Human PBMC Cytokine Response to Aggregatibacter actinomycetemcomitans

Objectives: The gram negative periodontal pathogen, Aggregatibacter actinomycetemcomitans (A.a.), has been strongly associated with Localized Aggressive Periodontitis (LAP). Our goal was to determine the nature of the T and B-cell responses to specific A.a. antigens in order to elucidate the nature of the host-oral pathogen interaction.

Methods: Blood samples were taken from 33 individuals with LAP and matched periodontally healthy individuals. The PBMC T-cells were stimulated in vitro with individual A.a. proteins, including purified leukotoxin (Lkt) and recombinant outer membrane 34 (OMP-34), GroES, GroEL, CdtB, CdtC and CagE. To assess T-cell activation, 3H-thymidine was added to the cultures and proliferation was measured. Cellular supernatants were collected at 72 hrs and the cytokines present were assessed for 5 of the subjects using a human CBA kit (BD). In addition, Western blots were used to measure A.a.-reactive antibodies in the sera.

Results: Most A.a.-positive subjects demonstrated antibody affinity for Lkt; several also had antibody to CagE, GroES and/or OMP-34. T-cell reactivity was seen in fewer subjects, but those that did respond reacted in vitro with most of the A.a. antigens tested. The cytokines produced in vitro were assessed for 2 controls and 3 LAP subjects. Interestingly, all showed an induction of IL-6 in response to Lkt, but not to the recombinant A.a. proteins. In one of the control subjects, very high levels of IFN-ƒ× were seen, regardless of the stimulus, suggesting an on-going infection.

Conclusion: The expression of particular cytokines in response to A.a. or its antigens did not seem to correlate with the presence of LAP. This may reflect the fact that disease in these subjects had occurred years previously so few specific T-cells remained; this is being investigated. This research was supported in part by NIH-DE12052.