Supplementary Material for: <b><i>P. gingivalis</i></b> Peptidyl Arginine Deiminase Can Modulate Neutrophil Activity via Infection of Human Dental Stem Cells

Periodontitis (PD) is a widespread chronic inflammatory disease in the human population. <i>Porphyromonas gingivalis</i> is associated with PD and can citrullinate host proteins via <i>P. gingivalis</i> peptidyl arginine deiminase (PPAD). Here, we hypothesized that infection of human dental follicle stem cells (hDFSCs) with <i>P. gingivalis</i> and subsequent interaction with neutrophils will alter the neutrophil phenotype. To test this hypothesis, we established and analyzed a triple-culture system of neutrophils and hDFSCs primed with <i>P. gingivalis</i>. Mitogen-activated pathway blocking reagents were applied to gain insight into stem cell signaling after infection. Naïve hDFSCs do not influence the neutrophil phenotype. However, infection of hDFSCs with <i>P. gingivalis</i> prolongs the survival of neutrophils and increases their migration. These phenotypic changes depend on direct cellular contacts and PPAD expression by <i>P. gingivalis</i>. Active JNK and ERK pathways in primed hDFSCs are essential for the phenotypic changes in neutrophils. Collectively, our results confirm that <i>P. gingivalis</i> modifies hDFSCs, thereby causing an immune imbalance.