Optimized procedures for generating an enhanced, near physiological 2D culture system from porcine intestinal organoids.

Abstract:

:An important practical limitation of the three-dimensional geometry of stem-cell derived intestinal organoids is that it prevents easy access to the apical epithelium for testing food components, microorganisms, bioactive and toxic compounds. To this end, we here report on a new robust method for generating confluent intestinal cell monolayers from single-cell suspensions of enzymatically-dissociated porcine organoids using modified culture conditions. With this method, cell seeding densities can be standardised, overcoming problems with methods based on mechanical dissociation of organoids. Confluent monolayers formed tight junctions with high transepithelial electrical resistance in three days and could be used in experiments for up to two weeks. Multilineage differentiation of ileal stem cells was demonstrated by immunohistochemistry and RT-qPCR of cell-specific transcripts, also unequivocally confirming the controversial existence of Paneth-like cells in the porcine small intestine. The method described here is useful to standardize primary epithelial monolayer formation from intestinal organoids and allows rapid and robust studies of intestinal physiology.

journal_name

Stem Cell Res

journal_title

Stem cell research

authors

van der Hee B,Loonen LMP,Taverne N,Taverne-Thiele JJ,Smidt H,Wells JM

doi

10.1016/j.scr.2018.02.013

subject

Has Abstract

pub_date

2018-04-01 00:00:00

pages

165-171

eissn

1873-5061

issn

1876-7753

pii

S1873-5061(18)30057-6

journal_volume

28

pub_type

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