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3D Cell Culture Models of Epithelial Tissues

  • Kai ZhangEmail author
  • Aki ManninenEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1926)

Abstract

Cells in tissues in vivo face a very different microenvironment than typical cultured cells plated on a plastic dish. Already several decades ago, cell biologists observed that cell lines show dramatically different morphology and growth characteristics when embedded into three-dimensional (3D) substrates or standard tissue culture plates (Montesano R, Schaller G, Orci L, Cell. 66:697–711, 1991; Barcellos-Hoff MH, Aggeler J, Ram TG, Bissell MJ, Development. 105:223–235, 1989; Simian M, Bissell MJ, J Cell Biol. 216:31–40, 2017). Despite its imminent benefit for cell biological studies, suspicion and prejudice toward more complicated sample preparation requirements limited the popularity of 3D culture techniques until recently, when it was shown that soft 3D gels made of basement membrane extracts (BME) allow prolonged culture of many types of primary epithelial cells (Clevers H, Cell. 165:1586–1597, 2016; Sato T, Vries RG, Snippert HJ, van de Wetering M, Barker N, Stange DE, et al., Nature. 459:262–265, 2009). These observations have brought 3D organoid culture systems into the mainstream. Here we describe two protocols for culturing epithelial cells in 3D substrates, the “blob culture” setup where cells are fully embedded into BME gel and the “overlay setup” where cells are seeded on top of BME gel and then overlaid with a thin layer of BME (Debnath J, Brugge JS, Nat Rev Cancer. 5:675–688, 2005; Bryant DM, Datta A, Rodriguez-Fraticelli AE, Peranen J, Martin-Belmonte F, Mostov KE, Nat Cell Biol. 12:1035–1045, 2010).

Key words

MDCK 3D culture Organoid 

Notes

Acknowledgments

Riitta Jokela is acknowledged for overall expert technical assistance, Jaana Träskelin for expert technical assistance at Biocenter Oulu Virus Core Laboratory, and Veli-Pekka Ronkainen for expert assistance in microscopy at Biocenter Oulu Tissue Imaging Center. This work was funded by the Academy of Finland (251314, 135560, 263770, and 140974/AM).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Center for Cellular Immunotherapies, Department of Pathology and Laboratory Medicine, Perelman School of MedicineUniversity of PennsylvaniaPhiladelphiaUSA
  2. 2.Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, Oulu Center for Cell-Matrix ResearchUniversity of OuluOuluFinland

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