Rapid accumulation of CD14+CD11c+ dendritic cells in gut mucosa of celiac disease after in vivo gluten challenge

Ann-Christin Røberg Beitnes, Melinda Ráki, Margit Brottveit, Knut Erik Aslaksen Lundin, Frode Lars Jahnsen, Ludvig Magne Sollid, Ann-Christin Røberg Beitnes, Melinda Ráki, Margit Brottveit, Knut Erik Aslaksen Lundin, Frode Lars Jahnsen, Ludvig Magne Sollid

Abstract

Background: Of antigen-presenting cells (APCs) expressing HLA-DQ molecules in the celiac disease (CD) lesion, CD11c(+) dendritic cells (DCs) co-expressing the monocyte marker CD14 are increased, whereas other DC subsets (CD1c(+) or CD103(+)) and CD163(+)CD11c(-) macrophages are all decreased. It is unclear whether these changes result from chronic inflammation or whether they represent early events in the gluten response. We have addressed this in a model of in vivo gluten challenge.

Methods: Treated HLA-DQ2(+) CD patients (n = 12) and HLA-DQ2(+) gluten-sensitive control subjects (n = 12) on a gluten-free diet (GFD) were orally challenged with gluten for three days. Duodenal biopsies obtained before and after gluten challenge were subjected to immunohistochemistry. Single cell digests of duodenal biopsies from healthy controls (n = 4), treated CD (n = 3) and untreated CD (n = 3) patients were analyzed by flow cytometry.

Results: In treated CD patients, the gluten challenge increased the density of CD14(+)CD11c(+) DCs, whereas the density of CD103(+)CD11c(+) DCs and CD163(+)CD11c(-) macrophages decreased, and the density of CD1c(+)CD11c(+) DCs remained unchanged. Most CD14(+)CD11c(+) DCs co-expressed CCR2. The density of neutrophils also increased in the challenged mucosa, but in most patients no architectural changes or increase of CD3(+) intraepithelial lymphocytes (IELs) were found. In control tissue no significant changes were observed.

Conclusions: Rapid accumulation of CD14(+)CD11c(+) DCs is specific to CD and precedes changes in mucosal architecture, indicating that this DC subset may be directly involved in the immunopathology of the disease. The expression of CCR2 and CD14 on the accumulating CD11c(+) DCs indicates that these cells are newly recruited monocytes.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. CD14 + CD11c + dendritic…
Figure 1. CD14+CD11c+ dendritic cells are selectively increased in duodenal mucosa of celiac disease patients after short-term gluten challenge.
Density of HLA-DQ+ antigen-presenting cell subsets in cryosections from celiac disease (CD) patients on gluten-free diet (GFD) before and after a three-day gluten challenge. The density of CD163+CD11c− macrophages is calculated by subtracting the number of CD163+CD11c+ cells from the total number of CD163+HLA-DQ+ cells. Paired data are connected by lines. ns = not significant (A). Three-color immunofluorescence staining for CD11c (green), CD14 (red) and cytokeratin (blue) in cryosection of duodenal mucosa from a CD patient on GFD before and after a three-day gluten challenge. Original magnification X 400 (B).
Figure 2. Expression of CCR2 on HLA-DR…
Figure 2. Expression of CCR2 on HLA-DR+ leukocytes in blood and duodenal mucosa.
Flow cytometric analysis of peripheral blood mononuclear cells from treated celiac disease (CD) patient (A) and viable single cells of duodenal mucosa from treated CD patient showing the expression of CCR2 (B) and DC-SIGN (C) on CD45+HLA-DR+ cells depending on the expression of CD11c and CD14. Dead cells were excluded by adding 0.2 µg/mL propidium iodide immediately before acquisition. The data are representative for two independent experiments.
Figure 3. CCR2 is highly expressed on…
Figure 3. CCR2 is highly expressed on CD14+CD11c+ dendritic cells in duodenal mucosa.
Percentage of CCR2+ cells in subsets of antigen-presenting cells from duodenal mucosa digests assessed by flow cytometry. Median is indicated by horizontal line. CD = celiac disease.
Figure 4. Neuthrophils are increased in duodenal…
Figure 4. Neuthrophils are increased in duodenal mucosa of celiac disease patients after short-term gluten challenge.
Density of CD3+ intraepithelial lymphocytes (IELs) per 100 epithelial cells (A); neutrophils per mm2 (B); and eosinophils per mm2 (C) in the lamina propria (LP) in sections of duodenal mucosa from celiac disease patients on gluten-free diet before and after a three-day gluten challenge. Paired data are connected by lines. ns = not significant.
Figure 5. Peripheral blood CD14 + monocytes…
Figure 5. Peripheral blood CD14+ monocytes efficiently present gluten to gluten-specific T cell clones.
Purity of CD14+CD11c+ monocytes isolated from peripheral blood mononuclear cells of two individuals are shown (A and B, upper panels). The monocytes were incubated with medium or two different gluten peptides ±100 U/ml IFN-γ for 24 hours, washed and incubated with a T-cell clone for 72 hours. The proliferative T-cell response (measured by thymindine-incorporation) is shown (A and B, lower panels). Experiments with IFN-γ are indicated (hatched columns).
Figure 6. Density of various leukocyte subsets…
Figure 6. Density of various leukocyte subsets remains unchanged in duodenal mucosa of gluten-sensitive control subjects after short-term gluten challenge.
Density of CD14+CD11c+ dendritic cells (DCs) per mm2 (A) and CD103+CD11c+ DCs per mm2 (B) in the lamina propria (LP); CD3+ intraepithelial lymphocytes (IELs) per 100 epithelial cells (C); neutrophils per mm2 (D) and eosinophils per mm2 (E) in the LP in sections of duodenal mucosa from gluten-sensitive control subjects on gluten-free diet before and after a three-day gluten challenge. Paired data are connected by lines. ns = not significant.

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