P075. Epithelial Toll-like receptor 3-dependent synthesis of complement factor B and local complement activation in inflammatory bowel disease
A.E. Østvik1,2,3, A.v.B. Granlund1,3, B.I. Gustafsson1,2, S.H. Torp4,5, T. Espevik1,3, J.K. Damås1,3,6, T.E. Mollnes3,7,8, A.K. Sandvik1,2,3, 1Norwegian University of Science and Technology, Department of Cancer Research and Molecular Medicine, Trondheim, Norway, 2St. Olav's University Hospital, Department of Gastroenterology and Hepatology, Trondheim, Norway, 3Norwegian University of Science and Technology, Centre of Molecular Inflammation Research, Trondheim, Norway, 4Norwegian University of Science and Technology, Department of Laboratory Medicine, Children and Women's Health, Trondheim, Norway, 5St. Olav's University Hospital, Department of Pathology and Medical Genetics, Trondheim, Norway, 6St. Olav's University Hospital, Department of Infectious Diseases, Trondheim, Norway, 7University of Oslo, 8Department of Immunology, Oslo University Hospital Rikshospitalet, and K.G. Jebsen IRC, Oslo, Norway, 8University of Tromsø, Research Laboratory, Nordland Hospital, Bodø, Tromsø, Norway
To verify local synthesis of complement factor B (fB) in a human material of colonic biopsies, after the fB gene CFB was found to be among the most upregulated genes in a gene expression study on an intestinal cell line stimulated with the Toll-like receptor 3 (TLR3) ligand polyinosinic:polycytidylic acid [poly(I:C)]. We also wanted to study if complement activation occurred locally in the intestinal mucosa.
In a microarray study using Illumina Bead Chip technology on poly(I:C) stimulated intestinal cell line we found CFB to be highly upregulated downstream of TLR3. Moreover, microarray gene expression analysis was done on colonic biopsies from an IBD population (n = 133). The expression of CFB was confirmed by PCR. Immunohistochemical staining and in situ hybridization of colonic biopsies were done to identify cellular sources of fB/CFB. Also, immunohistochemical staining for terminal complement complex (TCC) in colonic biopsies was done to assess complement activation.
Complement factor B mRNA and protein were abundantly expressed in the colonic epithelial cell line HT-29, and synthesis was enhanced by the TLR3 ligand (poly(I:C). CFB mRNA was the most significantly overexpressed gene and the diseased/non-diseased mRNA abundance ratio among the 50 highest in inflamed vs normal colon, of inflamed vs un-inflamed mucosa of UC and CD. Both the epithelial cells and infiltrating neutrophils/granulocytes of the inflamed mucosa expressed fB. Immunohistochemical staining showed positivity for TCC in submucosal vessels; local complement activation does take place in active IBD.
Complement factor B is expressed in colonic epithelial cells and regulated by the TLR3 agonist poly(I:C). It is the most significantly upregulated gene in a large human material of colonic biopsies in IBD, and together with local complement activation in colonic mucosa this strongly suggests a role for complement in IBD pathogenesis.