Genital infection by Chlamydia trachomatis is the most common bacterial sexually transmitted disease (STD) in the United States with an excess of $2 billion spent on over four million annual clinical cases. Like several other intracellular bacterial pathogens that require a robust T helper type 1 (Th1) immunity for control (e.g., Listeria and Mycobacteria), there are no vaccines against C. trachomatis. Progress in the immunobiology of Chlamydia has indicated that novel approaches to identify and target immunomodulatory factors that regulate the induction of Th1 cells are crucial for designing effective vaccines against these pathogens. Recently, it was found that chlamydia-pulsed, interleukin (IL)-10 deficient dendritic cells (DCs) were potent antigen-presenting cells that induced a rapid and robust Th1 response and the complementary humoral immune response which conferred sterilizing immunity against genital chlamydial infection in mice. The effectiveness of chlamydiapulsed IL-10 deficient DCs is not due merely to the absence of IL-10 but to acquisition of certain immunobiologic properties that include rapid maturation and expression of a unique set of immunomodulatory molecules. The main objective of this study is to elucidate the molecular and immunobiological basis for the potency of chlamydia-pulsed IL-10 deficient DCs, including defining novel molecular elements that can be applied in designing and delivering efficacious vaccines against Chlamydia. The central hypothesis to be tested is that chlamydia-primed IL-10 deficient DCs are quantitatively and qualitatively distinct in their metabolic characteristics relating to T cell activation compared to wild-type (WT) DCs. To investigate this hypothesis, we will use a combination of proteomics and immunological techniques, including twodimensional gel electrophoresis (2-DE), matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF), in vivo gene silencing via short interfering RNA (siRNA), and analysis of genetically-engineered specific gene knockout or transgenic mice, to identify and immunologically characterize certain gene products that contribute to early DC maturation and promote efficient APCs function for an enhanced Th1 activation. Additional in vivo and ex vivo biochemical techniques will be used to deliver chlamydia-specific vaccine constructs in the presence or absence of the relevant molecules identified, to determine the effect on vaccine efficacy in vivo, in a murine model of genital Chlamydia infection. The ultimate goal is to identify and characterize certain immunomodulatory molecules that can be used to design and deliver efficacious vaccines against Chlamydia.
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