Aberrant plasmacytoid dendritic cell distribution and function in patients with Crohn's disease and ulcerative colitis

Abstract

Dendritic cell (DC) function is believed to be of critical importance for the pathogenesis of inflammatory bowel disease (IBD). To date, most research in animal models and the few human data available is restricted to myeloid DC, while plasmacytoid DC (pDC) capable of controlling both innate and adaptive immune responses have not yet been investigated systematically in human Crohn's disease (CD) or ulcerative colitis (UC). CD11c(-) , CD303(+) /CD304(+) and CD123(+) pDC from peripheral blood (n = 90), mucosal tissue (n = 28) or mesenteric lymph nodes (n = 40) (MLNs) of patients with UC and CD or controls were purified and cultured. Thereafter, pDC were enumerated, phenotyped and cytokine secretion measured by flow cytometry (FACS), immunohistochemistry and/or cytometric bead array, respectively. Interferon (IFN)-α secretion following cytosine phosphatidyl guanine (CpG) A oligodeoxynucleotide (ODN) 2216 (5'-GGGGGACGATCGTCGGGGGG-3') stimulation was assessed by enzyme-linked immunosorbent assay (ELISA). We found a significantly higher frequency of pDC in the inflamed colonic mucosa and MLN of IBD patients. Moreover, the fraction of CD40 and CD86 expressing cultured peripheral blood pDC was significantly higher in flaring UC and CD patients and their secretion of tumour necrosis factor (TNF)-α, interleukin (IL)-6 and IL-8 were increased significantly compared with controls. In contrast, the IFN-α secretion of peripheral blood pDC isolated from flaring IBD, particularly in UC patients, was reduced significantly compared with controls. Our data suggest an aberrant distribution and function of pDC in IBD, contrary to their generally implicated role as inducers of tolerance. We speculate that the impaired IFN-α secretion may relate to the hypothesized defect in innate immunity in IBD and could also impact upon the generation of regulatory T cells (T(reg) ).

Fig. 1

Increased frequency of plasmacytoid dendritic cells (pDC) in mesenteric lymph nodes and colonic mucosa of flaring inflammatory bowel disease (IBD) patients. (a) Colonic mucosa: FACS plots from representative experiments. Quadrant thresholds were placed determined according to isotype controls. Bar graphs summarize data from all experiments with control mucosa (n = 16) or tissue from ulcerative colitis (UC) (n = 6) and Crohn's disease (CD) (n = 6) patients. (b) Mesenteric lymph node: FACS plots from representative experiments. Quadrant thresholds were placed determined according to isotype controls. Bar graphs summarize data from all experiments with control lymph node (n = 6) or tissue from UC (n = 14) and CD (n = 20) patients. (c) Mesenteric lymph node: sample immunofluorescence images from control, UC and CD. Asterisks denote statistical significance: **P < 0·01; ***P < 0·001.

Fig. 2

Distinct phenotype of peripheral blood plasmacytoid dendritic cells (pDC) in Crohn's disease (CD) and ulcerative colitis. (a) CD40: increased fraction of CD40 expressing pDC in flaring UC [remission (RM) n = 6, flare-up (FU) n = 3] and CD (RM n = 4, FU n = 3) patients compared with controls (n = 6). (b) CD86: increased fraction of CD86 expressing pDC in flaring UC (RM n = 3, FU n = 3) and CD (RM n = 4, FU n = 4) patients compared with controls (n = 5). (c) FACS plots from representative experiments. Quadrant thresholds were placed determined according to isotype controls. Asterisks denote statistical significance: **P < 0·01; ***P < 0·001.

Fig. 3

Increased secretion of key inflammatory cytokines by peripheral blood plasmacytoid dendritic cells (pDC) from inflammatory bowel disease (IBD) patients. (a) Tumour necrosis factor (TNF)-α: cultured pDC from ulcerative colitis [remission (RM) n = 7, flare-up (FU) n = 3] and Crohn's disease (CD) (RM n = 8, FU n = 10) patients secrete more tumour necrosis factor (TNF)-α than healthy controls (n = 14). (b) Interleukin (IL)-6: cultured pDC from UC (RM n = 7, FU n = 3) and CD (RM n = 8, FU n = 9) patients secrete more IL-6 than healthy controls (n = 13). (c) IL-8: cultured pDC from UC (RM n = 6, FU n = 3) and CD (RM n = 9, FU n = 9) patients secrete more IL-8 than healthy controls (n = 14). Asterisk denotes statistical significance *P < 0·05; **P < 0·01.

Fig. 4

Impaired secretion of interferon (IFN)-α by peripheral blood plasmacytoid dendritic cells (pDC) from inflammatory bowel disease (IBD) patients. Cytosine–phosphatidyl guanine (CpG) A oligodeoxynucleotide (ODN) 2216-stimulated pDC from flaring ulcerative colitis (UC) (RM n = 6, FU n = 3) and CD (RM n = 6, FU n = 6) patients secrete significantly less IFN-α than healthy controls (n = 14). Asterisks denote statistical significance: **P < 0·01.