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P003. Dendritic cell homing and immune cell function in Crohn's anal fistulae

P. Tozer1,2, H.O. Al-Hassi3, N. Rayment4, D. Bernardo3, A.U. Murugananthan1, N. Daulatzai1,2, T. Ansari5, K. Whelan4, R.K.S. Phillips1,2, S.C. Knight3, A.L. Hart1,2

1St Mark's Hospital, London, United Kingdom; 2Imperial College, London, United Kingdom; 3APRG, Imperial College, London, United Kingdom; 4Kings College, London, United Kingdom; 5NPIMR, London, United Kingdom

Aims: The aetiology of Crohn's disease, and particularly perianal fistulating disease, remains obscure but genetic, microbiological and immunological factors are thought to play a role.

Dendritic cells (DC) are antigen presenting cells which sample mucosal-associated bacteria and migrate to lymph nodes to stimulate an immune response via T cells. DC also express homing markers that they imprint on T cells to direct them to specific organs such as skin (expressing cutaneous lymphocyte-associated antigen, CLA) and gut (expressing α4β7 integrin). In IBD DCs express more Toll-like receptors which recognise microbes and increase production of proinflammatory cytokines.

We aimed to characterise the immune cell composition and cytokine milieu of Crohn's and idiopathic fistulae including assessment of DC phenotype and homing markers.

Methods: Biopsy samples were taken from anal fistula tracts of Crohn's and idiopathic fistula patients. Samples were incubated in medium overnight at 37°C with 5% CO2. ‘Walk out’ cells were isolated from the supernatant and analysed by flow cytometer. DC were identified as HLA-DR positive, lineage (CD3, CD14, CD16, CD19, CD34, CD56) negative and assessed for both subsets of DC; myeloid (CD11c +ve) or plasmacytoid (CD11c −ve), and the homing molecules CLA and α4β7. Proportions and staining intensity on CD14, CD16, CD19, CD3 and CD65 cells were also assessed. TH1/2 and 17 cytokines profile was determined using Multiplex analysis.

Results: Fifteen Crohn's anal fistula patients (CPD) and 12 idiopathic anal fistula patients (IPD) had samples taken for immunological analysis. Rectal biopsies were taken from 10 patients with endoscopically normal mucosa to act as control samples.

 NAgeFemaleDuration of perianal diseaseDuration of luminal diseaseLocation of luminal diseaseStomaImmuno­modulatorsSmoker
IPD1246.533.5 yearsNANA0NA3
CPD153275.5 years12 years4× L1p
5× L2p
4× L3p
2× p
32× oral steroids
5× thiopurines
5
Normal1056.55NANANA0NA2

DC in Crohn's fistula tracts had significantly reduced levels of α4β7 and CLA compared with idiopathic fistulae. There was no significant difference in proportions of either myeloid or plasmacytoid DC, or of CD14, CD16, CD19 or CD3 cells between Crohn's anal fistulae and idiopathic anal fistulae. Crohn's fistula tracts showed lower levels of CD65 than idiopathic fistula tracts (P = 0.04).

Levels of IL-2, IL-4, IL-6, IL-10, TNF and IFN-γ were similar in Crohn's and idiopathic anal fistulae and also the rectum of healthy controls. IL-17a levels were higher in Crohn's anal fistulae than normal rectum and idiopathic anal fistulae (P = 0.04).

Conclusions: Similarities between Crohn's and idiopathic anal fistula tract immune cells and cytokines may suggest that once a fistula tract has formed, the immunological processes in both types of fistula are similar. Differences in levels of IL-17 may provide the basis for strategies of diagnosis and treatment of Crohn's anal fistulae. Aberrant expression of homing molecules on DC in Crohn's disease perianal fistulae suggests a “non-directed” immune response in Crohn's disease which may contribute to the pathophysiology.