Release and activation of matrix metalloproteinase 8 from human neutrophils triggered by the leukotoxin 9 of Actinobacillus actinomycetemcomitans

Matrix metalloproteinase 8 (MMP 8) degrades type I collagen and may be involved in the pathogenesis of periodontitis. Latent MMP 8 is stored in neutrophil granules and can be activated when released extracellularly. The periodontitis-associated bacterium Actinobacillus actinomycetemcomitans produces an RTX-toxin, leukotoxin, that degranulates and lyses human neutrophils. This study deals with the ability of leukotoxic A. actinomycetemcomitans to trigger the release and activation of MMP 8. Whole bacteria of three A. actinomycetemcomitans strains or leukotoxin purified from the highly toxic strain HK 1519 were incubated with human neutrophils. The extracellularly released latent and active forms of MMP 8 were detected by an immunoblot technique using specific antibodies against the protease. The activity of MMP 8 was determined by a collagen degradation assay. All strains induced release and activation of MMP 8. The effect was more pronounced under aerobic than anaerobic conditions and correlated with the leukotoxicity of the strains. Pure leukotoxin also induced MMP 8 release and activation in a concentration-dependent manner. Under aerobic conditions, oxidising substances formed by the neutrophils contributed to the rapid activation of the latent enzyme. Upon anaerobic incubation, the activation was slow and mainly caused by other proteases released during neutrophil degranulation. The activation was totally abolished in the presence of serum, probably due to the serum-protease inhibitors. Compared to the calcium ionophore A 23187, a well-known stimulus of neutrophil degranulation, leukotoxin was a more powerful inducer of MMP 8 release, since it triggered the process at a 1000-fold lower concentration. The present findings reveal a specific mechanism that can be induced by A. actinomycetemcomitans leukotoxin and which may contribute to the degradation of periodontal tissues under certain conditions.