This application will study the interaction of Pl. aeruginosa mucoid exopolysaccharide (MEP) antigens with the mammalian immune system to characterize the cells and cell circuits involved which regulate production of opsonic-killing antibody to MEP. MEP is the major cell surface polysaccharide produced by mucoid strains of P. aeruginosa, recovered principally from the sputa of cystic fibrosis (CF) patients. Older (>12 years) CF patients lacking mucoid P.l. aeruginosa colonization have a high proportion of opsonic-killing antibody, while only low levels of MEP- specific opsonic-killing antibody are found in CF patients chronically colonized with P. aeruginosa. These colonized patients, along with normal humans and younger (<12) CF patients lacing P. aeruginosa colonization, have antibody which binds to MEP. However, this antibody is not functional, ie. it does not mediate opsonic-killing and in the presence of these antibodies it is difficult to elicit opsonic-killing antibody either via infection or immunization. We have duplicated in mice an immunologic state which mimics the human response to MEP. Immunization of mice with high doses of MEP, which elicit only non-opsonic-killing antibody, results in a state of non-responsiveness to production of opsonic-killing antibody following immunization with low doses of MEP. The nonresponsive state can be transferred with T-cells from mice immunized with high doses of MEP. We will study how these suppressor T-cells effect opsonic-killing antibody producing B-cells by looking for mechanisms such as idiotype recognition that could be involved in regulation. Methods will be employed to isolate the appropriate cells from mice immunized with opsonic-killing antibody- inducing doses of MEP (1 mg) or non-opsonic-killing antibody-inducing doses (50mug), transfer these cells to naive mice and immunize them with 1 mug to assess their responsiveness. Manipulations of the cells using appropriate reagents to cell surface markers and putative anti-idiotype recognition structures will provide information about the cell's characteristics. We will also explore the immunologic properties of protein-polysaccharide and protein oligosaccharide conjugates. We will isolate oligosaccharide fragments of MEP which bind to either opsonic-killing or non-opsonic- killing antibody to identify these structures. They will then be coupled to a toxoid of P. aeruginosa exotoxin A. Investigations in mice will determine if carrier specific epitopic suppression develops to these conjugates to ensure that immunization with conjugates does not lead to a state of nonresponsiveness after the initial series of immunizations. Conjugates will provide important information about the utility of this methodology for making a vaccine to elicit opsonic-killing antibody to MEP. Overall we are pursuing a program designed to define parameters of immunologic resistance of CF patients to mucoid P. aeruginosa infection, evaluate means of inducing the desired immune effectors in animals,, with the goal of producing a vaccine for this patient population.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI022806-06
Application #
3134339
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Project Start
1985-12-01
Project End
1995-02-28
Budget Start
1991-03-01
Budget End
1992-02-29
Support Year
6
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02115
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