Reeves, Emer Patricia; (2002) Killing of microbes by neutrophils, with particular reference to the role of proteolytic enzymes. Doctoral thesis (Ph.D), UCL (University College London).

Text out.pdf Download (17MB)

Abstract

According to the hitherto accepted view, neutrophils kill ingested microorganisms by subjecting them to highly toxic reactive oxygen species (ROS) and myeloperoxidase catalysed halogenation. Work presented in this project shows that this simple scheme, which for many years has served as a satisfactory working hypothesis, is inadequate. It was found that mice deficient in neutrophil granule proteases but normal in respect of superoxide production and iodination, are unable to resist staphyloccal and candidal infections. It was further shown that accumulative ROS concentrations could reach some 4 mols/l within the phagocytic vacuole. To compensate for the movement of charge incurred in making superoxide, a large concentration of potassium ions (K+) cross the membrane. The rate of superoxide generation stimulated by phorbol 12-myristate 13-acetate (PMA) is accelerated in the presence of valinomycin, a specific K+ ionophore, and reduced by 4-aminopyridine, a K+ channel inhibitor. Diphenylene iodonium (DPI), an inhibitor of the oxidase blocks the release K+ of and the activation of the K+ channel is pH-dependent, being inhibited at pHs above 8.0. The massive rise in ionic strength within the vacuole engenders the release of cationic granule proteins, including elastase and cathepsin G, from the anionic sulphated proteoglycan matrix. It is these proteases that destroy the bacterium. The inferred killing mechanism is confirmed by the lack of solubilized granule proteases in CGD neutrophils or those treated with DPI. In addition the bactericidal properties of hydrogen peroxide and hypochlorous acid were re-examined, illustrating, that neither oxidants possessed adequate effects against staphylococci and E. coli in vitro. Killing that is promoted by the addition of potassium to the granules is eliminated by protease inhibitors.

Download activity - last month

Export as XMLJSONCSV

Download activity - last 12 months

Export as JSONXMLCSV

Downloads by country - last 12 months

Export as CSVXMLJSON

Archive Staff Only

View Item