the main findings?

This EC-neural organoid model provides a useful tool to study the interactions between vascular endothelial cells and neural cells in early brain development.

These EC-neural organoids can potentially be used to model neural infectious disorders where endothelial cells are targets and used as mediators of neuronal damage.

Abstract: The 3D brain organoids have been widely used as a tool to study human brain development and disorders. Although angiogenesis and blood vascular endothelial cells play important roles in brain development and pathogenesis in neurological disorders, most 3D brain organoids lack inherent endothelial cells and need either the addition of differentiated endothelial cells or to be transplanted to animals to reconstitute such vascular structures. However, these approaches could miss the developmental interactions between angiogenesis and neurogenesis in the human brain. To reconstitute a 3D organoid mimicking the in vivo development of neural and vascular endothelial cells, we cultured iPSC-derived embryoid bodies and sequentially applied endothelial and neuronal induction media along with Matrigel embedding. The resulting 3D organoid consists of both neural cells and endothelial cells with vascular-like structures, as determined by immunostaining. With scRNA-Seq analysis, the organoid was confirmed to contain neural cell types similar to human brains, including a variety of excitatory and inhibitory neurons and glia. Furthermore, when compared with conventionally generated cerebral organoids without endothelial cells using RNA-Seq analysis, the vascular endothelial-containing neural organoids (EC-neural organoids) showed different gene profiles and favored angiogenesis and vasculogenesis. Of the differentially expressed genes, KRBA2 expression was found to be higher in neural cells and its inhibition by siRNA treatment resulted in decreased transcriptions of a variety of genes specific to neuronal differentiation but not genes specific to pluripotent stem cells such as OCT4. The EC-neural organoids also expressed receptors to SARS-CoV-2 at levels similar to human brains. This 3D EC-neural model provides a useful tool to study the interactions between vascular endothelial cells and neural cells in brain development and potentially for the study of neural infectious disorders where vascular endothelial cells are targets for infection and mediators for neural damage.

Reproduced under the paper's license (CC BY), from the paper cited above.

Code

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Data

Datasets cited

• geo:GSE330092, at NCBI GEO; found in “Data Availability Statement”

Data Availability Statement

All relevant data supporting the key findings of this study are available within the article and its Supplementary Information files. The RNA-Seq data are deposited in the public domain: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE330092, accessed on 15 June 2026. Software used in this study includes 10X Genomics Loupe Browser 4.0.0 (https://support.10xgenomics.com, accessed on 21 December 2021).

Reproduced under the paper's license (CC BY), from the paper cited above.

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Version 1, 27 September 2026: the first record

Recorded: type, language, journal, volume, issue, pages, dates, 10 authors, 5 keywords, 11 MeSH terms, 1 funder, 50 references.

Cite

This paper

Wang, T., Bagnell, A., McDonald, V., Gastfriend, B. D., Steiner, J. P., Elkahloun, A. G., Johnson, K., Langston, R. G., Cookson, M. R., & Nath, A. (2026). Generation of Blood Vascular Endothelial-Neural 3D Organoids by Serial Induction of Differentiation on Human iPSC-Derived Embryoid Bodies. Cells, 15(13), 1192. https://doi.org/10.3390/cells15131192

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