Arginine deiminase and cytokine generation by human monocytes

Objective: Extracellular arginine deiminase (ADI), produced by Streptococcus intermedius suppresses surface-attached growth (biofilm formation) of the periodontal pathogen Porphyromonas gingivalis.  pH can greatly effect the activity of this enzyme. We hypothesized that ADI also modulates the host immune response by enabling an environment that favors formation of a healthy biofilm. The aim of this study was to determine the functional aspects and mechanism of monocyte/macrophage response to ADI.

Method: Peripheral blood from systemically healthy donors was collected and purified monocytes were cultured in the presence/absence of ADI. Three pH conditions (6.6, 7.4, and 8.2) have been tested to assess the host response. Pro-inflammatory cytokine/chemokine levels (IL-1β, IL-6, TNF-α, GM-CSF, and IL-8) were measured via xMAP multiplex assays using supernatants after 6 and 24-hour incubation periods. Lipopolysaccharide from E. coli was used as the positive control and enzymatically inactive ADI was used for comparison. Western blot analysis of cell lysates from these cultures was performed to examine levels of NF-κB expression.

Result: Levels of IL-1β, IL-6, TNF-α, GM-CSF, and IL-8 were slightly elevated in response to ADI compared to control medium at neutral pH (7.4). Cells cultured in medium at pH 6.6 showed a substantial increase in cytokine release compared to those cultured at pH 7.4, with fold changes between 38x (IL-1 β) and 150x (IL-6). Alkaline pH (8.2) did not make any difference in cytokine generation by the monocytes. ADI completely suppressed cytokine release at pH 6.6. NF-κB expression showed a reduction of about 25% in response to treatment of primary monocytes with ADI after 6 hours (p<0.05).

Conclusion: ADI limits inflammation by suppressing the release of various pro-inflammatory cytokines/chemokines (IL-1β, IL-6, TNF-α, GM-CSF, and IL-8) possibly creating a favorable environment for biofilm growth. This response is regulated through NF-κB activation.