Chlamydia trachomatis infections are the leading cause of involuntary infertility and ectopic pregnancy in women. Although antibiotic treatment cures infection, it does not ameliorate the inflammatory process that leads to the development of disease. We have previously demonstrated that Toll-like receptor 2 (TLR2)-deficient mice fail to develop oviduct pathology after chlamydial infection, demonstrating that TLR2 signaling is directly involved in disease development. Identification of the bacterial product responsible for induction of TLR2-dependent signaling in response to chlamydial infection is essential to advance our ultimate goal of determining the molecular mechanisms that drive the development of chlamydia-induced immunopathology. We have derived novel, plasmid-deficient C. muridarum mutants that retain the ability to infect the murine genital tract but do not develop the characteristic upper tract scarring and pathology of chlamydial disease because they fail to stimulate TLR2-dependent signaling in cell culture and in vivo. We hypothesize that a gene product or regulatory element transcribed from the chlamydial plasmid influences expression or modification of pathogenic TLR2 ligand(s). In this application we propose careful, systematic proteomic and biochemical approaches to identify and characterize the chlamydial component that drives the development of upper tract disease. Specifically we will: (1) employ two-dimensional difference gel electrophoresis (2D-DIGE) to resolve differences in the expression profile of the plasmid-deficient strain when compared to its parent, and the use of a TLR2-Fc fusion construct as a way to identify candidate TLR2 ligands and to characterize ligand-receptor complexes biochemically;(2) examine the immunomodulatory properties of chlamydial glycogen which is no longer synthesized by plasmid-deficient chlamydiae and (3) determine the contribution of chlamydial lipoproteins to the induction of TLR2-dependent cytokine expression. Plasmid-deficient chlamydiae express Mip, a recently-identified candidate chlamydial TLR2 ligand, normally, suggesting that native chlamydial lipoproteins do not contribute to the development of reproductive tract disease. Identification of the pathogenic TLR2 ligand(s) will serve several purposes. We will be able to evaluate expression of the TLR2-dependent ligands by the plasmid-deficient strains. The genes or pathways involved in the expression of the ligand may serve as drug targets for therapies that limit tissue damage arising from infection. It may aid in the identification of endogenous TLR2 activating ligands that might contribute to the development of tubal scarring. Finally, it is possible that the TLR2 ligand of C. trachomatis could serve as an immunogen, eliciting protective responses that block or down-regulate TLR2-dependent signaling in response to chlamydial infection and provide a novel approach to protecting the upper reproductive tract from deleterious sequelae.

Public Health Relevance

Chlamydia trachomatis infections are the leading cause of involuntary infertility and ectopic pregnancy in women. Tissue damage and scarring of the upper reproductive tract that occurs is the result of an immune response triggered by an as yet unknown molecule, or ligand, made by the bacteria. This application is to use novel mutants of C. trachomatis that cannot stimulate this signaling process as tools to identify this ligand and to examine the stimulatory properties of a carbohydrate polymer and a lipoprotein made by chlamydiae to determine if they also contribute to this damaging signal pathway.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AI083657-02
Application #
7895053
Study Section
Special Emphasis Panel (ZRG1-IDM-A (90))
Program Officer
Hiltke, Thomas J
Project Start
2009-07-18
Project End
2012-06-30
Budget Start
2010-07-01
Budget End
2012-06-30
Support Year
2
Fiscal Year
2010
Total Cost
$227,250
Indirect Cost
Name
University of Pittsburgh
Department
Pediatrics
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213