Polymorphonuclear leukocyte degranulation induced by leukotoxin from Actinobacillus actinomycetemcomitans

The ability of leukotoxin from Actinobacillus actinamycetemcomitans to indu ce release of lysosomal constituents was studied with human polymorphonucle ar leukocytes (PMNL). Leukotoxin purified from A. actinomycetemcomitans or bacterial cells of a leukotoxic strain were mixed with human PMNL and the s uspension was incubated under anaerobic conditions. Samples were taken at c ertain time intervals to examine the cell morphology of PMNL by electron mi croscopy and the extracellular concentrations of the granule components lac toferrin and elastase by enzyme-linked immunosorbent assay (ELISA). Electro n microscopy revealed that within 10 min of exposure to leukotoxin, the num ber of intracellular granules was markedly reduced and the remaining granul es were translocated to the periphery in PMNL. At the same time, the extrac ellular concentrations of lactoferrin and elastase were elevated, while tha t of the cytosolic enzyme lactate dehydrogenase, an indicator of cell lysis , remained low. The lysosome molecules CD63 and CD66b were also exposed on the PMNL surface, indicating fusion of lysosomes with the plasma membrane. These effects were completely abolished by the addition of anti-leukotoxin serum. Pre-incubation of PMNL with monoclonal antibodies to CD11a and CD18 that recognize alpha- and beta-chains of the LFA-1 integrin, a leukotoxin r eceptor on PMNL, inhibited the cytolysis, but not the release of granule co mponents. The present results demonstrate the ability of A. actinomycetemco mitans leukotoxin to trigger a rapid release of lysosomal compounds in huma n PMNL. The release is due to an active process stimulated by the interacti on of PMNL with the toxin or toxin-carrying bacteria.