Abstract Body (Do not enter title and authors here): Background: Previous results indicate that inflammation induced by Porphyromonas gingivalis (P. gingivalis), a key pathogen in periodontal disease, enhances foam cell formation and atheromatous plaques development.
Objective: To determine the impact of P. gingivalis on foam cell formation in macrophages exposed to oxidized LDL (Ox-LDL) in atherosclerotic plaque development and to evaluate the pro-resolving actions of Resolvin D2 (RvD₂), a pro-resolution lipid mediator, on this process.
Methods: Peripheral blood mononuclear cells were isolated using Ficoll-Paque density gradient method (N=4 subjects). Monocytes were purified using CD14 microbeads and magnetic separation, differentiated into macrophages over six days, and treated across four groups: Control, Ox-LDL, Ox-LDL+P. gingivalis, and Ox-LDL+P. gingivalis + RvD₂. Cells were stained at 12, 24, and 48 hours using Oil Red O to assess lipid accumulation. Microscopic images of four areas in each well/group were taken. Quantification was performed using the lipid-laden macrophage index based on lipid staining intensity in cell cytoplasm: 0=no staining, 1=up to 25% stained, 2=25-50% stained, 3=50-75% stained, and 4=fully stained using ImageJ. Percentages were compared across groups and time points using multiple comparisons.
Results: Foam cell formation was increased with Ox-LDL alone or with P. gingivalis. Cells showed clustering and characteristics of becoming foam cells. Ox-LDL increased intensity of Grade 3 and 4 staining at all time points (31.52±13.78%, 51.51±15.67%, and 60.72±4.51%, respectively). P. gingivalis amplified clustering and staining intensity (38.54±3.84%, 63.88±16.27%, and 79.43±15.36%, p<0.05 compared to control). RvD₂ reduced lipid accumulation to 21.22±2.09%, 39.17±8.39%, and 58.1±19.52%, respectively. Over time, there was a significant increase in foam cell formation caused by P. gingivalis. RvD₂ specifically reduced the formation of higher-grade foam cells compared to P. gingivalis+OxLDL group (-17.33, -24.71, and -21.33, respectively, p<0.05).
Conclusion: P. gingivalis contributes to foam cell formation by increasing internalization of lipids and promoting clustering, contributing to plaque instability. RvD₂ reduces foam cell formation, clustering, and lipid droplet size suggesting that it has the potential to alleviate inflammation and atherosclerotic changes caused by P. gingivalis.
Keywords: Porphyromonas gingivalis, Foam Cells, Resolvin D2, Atherosclerosis, Oxidized LDL