Scientific Abstract

Proposal No.  NICE-03
Principal Investigator:  R. Balfour  Sartor, M.D.
Applicant Organization:  University of North Carolina at Chapel Hill (U.S.A.)
Project Title:  Identification of dominant bacterial antigens from IL-10 knockout mice and HLA-527 transgenic rats with colitis
Period of Award:  January 1, 2003 - December 31, 2004

Specific Aims

 1.    Collect cecal samples from specific pathogen-free (SPF) IL-10 knockout (ko) mice on two genetic backgrounds (129 SvEv and C57/Bl6) and HLA B27 transgenic rats with colitis.  These samples and aliquots of Bacteroides vugatus (which cause colitis in monoassociated HLA B27 TG rats) and Enterococcus faecalis (which cause disease in monoassociated IL-10 ko mice) will be sent to Corixa for expression cloning and sequencing.

 2.    Screen these libraries by responses of MLN CD4+ T cells from gnotobiotic (monoassociated with the bacterial species of interest) and SPF mice and rats with colitis cultured with  bacterial antigen-pulsed antigen presenting cells (APC) to isolate the dominant antigens.  Wildtype mice and rats will be used for specificity controls.

 3.    Screen these libraries by responses  of dendritic cells and macrophages from transgenic mice expressing GFP on an inducible NFκB promoter (using GFP as a readout) and rat and human epithelial cell lines (using chemokine secretion as a read out).

 4.    Express the dominant antigens in bacterial vectors that do not induce inflammation in these models, and delete the epitopes  from enteric bacterial species normally expressing these antigens.  Determine the ability of  these recombinant bacteria selectively populating (monoassociate) germ free  IL-10 ko and HLA B27 TG rats to induce colitis.

 5.    Collect serum from inflammatory bowel disease and control patients and measure serologic responses to selected recombinant bacterial antigens.