There is an emerging paradigm shift from coronary heart disease having a purely hereditary and nutritional causation to possibly having an infectious etiology. Recent epidemiological studies have shown a correlation between periodontal disease and coronary heart disease. However, to date, there is minimal information as to the possible disease mechanisms of this association. It is our hypothesis that invasion of the coronary artery cells by oral bacteria may start and/or exacerbate the inflammatory response in atherosclerosis. Since a few periodontal pathogens have been reported to invade oral epithelial tissues, we tested the ability of three putative periodontal pathogens— Eikenella corrodens , Porphyromonas gingivalis , and Prevotella intermedia —to invade human coronary artery endothelial cells and coronary artery smooth muscle cells. In this study we demonstrate by an antibiotic protection assay and electron microscopy that specific species and strains invade coronary artery cells at a significant level. Actin polymerization and eukaryotic protein synthesis in metabolically active cells were required since the corresponding inhibitors nearly abrogated invasion. Many intracellular P. gingivalis organisms were seen to be present in multimembranous vacuoles resembling autophagosomes by morphological analysis. This is the first report of oral microorganisms invading human primary cell cultures of the vasculature.

Porphyromonas gingivalis is a periodontal pathogen that also localizes to atherosclerotic plaques. Our previous studies demonstrated that P. gingivalis is capable of invading endothelial cells and that intracellular bacteria are contained in vacuoles that resemble autophagosomes. In this study, we have examined the trafficking of P. gingivalis 381 to the autophagic pathway. P. gingivalis 381 internalized by human coronary artery endothelial (HCAE) cells is located within vacuoles morphologically identical to autophagosomes. The progression of P. gingivalis 381 through intracellular vacuoles was analyzed by immunofluorescence microscopy. Vacuoles containing P. gingivalis colocalize with Rab5 and HsGsa7p early after internalization. At later times, P. gingivalis colocalizes with BiP and then progresses to a vacuole that contains BiP and lysosomal glycoprotein 120. Late endosomal markers and the lysosomal cathepsin L do not colocalize with P. gingivalis 381. The intracellular survival of P. gingivalis 381 decreases over 8 h in HCAE cells pretreated with the autophagy inhibitors 3-methyladenine and wortmannin. In addition, the vacuole containing P. gingivalis 381 lacks BiP but contains cathepsin L in the presence of wortmannin. These results suggest that P. gingivalis 381 evades the endocytic pathway to lysosomes and instead traffics to the autophagosome.

Bacteria have evolved a variety of mechanisms to invade eukaryotic cells and survive intracellularly. Once inside, bacterial pathogens often modulate their phagosome to establish an intracellular nic...

Invasion of oral epithelial cells by pathogenic oral bacteria may represent an important virulence factor in the progression of periodontal disease. Here we report that a clinical isolate of Prevotella intermedia , strain 17, was found to invade a human oral epithelial cell line (KB), whereas P. intermedia 27, another clinical isolate, and P. intermedia 25611, the type strain, were not found to invade the cell line. Invasion was quantified by the recovery of viable bacteria following a standard antibiotic protection assay and observed by electron microscopy. Cytochalasin D, cycloheximide, monodansylcadaverine, and low temperature (4°C) inhibited the internalization of P. intermedia 17. Antibodies raised against P. intermedia type C fimbriae and against whole cells inhibited invasion, but the anti-type-C-fimbria antibody inhibited invasion to a greater extent than the anti-whole-cell antibody. This work provides evidence that at least one strain of P. intermedia can invade an oral epithelial cell line and that the type C fimbriae and a cytoskeletal rearrangement are required for this invasion.

Quorum sensing is a phenomenon defined as gene regulation in response to cell density that regulates various functions in bacteria. The periodontopathogenPorphyromonas gingivalispossesses aluxSgene homologue that may encode a quorum-sensing system. In order to identify genes ofP. gingivalisthat are regulated byluxS, gene expression analysis was done using microarrays and RNA samples from the W83 wild-type strain and an isogenicluxSmutant, LY2001. The results indicated that 17 open reading frames (ORFs) in LY2001 are upregulated and two are downregulated. Real-time PCR was done to confirm the microarray results. Among the upregulated ORFs is a group of stress-related genes, includinghtrA,clpB,groEL,dnaK, and the F subunit of alkyl hydroperoxide reductase. This suggested thatluxSis involved in stress gene regulation inP. gingivalis. Stress response experiments, including high-temperature survival, resistance to hydrogen peroxide (H₂O₂), and survival during exposure to low and high pH, were performed on theP. gingivaliswild-type and LY2001 strains. LY2001 had a significantly higher survival rate than did W83 when stressed at 50°C. No difference was found at pH 5, but LY2001 had increased survival compared to W83 at pH 9. LY2001 also survived better than W83 when stressed with 0.35 mM H₂O₂. These results suggest thatluxSmight be involved in promoting survival ofP. gingivalisin the host by regulating its response to host-induced stresses such as temperature, H₂O₂, and pH.