3rd Annual BMRP Investigator Meeting - Abstract
Loop Ileostomy for Crohn’s Disease – Molecular Clues to Disease Pathogenesis
Tariq Ahmad1,a, Karine Maillard2, Maxine Allen2, Sue Goldthorpe1, Marco Daperno1, Fraser Cummings1, Bryan Warren1, Simon Travis1, Ken Welsh3, Sarah Marshall3 and Derek Jewell1,b
1University of Oxford (Oxford, England); 2Oxagen Ltd. (Abingdon, England); 3Department of Immunology, Imperial College (London, England)
Background: Crohn’s disease (CD) is characterized by an inappropriate response of the mucosal immune system to unidentified antigens in the fecal stream. Several genomic regions are believed to harbor susceptibility genes, but positional cloning strategies have been limited by the extent of linkage disequilibrium across these gene dense regions.
Aim: To identify genes involved in susceptibility to colonic CD by combining positional information with changes in gene expression in early disease activation.
Methods: Five patients with a defunctioning ileostomy for refractory colonic CD, who were in clinical remission for at least three months with a CRP<8mg/L, were studied. Patients were challenged on three consecutive days with autologous ileal effluent, instilled down the efferent ileostomy limb. Mucosal biopsies were taken from the ascending colon prior to and on day three of fecal challenge. Four individuals who had undergone loop ileostomy for rectal cancer acted as controls, allowing the discrimination of changes specific to CD. Gene expression was studied using a customized oligonucleotide array (Nimblegen) designed to include genes encoded in extended regions of replicated CD linkage (7,396 probe sets representing ≥ 3500 known genes from chromosomes 1p22–1p36, 3p24-3q12, 5q22–5q35, 6p, 14p13-14q22, 16p13–16q22, 17q12-17q25 and 19p. Hybridizations were carried out in duplicate. Genstat was used to normalize the data and test statistical models. Differential expression of selected genes was confirmed by Q-PCR.
Results: Four of five patients exhibited a rise in CRP, of whom three had clinical and histological evidence of disease activation. No response was seen in controls. Prior to challenge, significant differences in gene expression were observed between CD and controls. Upon challenge, significant changes in expression were seen in both groups. This response was significantly different (P<0.001) between CD and controls for 20 genes, all of which demonstrated reduced expression on challenge in CD, compared to controls.
Discussion: The loop ileostomy provides a unique model to study the complex pattern of gene expression characterizing the initiation and amplification of the aberrant immune response in CD. Combined with positional information, these data may help identify susceptibility genes involved in CD pathogenesis.
aCo-investigator and Presenter; bPrincipal Investigator