ABSTRACT
Several studies have shown a clear association between periodontal disease and increased risk of cardiovascular disease. Porphyromonas gingivalis (Pg), a key oral pathogen, and its cell surface-expressed gingipains, induce oedema in a zebrafish larvae infection model although the mechanism of these vascular effects is unknown. Here, we aimed to determine whether Pg-induced vascular damage is mediated by gingipains. In vitro, human endothelial cells from different vascular beds were invaded by wild-type (W83) but not gingipain-deficient (ΔK/R-ab) Pg. W83 infection resulted in increased endothelial permeability as well as decreased cell surface abundance of endothelial adhesion molecules PECAM-1 and VE-cadherin compared to infection with ΔK/R-ab. In agreement, when transgenic zebrafish larvae expressing fluorescently labelled PECAM-1 or VE-cadherin were systemically infected with W83 or ΔK/R-ab, a significant reduction in adhesion molecule fluorescence was observed specifically in endothelium proximal to W83 bacteria through a gingipain-dependent mechanism. Furthermore, this was associated with increased vascular permeability in vivo when assessed by dextran leakage microangiography. These data are the first to show that Pg directly mediates vascular damage in vivo by degrading PECAM-1 and VE-cadherin. Our data provide a molecular mechanism by which Pg might contribute to cardiovascular disease.
Subject(s)
Bacteroidaceae Infections/etiology , Cardiomegaly/etiology , Edema/etiology , Endothelial Cells/drug effects , Gingipain Cysteine Endopeptidases/toxicity , Porphyromonas gingivalis/pathogenicity , Animals , Animals, Genetically Modified , Antigens, CD/genetics , Antigens, CD/metabolism , Bacteroidaceae Infections/genetics , Bacteroidaceae Infections/metabolism , Bacteroidaceae Infections/pathology , Cadherins/genetics , Cadherins/metabolism , Capillary Permeability/drug effects , Cardiomegaly/genetics , Cardiomegaly/metabolism , Cardiomegaly/pathology , Edema/genetics , Edema/metabolism , Edema/pathology , Embryo, Nonmammalian , Endothelial Cells/metabolism , Endothelial Cells/pathology , Fluorescein Angiography , Gene Expression/drug effects , Genes, Reporter , Gingipain Cysteine Endopeptidases/biosynthesis , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Humans , Larva/drug effects , Larva/microbiology , Platelet Endothelial Cell Adhesion Molecule-1/genetics , Platelet Endothelial Cell Adhesion Molecule-1/metabolism , Porphyromonas gingivalis/growth & development , Porphyromonas gingivalis/metabolism , Primary Cell Culture , Signal Transduction , ZebrafishABSTRACT
The aim of this study was to explore the effects of platelet-rich plasma on gingipain-caused changes in cell morphology and apoptosis of osteoblasts. Mouse osteoblasts MC3T3-E1 cells were treated with gingipain extracts from Porphyromonas gingivalis in the presence or absence of platelet-rich plasma. Apoptosis was detected with terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining. F-actin was determined by phalloidin-fluorescent staining and observed under confocal microscopy. Western blot analysis was used to detect integrin ß1, F-actin, and G-actin protein expressions. A knocking down approach was used to determine the role of integrin ß1. The platelet-rich plasma protected osteoblasts from gingipain-induced apoptosis in a dose-dependent manner, accompanied by upregulation of integrin ß1. Platelet-rich plasma reversed the loss of F-actin integrity and decrease of F-actin/G-actin ratio in osteoblasts in the presence of gingipains. By contrast, the effects of platelet-rich plasma were abrogated by knockdown of integrin ß1. The platelet-rich plasma failed to reduce cell apoptosis and reorganize the cytoskeleton after knockdown of integrin ß1. In conclusion, platelet-rich plasma inhibits gingipain-induced osteoblast apoptosis and actin cytoskeleton disruption by upregulating integrin ß1 expression.
