Role of mitogen-activated protein kinase pathways in migration of gingival epithelial cells in response to stimulation by cigarette smoke condensate and infection by Porphyromonas gingivalis.

Abstract

Previous studies have shown that cigarette smoke (CS) and periodontal pathogens could alter wound healing responses of gingival epithelial cells. To elucidate molecular mechanisms leading to these epithelial changes, we studied the signaling pathway involved in the modulation of cell migration by CS condensate (CSC) and the infection by a prominent periodontal pathogen, Porphyromonas gingivalis. Human gingival epithelial cells (Ca9-22) were treated with CSC or vehicle control for 24 h. Activation of mitogen-activated protein kinases (MAPK) in cells with or without infection by P. gingivalis was assessed by polymerase chain reaction array and immunoblotting using phospho-specific antibodies. Cell migration was assessed using in vitro wound closure model, and specific pharmacologic inhibitors of MAPK pathways were used to characterize further the extent of involvement of the MAPK pathways. Polymerase chain reaction array showed that gene expression of several members of the MAPK, particularly p38 and JNK, was upregulated more than twofold in Ca9-22 cells stimulated with 10 μg/mL CSC. Coincubation with P. gingivalis induced a different pattern of gene expression for MAPK pathways, but it did not suppress the MAPK-related genes upregulated by CSC. A significant phosphorylation of ERK1/2 and p38 was observed in cells stimulated with 10 μg/mL CSC (p < 0.05), whereas coincubation with a higher concentration of CSC (250 μg/mL) evoked no such activation. P. gingivalis infection resulted in a tendency to reduce the phosphorylation of ERK1/2 and p38, which had been enhanced by stimulation with 10 μg/mL CSC. Incubation with ERK1/2 and p38 inhibitors significantly reduced the wound closure of CSC-stimulated cells, by approximately 43% and 46%, respectively (p < 0.05). CSC exerts effects on the migration of human gingival epithelial cells through the activation of the MAPK ERK1/2 and p38 signaling pathways. P. gingivalis infection attenuates the CSC-induced migration at least partly by suppressing the phosphorylation of ERK1/2 and p38, but other pathways are likely to be involved in this modulatory process.