Project description:We report the study of the mechanism of action of Porphyromonas gingivalis on human oral epithelial cells based on high-throughput sequencing technology. By acting Porphyromonas gingivalis and its metabolites on human oral epithelial cells separately, the mechanism of Porphyromonas gingivalis evading immune surveillance and causing local and deep tissue diffusion to induce systemic diseases was studied. This study provides a framework for studying the pathogenic mechanism of Porphyromonas gingivalis.
Project description:Recent epidemiological studies revealed a significant association between oral squamous cell carcinoma (OSCC) and Porphyromonas gingivalis, a major pathogen of periodontal disease. As a keystone pathogen, P. gingivalis is known not only to damage local periodontal tissue, but also to evade from the host immune system and eventually affect systemic health. However, its role in OSCC has yet to be well studied. To explore the underlying effect of chronic P. gingivalis infection on OSCC and to identify relevant biomarkers as promising targets for therapy and prevention, we established a novel model by exposing human immortalized oral epithelial cells (HIOECs) to P. gingivalis at a low MOI for 5 to 23 weeks. The P. gingivalis-infected HIOECs were monitored in tumor biological alteration. Bioinformatics analyses were performed on HIOECs infected for 15 weeks, and some selected data were validated by q-PCR and (or) Western blot on cells infected for 15 and 23 weeks. Accordingly, we found that persistent exposure to P. gingivalis induced tumor biological properties on HIOECs. Tumor-related genes such as NNMT, FLI1, GAS6, lncRNA CCAT1, PDCD1LG2 and CD274 were aberrantly expressed in response to long-term exposure of P. gingivalis. In addition, some clinical biomarkers and novel proteins were presented. In conclusion, chronic P. gingivalis infection may be a potential risk factor of OSCC. The key regulators and biomarkers might be used in monitoring OSCC with chronic periodontal infection. We applied lncRNA Microarray (Affymetrix) which could simultaneously detect coding and non-coding genes to explore the differences of cell gene expression after long-term exposure to P. gingivalis.
Project description:Transcriptional profiling was utilized to define the biological pathways of gingival epithelial cells modulated by co-culture with the oral pathogenic Porphyromonas gingivalis and Aggregatibacter (formerly actinobacillus) actinomycetemcomitans. We used microarrays to detail the global programme of gene expression underlying infection and identified distinct classes of up- and down-regulated genes during this process. Experiment Overall Design: Gingival epithelial HIGK cells were sham infected (CTRL) and infected with either the oral pathogenic P. gingivalis (Pg) or A. actinomycetemcomitans (Aa). These samples were hybridized to Affymetrix microarrays. Understanding how host cells have adapted to pathogens, and how barrier cells respond to limit their impact, provides a mechanistic biological basis of microbial disease in the mixed bacterial-human ecosystem of the oral cavity.
Project description:Porphyromonas gingivalis is a keystone oral pathogen that successfully manipulates the human innate immune defenses, resulting in a chronic pro-inflammatory state of periodontal tissues and beyond. Here we demonstrate that secreted outer membrane vesicles (OMVs) are deployed by P. gingivalis to selectively coat and activate human neutrophils, thereby provoking degranulation without neutrophil killing. Secreted granule components with antibacterial activity, especially LL-37 and MPO, are subsequently degraded by potent OMV-bound proteases known as gingipains, thereby ensuring bacterial survival. In contrast to neutrophils, the P. gingivalis OMVs are efficiently internalized by macrophages and epithelial cells. Importantly, we show that neutrophil coating is a conserved feature displayed by OMVs of at least one other oral pathogen, namely Aggregatibacter actinomycetemcomitans. Altogether, we conclude that P. gingivalis deploys its OMVs for a neutrophil-deceptive strategy to escape phagocytosis and to create a favorable inflammatory niche.
Project description:The human oral pathogen Porphyromonas gingivalis colonizes the gingival crevice and invades gingival epithelial cells. Multidimensional capillary high-performance liquid chromatography coupled with tandem mass spectrometry and two-dimensional gel electrophoresis were used to analyze the proteome of P. gingivalis as it adapts to a set of experimental conditions designed to reflect important features of an epithelial cell environment. 1014 proteins (46% of the total theoretical proteome) were identified in four independent analyses; 479 of these proteins showed evidence of differential expression after exposure of P. gingivalis to either conditioned epithelial cell growth medium or control conditions: i.e., they were only detected under one set of conditions. Moreover, 276 genes annotated as hypothetical were found to encode expressed proteins. Among the proteins up-regulated in the presence of epithelial cell components were a homolog of the internalin proteins of Listeria monocytogenes and subunits of the ATP-dependent Clp protease complex. Insertional inactivation of clpP, encoding the Clp proteolytic subunit, resulted in approximately a 50% reduction in invasion of P. gingivalis. These results suggest that adaptation to an epithelial cell environment induces a major shift in the expressed proteome of the organism. Furthermore, ClpP, that is up-regulated in this environment, is required for optimal invasive activity of P. gingivalis. Keywords: proteome analysis of P. gingivalis
Project description:Transcriptional profiling was utilized to define the biological pathways of gingival epithelial cells modulated by co-culture with the oral pathogenic Porphyromonas gingivalis and Aggregatibacter (formerly actinobacillus) actinomycetemcomitans. We used microarrays to detail the global programme of gene expression underlying infection and identified distinct classes of up- and down-regulated genes during this process. Keywords: infection state
Project description:We use high through put RNA sequenceing technology to study the genome-wide expression profile in an oral pathogen Filifactor alocis when co-cultured with another key-stone oral pathogen Porphyromonas gingivalis under anaerobic conditions and oxidative stress conditions.
Project description:Porphyromonas gingivalis secretes cysteine proteases named gingipains which can cleave an array of proteins and importantly contribute to the development of periodontitis. In this study we focused on gingipain-exerted proteolysis at the cell surface of human gingival epithelial cells (telomerase immortalized gingival keratinocytes [TIGK]). We examined whether gingipains have sheddase activity or if their main activity is degradation of membrane proteins into small fragments. Using mass spectrometry, we investigated the whole sheddome/degradome of TIGK cell surface proteins by P. gingivalis strains differing in gingipain expression. We observed extensive degradation of TIGK surface proteins, suggesting that gingipains could in fact be the major cause of damage to the gingival epithelium. Most of the identified gingipain substrates were molecules involved in adhesion, suggesting that gingipains may cause tissue damage through cleavage of cell contacts, resulting in cell detachment and rounding, and consequently leading to anoikis. These results reveal a molecular underpinning to P. gingivalis-induced tissue destruction and enhance our knowledge of the role of P. gingivalis’ proteases in the pathobiology of periodontitis.
Project description:Mazumdar2008 - Genome-scale metabolic network
of Porphyromonas gingivalis (iVM679)
This model is described in the article:
Metabolic network model of a
human oral pathogen.
Mazumdar V, Snitkin ES, Amar S,
Segrè D.
J. Bacteriol. 2009 Jan; 191(1):
74-90
Abstract:
The microbial community present in the human mouth is
engaged in a complex network of diverse metabolic activities.
In addition to serving as energy and building-block sources,
metabolites are key players in interspecies and host-pathogen
interactions. Metabolites are also implicated in triggering the
local inflammatory response, which can affect systemic
conditions such as atherosclerosis, obesity, and diabetes.
While the genome of several oral pathogens has been sequenced,
quantitative understanding of the metabolic functions of any
oral pathogen at the system level has not been explored yet.
Here we pursue the computational construction and analysis of
the genome-scale metabolic network of Porphyromonas gingivalis,
a gram-negative anaerobe that is endemic in the human
population and largely responsible for adult periodontitis.
Integrating information from the genome, online databases, and
literature screening, we built a stoichiometric model that
encompasses 679 metabolic reactions. By using flux balance
approaches and automated network visualization, we analyze the
growth capacity under amino-acid-rich medium and provide
evidence that amino acid preference and cytotoxic by-product
secretion rates are suitably reproduced by the model. To
provide further insight into the basic metabolic functions of
P. gingivalis and suggest potential drug targets, we study
systematically how the network responds to any reaction
knockout. We focus specifically on the lipopolysaccharide
biosynthesis pathway and identify eight putative targets, one
of which has been recently verified experimentally. The current
model, which is amenable to further experimental testing and
refinements, could prove useful in evaluating the oral
microbiome dynamics and in the development of novel biomedical
applications.
This model is hosted on
BioModels Database
and identified by:
MODEL1507180038.
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To the extent possible under law, all copyright and related or
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Project description:EXPERIMENT: Microarray expression profiles derived from the human primary gingival epithelial cells 24.0h after exposure to heat inactivated P. gingivalis ANIMAL MODEL: NON EXPOSURE: Human primary gingival epithelial cells (at 3rd passage) were exposed to heat inactivated P. gingivalis (MOI:100) at 90% confluence. Two types of gingival epithelial cells were used. One with Normal cytokine inducer type (at least 2 fold IL-6/TNF-alpha/IL-1à when challenged with TLR2/4 agonists) and the other with diminished cytokine inducer type (no change in IL-6/TNF-alpha/IL-1à when challenged with TLR2/4 agonists). INTERVAL: NON. PLATFORM: microRNA expression profile in gingival epithelial cells - miRCURY LNA⢠microRNA Arrays (Exiqon). The RNA samples were subjected to microarray on 8/9/2007 Keywords = Human primary gingival epithelial cells Keywords = P. gingivalis Keywords = Periodontitis Keywords: Ordered The effect of heat inactivated P. gingivalison human primary gingival epithelial cells were assayed.