We and others have recently shown that the time of onset, incidence and severity of colitis is exacerbated in IL-10-/- mice that also bear a mutation in TLR-4. By a mechanism apparently unrelated to its role in innate immune signaling by antigen presenting cells (APC), TLR-4 signaling by CD4+ T cells regulates the response to subsequent TCR activation. CD4+ T cells from TLR-4-/- x IL-10-/- (DKO) mice exhibit enhanced proinflammatory colitiogenic effector responses when measured either directly ex vivo or after adoptive transfer into Rag1-/- recipients. Moreover, Foxp3+ Tregs isolated from TLR-4-/- mice are impaired in their ability to secrete IL-10. TLR-4 signaling in CD4+ T cells therefore enhances Treg function and inhibits inflammatory T effector function;these two roles are likely to synergize to prevent intestinal inflammation in healthy mice. In DKO mice, however, Foxp3+ Tregs accumulate in the inflamed lamina propria (LP), secrete IFN-? and IL-17, and fail to control disease. A role for MyD88 dependent TLR-4 mediated signals in the homeostatic maintenance of the intestinal epithelial barrier is already well established. We found that apoptotic remnants apparently derived from the intestinal epithelium accumulate in the colonic LP of Helicobacter-positive (Hh+) TLR-4-/- mice (in the absence of inflammation) as well as in the inflamed LP of both IL-10-/- and DKO mice. Apoptotic cells were not detectable in the colonic LP of WT C57Bl/6 mice or Helicobacter free (Hh-) TLR-4-/- or DKO mice. How does the absence of TLR-4 mediated signals exacerbate intestinal inflammation in IL-10 deficient mice? Our working hypothesis is that apoptotic epithelial cells accumulate in the colonic LP of Helicobacter-colonized DKO mice and induce an inflammatory Th17 response. IFN-? is also induced in the absence of tonic TLR-4 signaling in CD4+ effector T cells. The apoptosis induced IL-17 response leads to an earlier and increased recruitment of inflammatory cells to the intestines that results ultimately in chronic colitis. In Helicobacter-colonized TLR-4-/- mice this chronic inflammatory response is held in check by the immunoregulatory effects of IL-10. In DKO mice, however, T cell activation is dysregulated in the absence of TLR-4 signaling, potentiating the inflammatory T effector response. Treg function and/or stability is also compromised in DKO mice and the Tregs themselves become pathogenic effector cells.
In Aim 1, both in vitro and in vivo approaches will be used to examine whether Helicobacter induced apoptotic epithelial remnants phagocytosed by APC in the colonic LP induce IL-17 producing Foxp3- and Foxp3+ T cells and drive intestinal inflammation in DKO mice.
In Aim 2 we will further characterize the putative pathogenic Tregs in the LP of DKO mice and the triggers that destabilize or convert this Treg subset. We will also examine the functional suppressive and/or effector capabilities of these cells.

Public Health Relevance

We will use a novel murine model of spontaneous intestinal inflammation to examine the triggers that lead to the failure of the responses that protect healthy individuals from disease. A clearer understanding of the mechanisms regulating this protection may inform the development of new immunotherapeutic approaches to treat or prevent inflammatory bowel disease (IBD).

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21DK089999-02
Application #
8102070
Study Section
Special Emphasis Panel (ZRG1-DKUS-C (04))
Program Officer
Hamilton, Frank A
Project Start
2010-07-01
Project End
2012-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
2
Fiscal Year
2011
Total Cost
$193,050
Indirect Cost
Name
University of Chicago
Department
Pathology
Type
Schools of Medicine
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
Evans-Marin, Heather L; Cong, Yingzi (2014) Microbiota regulation of inflammatory bowel disease. Inflamm Allergy Drug Targets 13:65-73
Cao, Anthony T; Yao, Suxia; Stefka, Andrew T et al. (2014) TLR4 regulates IFN-γ and IL-17 production by both thymic and induced Foxp3+ Tregs during intestinal inflammation. J Leukoc Biol 96:895-905