Wolinella recta is a Gram-negative, anaerobic bacterium that has been implicated in oral and extraoral infections and that is frequently isolated from periodontal lesions of patients with immune deficiencies, such as diabetics and AIDS patients. The W. recta lipopolysaccharide (LPS) has a distinctive chemical composition with an enteric type lipid A and a polysaccharide like that of phytopathogenic bacteria. The LPS and its lipid A and polysaccharide fractions are potentially significant virulence factors. They have exceptional capacity to elicit the secondary immune mediators, prostaglandin E (PGE), interleukin-1 (Il-1) and tumor necrosis factor (TNF), from monocytes and macrophages and to indirectly induce PGE production in human gingival fibroblasts. Among a number of endogenous immune mediators that are over-produced by host cells in response to challenge with LPS, PGE is of interest in the study of periodontopathogenic bacteria, because it mediates activities that contribute to the pathogenesis of periodontal disease. The goal of this project is to locate the site of interaction of the W. recta LPS with the phosphatidyl inositol (PI) cycle and to determine the biologically active part of the LPS. The research will test two hypotheses: 1. the W. recta LPS is a major virulence factor which orchestrates the tissue destruction that is characteristic of periodontal disease through initiation of the arachidonic acid cascade; 2. the LPS mediates this activity through specific enzymes in the biochemical pathway of the PI cycle.
The specific aims of the project are to: I. purify the W. recta lipid A, O-antigen and core oligosaccharide. II. determine the linkages of the fatty acids present in the lipid A. III. locate the site of interaction of the LPS and LPS fractions with the PI cycle of human peripheral monocytes, platelets, murine macrophages, fibroblasts and HL-60 cells. IV. prepare monoclonal antibodies against the polysaccharide and lipid A moieties of the W. recta LPS. Radioimmunoassay techniques will be used to determine the mammalian cell types from which purified LPS and LPS fractions elicit arachidonic acid metabolites. Subsequently, virulence factor stimulation of the PI cycle and arachidonic acid cascade will be examined in radiolabeled target cells in the presence of effectors specific to the enzymes of those metabolic pathways and monoclonal antibodies to the LPS. These studies will contribute to the evaluation of monoclonal anti-LPS antibodies and inhibitors of arachidonic acid metabolism as agents in the treatment, prevention and study of periodontal disease and other inflammatory disorders. They will advance our understanding of how bacterial virulence factors interact with regulatory biochemical pathways to manipulate host cell responses, and they will provide the base for future long term studies on the feasibility of using LPS derived reagents, that are capable of immune modulation, in therapy or research.
| Gillespie, M J; Wright, L; Barton, L L (1995) Energetics of molecular hydrogen oxidation in the oral pathogen Campylobacter rectus. Clin Infect Dis 20 Suppl 2:S172-3 |
| Gillespie, M J; Smutko, J; Haraszthy, G G et al. (1993) Isolation and partial characterization of the Campylobacter rectus cytotoxin. Microb Pathog 14:203-15 |
| Gillespie, J; De Nardin, E; Radel, S et al. (1992) Production of an extracellular toxin by the oral pathogen Campylobacter rectus. Microb Pathog 12:69-77 |
