title: The effects of P. gingivalis and E. coli LPS on the expression of proinflammatory mediators in human mast cells and their relevance to periodontal disease.
abstract: Mast cells (MCs) are tissue-resident immune cells that participate in a variety of allergic and inflammatory conditions, including periodontal disease, through the release of cytokines, chemokines and proteolytic enzymes. Porhyromonas gingivalis (P. g) is widely recognized as a major pathogen in the development and progression of periodontitis. Here we compared the differential effects of lipopolysaccharides (LPS) from P. g and E. coli on the expression and production of tumor necrosis factor (TNF), vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein (MCP-1) by human MCs. Human LAD2 MCs were stimulated with LPS from either P. g or E. coli (1-1000 ng/ml). MCs were also stimulated with SP (2μM) serving as the positive control or media alone as the negative control. After 24 h, the cells and supernatant fluids were collected and analyzed for β-Hexosaminidase (β-hex) spectrophotometrically, TNF, VEGF and MCP-1 release by ELISA and real-time polymerase chain reaction (PCR) for mediator gene expression, respectively. To assess the functional role of tolllike receptors (TRL) in mediator release, MCs were pre-incubated with either anti-TLR2 or anti- TLR4 (2 μg/ml) polyclonal antibody for 1 h before stimulation with LPS. When MCs were stimulated with SP (2 μM), there was a statistically significant β-hex release as well as release of TNF, VEGF and MCP-1. Stimulation of MCs with either type of LPS did not induce degranulation based on the lack of β-hex release. However, both types of LPS stimulated expression and release of TNF, VEGF and MCP-1. Although, P. g LPS induced significant release of TNF, VEGF and MCP-1, the effect was not concentration-dependent. There was no statistically significant difference between the effects of P. g and E. coli LPS. P. g LPS stimulated TNF through TLR-2 while E. coli utilized TRL-4 instead. In contrast, VEGF release by P. g LPS required both TRL-2 and TRL-4 while E. coli LPS required TLR-4. Release of MCP-1 was independent of TLR-2 or TLR-4. P. g LPS activates human MCs to generate and release TNF, VEGF and MCP-1 through different TLRs than E. coli LPS. MCs may, therefore, be involved in the inflammatory processes responsible for periodontal disease.
authors.affiliation: Molecular Immunopharmacology and Drug Discovery Laboratory, Department of Integrative Physiology and Pathobiology, Tufts University School of Medicine, Boston, MA, USA.
authors.forename: I
authors.initials: I
authors.lastname: Palaska
authors.affiliation: Oral Medicine Clinics, A. Syggros Hospital of Dermatologic and Venereal Diseases, Department of Dermatology, School of Medicine, University of Athens, Greece.
authors.forename: E
authors.initials: E
authors.lastname: Gagari
authors.affiliation: Molecular Immunopharmacology and Drug Discovery Laboratory, Department of Integrative Physiology and Pathobiology, Tufts University School of Medicine, Boston, MA, USA; Oral Medicine Clinics, A. Syggros Hospital of Dermatologic and Venereal Diseases, Department of Dermatology, School of Medicine, University of Athens, Greece; Department of Internal Medicine, Tufts University School of Medicine, MA, USA.
authors.forename: T C
authors.initials: TC
authors.lastname: Theoharides
mesh_terms: D015550:Cell Degranulation / Q000187:drug effects; D045744:Cell Line, Tumor; D018932:Chemokine CCL2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D004731:Endotoxins / Q000494:pharmacology*; D004926:Escherichia coli / Q000737:chemistry / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D018836:Inflammation Mediators / Q000502:physiology*; D007946:Leukemia, Mast-Cell / Q000473:pathology; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008407:Mast Cells / Q000187:drug effects* / Q000502:physiology; D010518:Periodontitis / Q000276:immunology* / Q000382:microbiology; D016966:Porphyromonas gingivalis / Q000737:chemistry / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D013373:Substance P / Q000494:pharmacology; D051195:Toll-Like Receptor 2 / Q000037:antagonists & inhibitors / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000037:antagonists & inhibitors / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism
chemical_list: C111835:CCL2 protein, human; D018932:Chemokine CCL2; D004731:Endotoxins; D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D012333:RNA, Messenger; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D013373:Substance P; C044101:endotoxin, Escherichia coli