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Veterinary research2021; 52(1); 108; doi: 10.1186/s13567-021-00976-0

Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics.

Abstract: Enteroid cultures are three-dimensional in vitro models that reflect the cellular composition and architecture of the small intestine. One limitation with the enteroid conformation is the enclosed lumen, making it difficult to expose the apical surface of the epithelium to experimental treatments. The present study was therefore conducted to generate cultures of equine enteroids and to develop methods for culture of enteroid-derived cells on a two-dimensional plane, enabling easy access to the apical surface of the epithelium. Equine enteroids were established from small intestinal crypts within 7-9 days of culture. Transcriptional analysis of cell type markers confirmed the presence of enterocytes, stem-, Paneth-, proliferative-, enteroendocrine-, goblet- and tuft cells. This cellular composition was maintained over multiple passages, showing that the enteroids can be kept for prolonged periods. The transfer from 3D enteroids to 2D monolayers slightly modified the relative expression levels of the cell type markers, indicating a decrease of goblet- and Paneth cells in the monolayers. Stimulation with the TLR2, 3 and 4 agonists Pam3CSK4, Poly I:C and LPS, respectively, induced the pro-inflammatory cytokines TNF-α and IL-8, while the TLR5 agonist FliC only induced TNF-α. In addition, an up-regulation of TGF-β, IL-33 and IFN-β was recorded after exposure to lipofected Poly I:C that also affected the monolayer integrity. Thus, the equine enteroid-derived 2D monolayers described in the present study show both genetic and functional similarities with the equine intestine making it an interesting in vitro model for studies demanding access to the apical surface, e.g. in studies of host-microbe interactions.
© 2021. The Author(s).
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Publication Date: 2021-08-14 PubMed ID: 34391473PubMed Central: PMC8364015DOI: 10.1186/s13567-021-00976-0Google Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research article discusses the creation of 3D structures called enteroids, particularly equine enteroids, from the horses’ small intestine cells, and their conversion into 2D monolayers. These 2D monolayers are useful in research as they allow easy access to the epithelial layer of the intestine for experiments.

Equine Enteroids and 2D Monolayers

  • The researchers were able to successfully make and grow equine enteroids from small intestinal crypts, tiny invaginations of the intestine’s lining, within 7 to 9 days. These enteroids are 3D cultures that mimic the structure and cellular makeup of a horse’s small intestine. They contain enterocytes (main cells of the intestinal lining), stem cells, proliferative cells, and several specialized cell types such as Paneth, goblet, and tuft cells.
  • To overcome the difficulty of accessing the inner surface (apical surface) of these 3D enteroids, the researchers converted them into 2D monolayers. These are simpler structures that provide easy access to the apical surface for experimental treatments.
  • When they transferred enteroids to 2D monolayers, the researchers observed some changes in the relative levels of cellular composition, particularly a decrease in numbers of goblet cells and Paneth cells. These cells are involved in intestinal immunity and secretion.

Response to Microbial Mimics

  • The researchers tested the responsiveness of these 2D monolayers to microbial mimics or agonists. These are substances that can activate receptors on the cells to trigger an immune response, mimicking the body’s response to actual pathogens.
  • When exposed to the agonists Pam3CSK4, Poly I:C, and LPS (which activate TLR2, TLR3, and TLR4 receptors), the monolayers successfully produced inflammation-related substances TNF-alpha and IL-8.
  • The TLR5 agonist, FliC, was capable of inducing the production of TNF-alpha only.
  • When lipofected Poly I:C (a form of Poly I:C that can enter the cells more effectively) was used, it not only affected the structure of the 2D layer but also led to the up-regulated production of TGF-beta, IL-33, and IFN-beta, further demonstrating the 2D layers’ potential for immune response studies.

Relevance of the Study

  • The successful production of equine enteroid-derived 2D monolayers that respond to microbial mimics highlights their potential as in vitro models for further studies, particularly those that require access to the apical surface.
  • The 2D monolayers can be utilized for intricate studies of host-microbe interactions, providing insights about the intestinal immune response, the effects of diet or drugs on intestinal health, or even the pathogenesis of intestinal disorders.

Cite This Article

APA
Hellman S. (2021). Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics. Vet Res, 52(1), 108. https://doi.org/10.1186/s13567-021-00976-0

Publication

Veterinary research
ISSN: 1297-9716
NlmUniqueID: 9309551
Country: England
Language: English
Volume: 52
Issue: 1
Pages: 108
PII: 108

Researcher Affiliations

Hellman, Stina
  • Department of Biomedical Sciences and Veterinary Public Health, Swedish University of Agricultural Sciences, SLU, P.O. Box 7028, 750 07, Uppsala, Sweden. Stina.Hellman@slu.se.

MeSH Terms

  • Animals
  • Cell Culture Techniques / methods
  • Cell Culture Techniques / veterinary
  • Female
  • Horses
  • Intestine, Small / cytology
  • Organoids / cytology

Grant Funding

  • 2019-00809 / Svenska Forskningsru00e5det Formas
  • H-16-47-193 / Swedish-Norwegian Foundation for Equine Research
  • SLU ua 2019.4.2 - 3814 / Sveriges Lantbruksuniversitet (SE) SLU Future animals, nature and health

Conflict of Interest Statement

The author declare that they have no competing interests.

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Citations

This article has been cited 14 times.
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