Neutrophils in chronic and aggressive periodontitis in interaction with Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans

Abstract

Background and objective: This study analyzed the interaction of Porphyromonas gingivalis ATCC 33277 and Aggregatibacter actinomycetemcomitans Y4 with peripheral blood polymorphonuclear neutrophils taken from patients with aggressive periodontitis and chronic periodontitis.

Material and methods: Peripheral blood polymorphonuclear neutrophils obtained from 12 patients with chronic periodontitis, six patients with aggressive periodontitis and 12 healthy controls were exposed to P. gingivalis and A. actinomycetemcomitans following opsonization of the bacteria using the patient's own serum. Serum immunoglobulin G (IgG) levels against both periodontopathogens were measured. Phagocytosis and killing of the bacteria, as well as the extracellular human neutrophil elastase activity, were quantified. The total amount and the extracellular release of reactive oxygen species were measured using luminol-dependent and isoluminol-dependent chemiluminescence.

Results: Polymorphonuclear neutrophils from patients with chronic (62.16 +/- 19.39%) and aggressive (43.26 +/- 26.63%) periodontitis phagocytosed more P. gingivalis than the healthy controls (24.43 +/- 19.87%) at the 30-min time point after exposure to the bacteria (p < 0.05). High serum IgG levels against P. gingivalis and A. actinomycetemcomitans were detected in subjects with periodontitis. Polymorphonuclear neutrophils from subjects with chronic and aggressive periodontitis released significantly more reactive oxygen species and demonstrated greater human neutrophil elastase activity in the absence of any stimulus than polymorphonuclear neutrophils from healthy controls (p < 0.05). Polymorphonuclear neutrophils in chronic periodontitis released significantly more reactive oxygen species when exposed to P. gingivalis and A. actinomycetemcomitans than polymorphonuclear neutrophils in aggressive periodontitis.

Conclusion: High serum IgG levels against P. gingivalis and A. actinomycetemcomitans promote phagocytosis in periodontitis. The extracellular release of reactive oxygen species and neutrophil elastase by polymorphonuclear neutrophils may also contribute to damage of the surrounding periodontal tissues.

Figure 1

Phagocytosis and killing of P. gingivalis. PMNs from patients with chronic periodontitis phagocytosed significantly more P. gingivalis at the 30 min and 60 min time point than PMNs from controls (* p<0.05) (A). The percentage of killed bacteria followed a similar pattern (B). PMNs in chronic periodontitis killed more of the phagocytosed bacteria than PMNs from subjects with aggressive periodontitis or controls at 30 and 120 min (C).

Figure 2

Phagocytosis and Killing of Aggregatibacter actinomycetemcomitans. PMNs from aggressive periodontitis patients phagocytosed more A. actinomycetemcomitans than PMNs from controls at the 30 min and 120 min observations (* p<0.05) and more than PMNs of patients with chronic periodontitis after 120 minutes (A). The killing of the phagocytosed bacteria correlated with this observation (B). Nearly all phagocytosed bacteria were also killed by the PMNs, as indicated by the high killed/phagocytosed ratios (C).

Figure 3

Systemic antibodies to P. gingivalis (A) and A. actinomycetemcomitans (B) in aggressive and chronic periodontitis in comparison to periodontally healthy subjects. Patients with periodontitis had significantly higher titres of anti-P. gingivalis IgG than the controls (A). Only in subjects with aggressive periodontitis was the IgG titre for anti-A. actinomycetemcomitans significantly higher than in the periodontally-healthy controls (B).

Figure 4

Total (A) and extracellular (B) release of reactive oxygen species (ROS) by peripheral blood PMNs from chronic and aggressive periodontitis patients in comparison to healthy controls determined by chemoluminescence (counts). (A) PMNs in chronic and aggressive periodontitis release significantly more ROS than PMNs from controls in the absence of stimulation (* p<0.05). PMNs in aggressive periodontitis release significantly less ROS than PMNs from subjects with chronic periodontitis (#) and controls (*) after stimulation with P. gingivalis (P.g.) and A. actinomycetemcomitans (A.a.). (B) The extracellular release of ROS by PMNs from patients with aggressive periodontitis was also lower than that from PMNs of subjects with chronic periodontitis (# p<0.05). PMNs in chronic periodontitis released significantly more ROS after interaction with A. actinomycetemcomitans than PMNs from the controls (* p<0.05).