The intracellular bacterium, Chlamydia (C.) pneumoniae, has emerged over the last decade as a major human respiratory pathogen. C. pneumoniae infection also has been associated with atheromatous plaques in coronary heart disease. Much of the tissue injury in chlamydial disease presumably results from a deleterious immune response to the chlamydial protein hsp60 of the 60 kDa family of heat shock proteins. We propose to define the pathogenetic role of chlamydial hsp60 in a mouse model of C. pneumoniae pneumonitis. We will generate mice immunotolerant to hsp60 with two novel approaches, and expect these animals to be, in part, resistant to immunopathology induced by infection with whole organisms. We also will prime mice to T-Helper-cell 1 (TH1)- or TH2- dominated immunity against hsp60 by DNA-vaccination with plasmid vectors simultaneously expressing chlamydial hsp60 and murine interleukin-12 (IL- 12) or IL- 10, and anticipate differentially modulated immunopathology. These experiments will help to understand the regulation of immunopathological versus protective responses to chlamydial infections, and thus aid in the design of effective chlamydial vaccines, and of strategies to ameliorate the sequelae of these infections.
The specific aims of the proposed studies are: (1) Generate a homozygous, inbred mouse line immunotolerant to C. pneumoniae hsp60 through transgenic thymic expression of hsp6O during embryonic development (hsp60+ mouse). (2) Induce peripheral immunological tolerance or hyporesponsiveness to C. pneumoniae hsp60 in mice through oral administration of hsp60 covalently conjugated to cholera toxin B subunit (hsp60-orally tolerized mouse). (3) Induce immunological hyperresponsiveness to C. pneumoniae hsp60 in mice through intradermal injection of free plasmid DNA capable of coexpressing hsp60 with murine IL-12 or IL-10 in epithelial cells (IL-12 or IL-10 hsp60 DNA-vaccinated mouse). (4) Characterize the immune response to C. pneumoniae hsp60 and (5) analyze and compare disease pathogenesis and immune responses to experimental C. pneumoniae pneumonitis in unmanipulated mice, in hsp60+ mice, in hsp60-orally tolerized mice, and in IL-12 and IL- 10 hsp6O DNA-vaccinated mice. Humoral immunity will be assessed by enzyme immunoassays and immunoblot assays, and cell mediated immune responses will be analyzed by delayed type hypersensitivity and lymphoproliferation tests. Tissue sections of lung will be scored for seventy of lesions. Distribution of chlamydiae will be evaluated by cell culture isolation and in situ polymerase chain reaction (PCR). Inflammatory cell types, and TH1, TH2, and fibrogenic cytokine expression will be determined by immunohistochemical staining and reverse transcriptase in situ PCR.
