The presence of a serum factor that suppresses neutrophil chemiluminescence has been detected following trauma. This factor is present in significant amounts only in the serum of patients who subsequently develop sepsis. This proposal is designed to test the hypothesis that trauma generates a serum suppressive factor responsible for decreased neutrophil bactericidal activity and that this factor contributes to reduced host-resistance to infection after injury. This hypothesis will be tested in achieving the following specific aims:
Aim #1 is to purify the suppressive factor to homogeneity. A progressive purification scheme beginning with Sephadex G100 chromatography and ending with SDS-PAGE will be used for this purpose.
Aim #2 is to biochemically characterize this suppressor. Following its isolation and purification, monospecific antiserum will be produced and confirmed using immunoelectrophoresis. Using immunoelectrophoresis and Ouchterlony double diffusion, the similarity of this factor to other known proteins will be assessed. In addition, this antisera will be used for quantitative measurement of this factor and to facilitate its removal in vitro.
Aim #3 is to determine the mechanism of action of this serum factor.
This aim i s to identify the cellular site of action of this suppressive factor by step-wise analysis of its effect on membrane binding, depolarization, NADPH oxidase, hexose monophosphate shunt activity, superoxide formation, and bacterial killing. The ultimate goal of this project is to establish the biological significance of a newly detected serum-suppressor of neutrophil bactericidal activity in decreasing host resistance to sepsis following injury.
