Stefan E. Spirig

University of Basel

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Magdalena Renner

Institute of Molecular and Clinical Ophthalmology Basel

Yanyan Hou

University of Basel

Cameron S. Cowan

Institute of Molecular and Clinical Ophthalmology Basel

Adrienn Volak

Institute of Molecular and Clinical Ophthalmology Basel

Botond Roska

Institute of Molecular and Clinical Ophthalmology Basel

Valeria J. Arteaga-Moreta

University of Basel

Simone Picelli

University of Basel

Daniel K. Baeschlin

Novartis (Switzerland)

Jannick Imbach

Institute of Molecular and Clinical Ophthalmology Basel

Alissa Muller

University of Basel

Cooperative Institutions

Institute of Molecular and Clinical Ophthalmology Basel

University of Basel

Friedrich Miescher Institute

Novartis (Switzerland)

Novartis Institutes for BioMedical Research

Massachusetts General Hospital

Harvard University

University Hospital of Basel

Center for Cancer Research

Harvard NeuroDiscovery Center

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Cell Types of the Human Retina and Its Organoids at Single-Cell Resolution

Cell (2020)

Cameron S. Cowan Magdalena Renner Martina De Gennaro Brigitte Gross-Scherf David Goldblum

Human organoids recapitulating the cell-type diversity and function of their target organ are valuable for basic and translational research. We developed light-sensitive human retinal organoids with multiple nuclear and synaptic layers and functional synapses. We sequenced the RNA of 285,441 single cells from these organoids at seven developmental time points and from the periphery, fovea, pigment epithelium and choroid of light-responsive adult human retinas, and performed histochemistry. Cell types in organoids matured in vitro to a stable "developed" state at a rate similar to human retina development in vivo. Transcriptomes of organoid cell types converged toward the transcriptomes of adult peripheral retinal cell types. Expression of disease-associated genes was cell-type-specific in adult retina, and cell-type specificity was retained in organoids. We implicate unexpected cell types in diseases such as macular degeneration. This resource identifies cellular targets for studying disease mechanisms in organoids and for targeted repair in human retinas.

Organoid

Cell type

10.1016/j.cell.2020.08.013

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Citations (476)

Cell type-focused compound screen in human organoids reveals molecules and pathways controlling cone photoreceptor death

bioRxiv (Cold Spring Harbor Laboratory) (2023)

Stefan E. Spirig Valeria J. Arteaga-Moreta Zoltán Raics Susana Posada-Céspedes Stéphanie Chreng

SUMMARY Human organoids that mirror their corresponding organs in cell-type diversity present an opportunity to perform large-scale screens for compounds that protect disease-affected or damage healthy cell types. However, such screens have not yet been performed. Here, we generated 20,000 human retinal organoids with GFP-labeled cone photoreceptors. Since degeneration of cones is a leading cause of blindness, we induced cone death and screened 2,707 compounds with known targets, for those that saved cones or those that further damaged cones. We identified kinase inhibitors that protected cones in both the short and longer term, HSP90 inhibitors that saved cones in the short term but damaged them in the longer term, and broad HDAC inhibition by many compounds that significantly damaged cones. This resource provides a database for cone-damaging compounds, and it describes compounds that can be starting points to develop neuroprotection for cones in diseases such as macular degeneration.

Organoid

Cone (formal languages)

Photoreceptor cell

10.1101/2023.10.09.561525

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Citations (6)

Toward Retinal Organoids in High-Throughput

Cold Spring Harbor Perspectives in Medicine (2023)

Stefan E. Spirig Magdalena Renner

Stefan Erich Spirig1,2 and Magdalena Renner1,2 1Institute of Molecular and Clinical Ophthalmology Basel, Basel, Switzerland 2Department of Ophthalmology, University of Basel, Basel, Switzerland Correspondence: magdalena.renner{at}iob.ch

Organoid

10.1101/cshperspect.a041275

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Citations (3)

PEGDA-based HistoBrick for increasing throughput of cryosectioning and immunohistochemistry in organoid and small tissue studies

Scientific Reports (2025)

Emilie Vuille-dit-Bille Larissa Utz Fiona E. Müllner Valeria J. Arteaga-Moreta Yanyan Hou

Histology is the gold standard for analyzing tissue structure and cell morphology. Immunostaining on thin tissue sections enables precise visualization of antigens and proteins. However, for cryosectioning small tissues such as organoids, spheroids, and tumoroids there is a lack of standardized, time- and cost-effective methods, limiting the throughput of analysis. Here, we have adapted to cryosectioning our previously developed HistoBrick approach, in which small tissue arrangement is spatially controlled within arrayed mini-wells. By testing various embedding matrices, we show that an 8% PEGDA and 2.5% gelatine mixture is optimal, providing essential structural support to maintain sample integrity during cryosectioning. This embedding matrix preserves fragile substructures of human retinal organoids, which are particularly susceptible to damage during sample preparation. Using PEGDA-gelatine HistoBricks for the simultaneous embedding of 16 retinal organoids, we analyzed a time course of retinal organoid development. We observed the maintenance of photoreceptors cell bodies up to week 98 in culture, while photoreceptor outer segments were gradually lost. Further, we observed displaced photoreceptors in the region of outer segments. The PEGDA-gelatine HistoBrick is a cost-efficient tool that can be implemented for small tissue studies to increase throughput in experiments such as large-scale screenings or toxicology research.

Organoid

Immunostaining

10.1038/s41598-024-83164-2

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High levels of AAV vector integration into CRISPR-induced DNA breaks

Nature Communications (2019)

Killian S. Hanlon Benjamin P. Kleinstiver Sara P. Garcia Mikołaj Zaborowski Adrienn Volak

Abstract Adeno-associated virus (AAV) vectors have shown promising results in preclinical models, but the genomic consequences of transduction with AAV vectors encoding CRISPR-Cas nucleases is still being examined. In this study, we observe high levels of AAV integration (up to 47%) into Cas9-induced double-strand breaks (DSBs) in therapeutically relevant genes in cultured murine neurons, mouse brain, muscle and cochlea. Genome-wide AAV mapping in mouse brain shows no overall increase of AAV integration except at the CRISPR/Cas9 target site. To allow detailed characterization of integration events we engineer a miniature AAV encoding a 465 bp lambda bacteriophage DNA (AAV-λ465), enabling sequencing of the entire integrated vector genome. The integration profile of AAV-465λ in cultured cells display both full-length and fragmented AAV genomes at Cas9 on-target sites. Our data indicate that AAV integration should be recognized as a common outcome for applications that utilize AAV for genome editing.

Transduction (biophysics)

Adeno-associated virus

Genome Engineering

10.1038/s41467-019-12449-2

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Citations (327)

Corrigendum: Toward Retinal Organoids in High-Throughput

Cold Spring Harbor Perspectives in Medicine (2023)

Stefan E. Spirig Magdalena Renner

Organoid

10.1101/cshperspect.a041664

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Citations (0)