Abstract
Background & Aims The intestinal epithelium is the first line of defense against enteric pathogens. We investigated the response of small intestinal and colonic crypt cultures to a panel of toll-like receptor ligands to assess the impact of microbial pattern recognition on epithelial growth. Methods Primary murine jejunal enteroids and colonoids were cultured with lipopeptide Pam3CSK4, lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (Poly I:C) for 4 to 6 days. Surface area, budding and survival were assessed. Proliferation and numbers of lysozyme positive cells were quantified by flow cytometry. Gene expression was assessed by Nanostring and qRT-PCR. Results Exposure to Pam3CSK4 and LPS hadminimal impact on either enteroids or colonoids. In contrast, Poly I:C increased the surface area of enteroids, while colonoids demonstrated decreased budding. Survival was decreased by Poly I:C in enteroids but not in colonoids. Both enteroids and colonoids exhibited upregulated gene expression of chemokines, but these were increased in magnitude in enteroids. Decreases in gene expression associated with epithelial differentiation and lysozyme positive cells weremore apparent in enteroids than in colonoids. Baseline gene expression between enteroids and colonoids differed markedly in levels of stem cell and inflammatorymarkers. The changes inmorphology induced by Poly I:C were mediated by the toll-like receptor adaptor molecule 1 (Ticam1) in enteroids but not in colonoids. Conclusions Poly I:C alters the molecular program of epithelial cells and shifts from absorption and digestion towards defense and inflammation. Diversity of responses to microbial patterns in enteroids and colonoids may underlie differences in susceptibility to infection along the intestinal tract.
| Original language | English (US) |
|---|---|
| Article number | e0138531 |
| Journal | PLoS One |
| Volume | 10 |
| Issue number | 9 |
| DOIs | |
| State | Published - Sep 28 2015 |
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ASJC Scopus subject areas
- Agricultural and Biological Sciences(all)
- Biochemistry, Genetics and Molecular Biology(all)
- Medicine(all)
Cite this
The viral mimetic polyinosinic : Polycytidylic acid alters the growth characteristics of small intestinal and colonic crypt cultures. / Davies, Julie M.; Santaolalla, Rebeca; Von Furstenberg, Richard J.; Henning, Susan J.; Abreu, Maria T.
In: PLoS One, Vol. 10, No. 9, e0138531, 28.09.2015.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - The viral mimetic polyinosinic
T2 - Polycytidylic acid alters the growth characteristics of small intestinal and colonic crypt cultures
AU - Davies, Julie M.
AU - Santaolalla, Rebeca
AU - Von Furstenberg, Richard J.
AU - Henning, Susan J.
AU - Abreu, Maria T
PY - 2015/9/28
Y1 - 2015/9/28
N2 - Background & Aims The intestinal epithelium is the first line of defense against enteric pathogens. We investigated the response of small intestinal and colonic crypt cultures to a panel of toll-like receptor ligands to assess the impact of microbial pattern recognition on epithelial growth. Methods Primary murine jejunal enteroids and colonoids were cultured with lipopeptide Pam3CSK4, lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (Poly I:C) for 4 to 6 days. Surface area, budding and survival were assessed. Proliferation and numbers of lysozyme positive cells were quantified by flow cytometry. Gene expression was assessed by Nanostring and qRT-PCR. Results Exposure to Pam3CSK4 and LPS hadminimal impact on either enteroids or colonoids. In contrast, Poly I:C increased the surface area of enteroids, while colonoids demonstrated decreased budding. Survival was decreased by Poly I:C in enteroids but not in colonoids. Both enteroids and colonoids exhibited upregulated gene expression of chemokines, but these were increased in magnitude in enteroids. Decreases in gene expression associated with epithelial differentiation and lysozyme positive cells weremore apparent in enteroids than in colonoids. Baseline gene expression between enteroids and colonoids differed markedly in levels of stem cell and inflammatorymarkers. The changes inmorphology induced by Poly I:C were mediated by the toll-like receptor adaptor molecule 1 (Ticam1) in enteroids but not in colonoids. Conclusions Poly I:C alters the molecular program of epithelial cells and shifts from absorption and digestion towards defense and inflammation. Diversity of responses to microbial patterns in enteroids and colonoids may underlie differences in susceptibility to infection along the intestinal tract.
AB - Background & Aims The intestinal epithelium is the first line of defense against enteric pathogens. We investigated the response of small intestinal and colonic crypt cultures to a panel of toll-like receptor ligands to assess the impact of microbial pattern recognition on epithelial growth. Methods Primary murine jejunal enteroids and colonoids were cultured with lipopeptide Pam3CSK4, lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (Poly I:C) for 4 to 6 days. Surface area, budding and survival were assessed. Proliferation and numbers of lysozyme positive cells were quantified by flow cytometry. Gene expression was assessed by Nanostring and qRT-PCR. Results Exposure to Pam3CSK4 and LPS hadminimal impact on either enteroids or colonoids. In contrast, Poly I:C increased the surface area of enteroids, while colonoids demonstrated decreased budding. Survival was decreased by Poly I:C in enteroids but not in colonoids. Both enteroids and colonoids exhibited upregulated gene expression of chemokines, but these were increased in magnitude in enteroids. Decreases in gene expression associated with epithelial differentiation and lysozyme positive cells weremore apparent in enteroids than in colonoids. Baseline gene expression between enteroids and colonoids differed markedly in levels of stem cell and inflammatorymarkers. The changes inmorphology induced by Poly I:C were mediated by the toll-like receptor adaptor molecule 1 (Ticam1) in enteroids but not in colonoids. Conclusions Poly I:C alters the molecular program of epithelial cells and shifts from absorption and digestion towards defense and inflammation. Diversity of responses to microbial patterns in enteroids and colonoids may underlie differences in susceptibility to infection along the intestinal tract.
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U2 - 10.1371/journal.pone.0138531
DO - 10.1371/journal.pone.0138531
M3 - Article
C2 - 26414184
AN - SCOPUS:84946924422
VL - 10
JO - PLoS One
JF - PLoS One
SN - 1932-6203
IS - 9
M1 - e0138531
ER -