Exploring the human lacrimal gland using organoids and single-cell sequencing
Bannier-Hélaouët, Marie; Post, Yorick; Korving, Jeroen; Trani Bustos, Marc; Gehart, Helmuth; Begthel, Harry; Bar-Ephraim, Yotam E.; van der Vaart, Jelte; Kalmann, Rachel; Imhoff, Saskia M.; Clevers, Hans
(2021) Cell stem cell, volume 28, issue 7, pp. 1221 - 1232.e7
(Article)
Abstract
The lacrimal gland is essential for lubrication and protection of the eye. Disruption of lacrimal fluid production, composition, or release results in dry eye, causing discomfort and damage to the ocular surface. Here, we describe the establishment of long-term 3D organoid culture conditions for mouse and human lacrimal gland. Organoids
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can be expanded over multiple months and recapitulate morphological and transcriptional features of lacrimal ducts. CRISPR-Cas9-mediated genome editing reveals the master regulator for eye development Pax6 to be required for differentiation of adult lacrimal gland cells. We address cellular heterogeneity of the lacrimal gland by providing a single-cell atlas of human lacrimal gland tissue and organoids. Finally, human lacrimal gland organoids phenocopy the process of tear secretion in response to neurotransmitters and can engraft and produce mature tear products upon orthotopic transplantation in mouse. Together, this study provides an experimental platform to study the (patho-)physiology of the lacrimal gland.
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Keywords: adult stem cells, lacrimal gland, organoids, regeneration, single-cell sequencing, swelling assay, tear gland, tear secretion, transplantation, Molecular Medicine, Genetics, Cell Biology
ISSN: 1934-5909
Publisher: Cell Press
Note: Funding Information: We thank Stefan van der Elst, Reinier van der Linden, Rosemary Millen, and Cayetano Pleguezuelos-Manzano for flow cytometry assistance; Anko de Graaff and the Hubrecht Imaging Centre (HIC) for microscopy assistance; Single Cell Discoveries for the provided single-cell sequencing service and support; Utrecht Sequencing (USEQ) for bulk RNA-sequencing services; Gijs van Son for help with bulk RNA-sequencing data mapping; and Adriana Martínez-Silgado for help with Illustrator. Y.P. was supported by a grant from the Leducq Foundation ( 14CVD03 ); M.B.-H. was supported by a NWO-ZonW grant ( 114021013 ); H.G., Y.E.B.E., and H.C. were supported by an ERC Advanced Grant (agreement 67013e ); H.G. was supported by a VENI grant from NWO-ZonMW ( 016.166.119 ); and J.v.d.V. was supported by the gravitation program CancerGenomiCs.nl from the Netherlands Organisation for Scientific Research (NWO). Funding Information: We thank Stefan van der Elst, Reinier van der Linden, Rosemary Millen, and Cayetano Pleguezuelos-Manzano for flow cytometry assistance; Anko de Graaff and the Hubrecht Imaging Centre (HIC) for microscopy assistance; Single Cell Discoveries for the provided single-cell sequencing service and support; Utrecht Sequencing (USEQ) for bulk RNA-sequencing services; Gijs van Son for help with bulk RNA-sequencing data mapping; and Adriana Mart?nez-Silgado for help with Illustrator. Y.P. was supported by a grant from the Leducq Foundation (14CVD03); M.B.-H. was supported by a NWO-ZonW grant (114021013); H.G. Y.E.B.E. and H.C. were supported by an ERC Advanced Grant (agreement 67013e); H.G. was supported by a VENI grant from NWO-ZonMW (016.166.119); and J.v.d.V. was supported by the gravitation program CancerGenomiCs.nl from the Netherlands Organisation for Scientific Research (NWO). M.B.-H. Y.P. and H.C. conceived and designed the project. M.B.-H. Y.P. and M.T.B. performed experiments. M.B.-H. performed analysis of RNA-sequencing data with help from H.G. In vivo transplantation was done by J.K. Y.E.B.-E. assisted with fluorescence-activated cell sorting (FACS) for single-cell sequencing. J.v.d.V. helped generate RNA-sequencing data. M.B.-H. H.B. and J.K. generated IHC data. R.K. and S.M.I. provided access to human tissue. M.B.-H. Y.P. and H.C. wrote the manuscript, with input from all authors. H.C. is the inventor on several patents related to organoid technology; his full disclosure is given at https://www.uu.nl/staff/JCClevers/. Y.P. is an employee of Surrozen and was an employee of the Hubrecht Institute when this work was performed and completed. Publisher Copyright: © 2021 Elsevier Inc.
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