CelltypeR: A flow cytometry pipeline to characterize single cells from brain organoids

Rhalena A. Thomas; Julien Sirois; Shuming Li; Alexandre Gestin; Ghislaine Deyab; Valerio E.C. Piscopo; Paula Lépine; Meghna Mathur; Carol X.-Q. Chen; Vincent Soubannier; Taylor M. Goldsmith; Lama Fawaz; Thomas M. Durcan; Edward A. Fon

iScience (Sep 2024)

CelltypeR: A flow cytometry pipeline to characterize single cells from brain organoids

Rhalena A. Thomas,
Julien Sirois,
Shuming Li,
Alexandre Gestin,
Ghislaine Deyab,
Valerio E.C. Piscopo,
Paula Lépine,
Meghna Mathur,
Carol X.-Q. Chen,
Vincent Soubannier,
Taylor M. Goldsmith,
Lama Fawaz,
Thomas M. Durcan,
Edward A. Fon

Affiliations

Rhalena A. Thomas: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada; Corresponding author
Julien Sirois: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Shuming Li: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Alexandre Gestin: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; Université Paris-Saclay, 91190 Gif-sur-Yvette, France
Ghislaine Deyab: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada
Valerio E.C. Piscopo: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Paula Lépine: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Meghna Mathur: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Carol X.-Q. Chen: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Vincent Soubannier: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Taylor M. Goldsmith: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Lama Fawaz: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada
Thomas M. Durcan: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada
Edward A. Fon: Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, QC H3A 2B4, Canada; The Neuro's Early Drug Discovery Unit (EDDU), McGill University, Montreal, QC H3A 2B4, Canada

Journal volume & issue: Vol. 27, no. 9
p. 110613

Abstract

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Summary: Motivated by the cellular heterogeneity in complex tissues, particularly in brain and induced pluripotent stem cell (iPSC)-derived brain models, we developed a complete workflow to reproducibly characterize cell types in complex tissues. Our approach combines a flow cytometry (FC) antibody panel with our computational pipeline CelltypeR, enabling dataset aligning, unsupervised clustering optimization, cell type annotating, and statistical comparisons. Applied to human iPSC derived midbrain organoids, it successfully identified the major brain cell types. We performed fluorescence-activated cell sorting of CelltypeR-defined astrocytes, radial glia, and neurons, exploring transcriptional states by single-cell RNA sequencing. Among the sorted neurons, we identified subgroups of dopamine neurons: one reminiscent of substantia nigra cells most vulnerable in Parkinson’s disease. Finally, we used our workflow to track cell types across a time course of organoid differentiation. Overall, our adaptable analysis framework provides a generalizable method for reproducibly identifying cell types across FC datasets in complex tissues.

Published in iScience

ISSN: 2589-0042 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science
Website: http://www.cell.com/iscience/home

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