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Toll-like receptor expression in crypt epithelial cells, putative stem cells and intestinal myofibroblasts isolated from controls and patients with inflammatory bowel disease.

Brown M, Hughes KR, Moossavi S, Robins A, Mahida YR - Clin. Exp. Immunol. (2014)

Bottom Line: In conclusion, enhanced TLR-2 and TLR-4 expression by crypt epithelial cells in active inflammatory bowel disease likely reflects greater ability to respond to microbial products.Results from our studies using mucosal samples from patients with distal ulcerative colitis suggest that the enhanced expression of these TLRs could be constitutive.TLR-2, TLR-4 and TLR-5 expression by stem cells imply ability to respond to distinct bacterial products.

View Article: PubMed Central - PubMed

Affiliation: Institute of Infection, Immunity and Inflammation, University of Nottingham, UK; Nottingham Digestive Diseases Centre, University of Nottingham, UK.

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Expression of BerEP4 (a), Toll-like receptor (TLR)-2 (b), TLR-4 (c) and TLR-5 (d) protein in co-cultures of myofibroblasts and adherent crypt epithelial cells [(e) is negative control]. Isolated and disaggregated colonic crypt epithelial cells were cultured (at 37°C for 30 min) on monolayers of primary human colonic myofibroblasts. After washing, the myofibroblasts and adherent crypt epithelial cells (which are enriched for side population cells) were fixed and used for immunocytochemistry using relevant specific monoclonal antibodies in (a–d), or control buffer (e). Immunolabelled crypt epithelial cells (arrowed) are seen adherent to the much larger underlying myofibroblasts. Myofibroblasts (majority indicated by #) are negative for epithelial cell-specific BerEP4 (hence only their nuclei are seen) and weakly positive for TLR-2, TLR-4 and TLR-5. Each figure is representative of co-cultures using cells isolated from > 5 resection specimens.
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fig07: Expression of BerEP4 (a), Toll-like receptor (TLR)-2 (b), TLR-4 (c) and TLR-5 (d) protein in co-cultures of myofibroblasts and adherent crypt epithelial cells [(e) is negative control]. Isolated and disaggregated colonic crypt epithelial cells were cultured (at 37°C for 30 min) on monolayers of primary human colonic myofibroblasts. After washing, the myofibroblasts and adherent crypt epithelial cells (which are enriched for side population cells) were fixed and used for immunocytochemistry using relevant specific monoclonal antibodies in (a–d), or control buffer (e). Immunolabelled crypt epithelial cells (arrowed) are seen adherent to the much larger underlying myofibroblasts. Myofibroblasts (majority indicated by #) are negative for epithelial cell-specific BerEP4 (hence only their nuclei are seen) and weakly positive for TLR-2, TLR-4 and TLR-5. Each figure is representative of co-cultures using cells isolated from > 5 resection specimens.

Mentions: Side population cells present in isolated and disaggregated crypt cell preparations from normal control colon were characterized by flow cytometry (Fig. 5a), as described previously 24. Sorted side population cells were labelled by anti-BerEP4 (Fig. 5b), anti-TLR-2 and anti-TLR-4 (Fig. 5c) antibodies. When studied by RT–PCR, sorted side population cells also expressed transcripts for TLR-2, TLR-4 and TLR-5 (Fig. 6). In contrast to other disaggregated crypt epithelial cells, side population/putative stem cells adhere readily to monolayers of intestinal myofibroblasts 24. Such co-cultures were used to demonstrate immunoreactivity for not only BerEP4 (Fig. 7a), but also TLR-2 (Fig. 7b), TL-4 (Fig. 7c) and TLR-5 (Fig. 7d). In contrast to the epithelial cells, myofibroblast immunoreactivity for TLR-2, TLR-4 and TLR-5 in these co-cultures was weak.


Toll-like receptor expression in crypt epithelial cells, putative stem cells and intestinal myofibroblasts isolated from controls and patients with inflammatory bowel disease.

Brown M, Hughes KR, Moossavi S, Robins A, Mahida YR - Clin. Exp. Immunol. (2014)

Expression of BerEP4 (a), Toll-like receptor (TLR)-2 (b), TLR-4 (c) and TLR-5 (d) protein in co-cultures of myofibroblasts and adherent crypt epithelial cells [(e) is negative control]. Isolated and disaggregated colonic crypt epithelial cells were cultured (at 37°C for 30 min) on monolayers of primary human colonic myofibroblasts. After washing, the myofibroblasts and adherent crypt epithelial cells (which are enriched for side population cells) were fixed and used for immunocytochemistry using relevant specific monoclonal antibodies in (a–d), or control buffer (e). Immunolabelled crypt epithelial cells (arrowed) are seen adherent to the much larger underlying myofibroblasts. Myofibroblasts (majority indicated by #) are negative for epithelial cell-specific BerEP4 (hence only their nuclei are seen) and weakly positive for TLR-2, TLR-4 and TLR-5. Each figure is representative of co-cultures using cells isolated from > 5 resection specimens.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231243&req=5

fig07: Expression of BerEP4 (a), Toll-like receptor (TLR)-2 (b), TLR-4 (c) and TLR-5 (d) protein in co-cultures of myofibroblasts and adherent crypt epithelial cells [(e) is negative control]. Isolated and disaggregated colonic crypt epithelial cells were cultured (at 37°C for 30 min) on monolayers of primary human colonic myofibroblasts. After washing, the myofibroblasts and adherent crypt epithelial cells (which are enriched for side population cells) were fixed and used for immunocytochemistry using relevant specific monoclonal antibodies in (a–d), or control buffer (e). Immunolabelled crypt epithelial cells (arrowed) are seen adherent to the much larger underlying myofibroblasts. Myofibroblasts (majority indicated by #) are negative for epithelial cell-specific BerEP4 (hence only their nuclei are seen) and weakly positive for TLR-2, TLR-4 and TLR-5. Each figure is representative of co-cultures using cells isolated from > 5 resection specimens.
Mentions: Side population cells present in isolated and disaggregated crypt cell preparations from normal control colon were characterized by flow cytometry (Fig. 5a), as described previously 24. Sorted side population cells were labelled by anti-BerEP4 (Fig. 5b), anti-TLR-2 and anti-TLR-4 (Fig. 5c) antibodies. When studied by RT–PCR, sorted side population cells also expressed transcripts for TLR-2, TLR-4 and TLR-5 (Fig. 6). In contrast to other disaggregated crypt epithelial cells, side population/putative stem cells adhere readily to monolayers of intestinal myofibroblasts 24. Such co-cultures were used to demonstrate immunoreactivity for not only BerEP4 (Fig. 7a), but also TLR-2 (Fig. 7b), TL-4 (Fig. 7c) and TLR-5 (Fig. 7d). In contrast to the epithelial cells, myofibroblast immunoreactivity for TLR-2, TLR-4 and TLR-5 in these co-cultures was weak.

Bottom Line: In conclusion, enhanced TLR-2 and TLR-4 expression by crypt epithelial cells in active inflammatory bowel disease likely reflects greater ability to respond to microbial products.Results from our studies using mucosal samples from patients with distal ulcerative colitis suggest that the enhanced expression of these TLRs could be constitutive.TLR-2, TLR-4 and TLR-5 expression by stem cells imply ability to respond to distinct bacterial products.

View Article: PubMed Central - PubMed

Affiliation: Institute of Infection, Immunity and Inflammation, University of Nottingham, UK; Nottingham Digestive Diseases Centre, University of Nottingham, UK.

Show MeSH
Related in: MedlinePlus