3rd Annual BMRP Investigator Meeting - Abstract
Cdcs1, a Major Colitis Susceptibility Gene in Mice, Regulates Innate and Adaptive Immune Response to Enteric Bacterial Antigens
Edward H. Leiter1,a, John P. Sundberg1, Jason Beckwith1, Yingzi Cong2 and Charles O. Elson2
1The Jackson Laboratory (Bar Harbor, Maine, U.S.A.); 2Division of Gastroenterology, University of Alabama (Birmingham, Alabama, U.S.A.)
Inflammatory bowel disease (IBD) in humans entails interaction between a genetically susceptible genotype and the enteric flora. We have modeled this interaction in mice to understand IBD etiopathogenesis. Mice of the C3H/HeJBir (C3Bir) substrain were originally selected for spontaneous development of juvenile enterocolitis. Changes in animal husbandry at The Jackson Laboratory have currently suppressed spontaneous colitis development. However, the greater C3Bir susceptibility is readily apparent by comparing the effect of IL-10 deficiency on this background versus on the C57BL/6J (B6) background. Absence of IL-10, a key cytokine in gut homeostasis, causes severe colitis in C3Bir, but only mild colitis in B6 mice. Genetic segregation analysis showed a complex genetic control with at least six quantitative trait loci discovered. Among these, the major C3Bir-derived colitogenic modifier, designated Cytokine deficiency induced colitis susceptibility-1 (Cdcs1), was mapped on Chromosome 3. This locus contributed a sufficient proportion of the variance in both cecum and colon histopathology such that it could be analyzed as a Mendelian trait in reciprocal congenic stocks.
C3Bir mice congenic for the B6-derived Cdcs1 allele and reciprocal B6 mice congenic for the C3Bir allele were analyzed histopathologically for colitis development. Analysis of interval-specific recombinants allowed positioning of Cdcs1 within a minimum seven megabase interval around the Nfkb1 gene. Functional differences were found in innate and adaptive immune responses of parental and congenic stocks following activation with Toll-like receptor (TLR) 5 and nine bacterial ligands in vitro (cytokine release from bone marrow-derived macrophages and dendritic cells) and in vivo (serum cytokines and primed CD4+ T cell proliferation). C3Bir mice showed impaired innate responses, but higher adaptive T cell responses in comparison to B6. Comparison of the reciprocal congenic stocks showed these immunophenotypes were regulated by the Cdcs1 allele. Importantly, C3Bir colitis was significantly diminished by B6 genome in this interval. Reciprocally, colitis in B6 was significantly exacerbated the C3Bir Cdcs1 allele. Parental strains were compared by electrophoretic mobility shift assay to assess the candidacy of Nfkb1 (NF-κB p50) in the Cdcs1 interval. C3Bir macrophages constitutively expressed higher nuclear NF-κB p50 content in what appeared to be a homodimer. This phenotype in cultured macrophages from male congenic donors was responsive to the respective allele at Cdcs1.
We conclude that the C3Bir allele at Cdcs1 imparts a reduced innate responsiveness both in vivo and in vitro to bacterial ligands that, in turn, produces a compensatory increase in CD4 T cell responses to the enteric flora. These findings, although unexpected, are consistent with what is being learned about mutations in the human NOD2 gene and increased susceptibility to Crohn’s disease. Because the Cdcs1 defect is expressed in cultured C3Bir bone marrow-derived macrophages and dendritic cells, we propose gene expression studies to understand the specific Cdcs1-mediated changes in C3Bir signaling pathways that lead to impaired innate responses to the enteric flora and hence, increased IBD susceptibility.
aPrincipal Investigator