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Caroline A Genco - One of the best experts on this subject based on the ideXlab platform.
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oral infection with a periodontal pathogen alters oral and gut microbiomes
Anaerobe, 2021Co-Authors: Alexandra M Simas, Carolyn D Kramer, Ellen O Weinberg, Caroline A GencoAbstract:Abstract Periodontal disease, an inflammatory bone disease of the oral cavity, affects more than 50% of the United States population over the age of 30. The Gram-negative, anaerobic bacterium Porphyromonas Gingivalis, the etiological agent of periodontal disease, is known to induce dysbiosis of the oral microbiome while promoting inflammatory bone loss. We have recently reported that P. Gingivalis can also alter the gut microbiota of mice prone to develop inflammatory atherosclerosis. However, it is still unknown whether P. Gingivalis induces similar changes to the gut microbiome as it does to oral microbiome. In this study, we demonstrate that P. Gingivalis infection increases the diversity of the oral microbiome, allowing for colonization of potentially opportunistic species in the oral microbiome and overgrowth of commensal species in both the oral and gut microbiomes. Since periodontal disease treatment in humans typically involves antibiotic treatment, we also examined the combined effect of P. Gingivalis infection on mice pretreated with oral antibiotics. By correlating the oral and cecal microbiota of P. Gingivalis-infected mice fed a normal chow diet, we identified blooms of the Gram-negative genera Barnesiella and Bacteroides and imbalances of mucin-degrading bacteria. These disrupted community structures were predicted to have increased detrimental functional capacities including increased flavonoid degradation and l -histidine fermentation. Though antibiotic pretreatment (without P. gingivlais) had a dominant impact on the cecal microbiome, P. Gingivalis infection of mice with or without antibiotic pretreatment increased the abundance of the phylum Firmicutes and the Porphyromonadaceae family in the cecum. Collectively, our study demonstrates that P. Gingivalis oral infection disrupted the oral and cecal microbiomes of otherwise unperturbed mice, altering their community membership and functional potential.
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fimbria dependent activation of pro inflammatory molecules in porphyromonas Gingivalis infected human aortic endothelial cells
Cellular Microbiology, 2006Co-Authors: Frank C Gibson, Yusuke Takahashi, Michael Davey, Hiromichi Yumoto, Caroline A GencoAbstract:: Epidemiological studies support that chronic periodontal infections are associated with an increased risk of cardiovascular disease. Previously, we reported that the periodontal pathogen Porphyromonas Gingivalis accelerated atherosclerotic plaque formation in hyperlipidemic apoE-/- mice, while an isogenic fimbria-deficient (FimA-) mutant did not. In this study, we utilized 41 kDa (major) and 67 kDa (minor) fimbria mutants to demonstrate that major fimbria are required for efficient P. Gingivalis invasion of human aortic endothelial cells (HAEC). Enzyme-linked immunosorbent assay (ELISA) revealed that only invasive P. Gingivalis strains induced HAEC production of pro-inflammatory molecules interleukin (IL)-1beta, IL-8, monocyte chemoattractant protein (MCP)-1, intracellular adhesion molecule (ICAM)-1, vascular cellular adhesion molecule (VCAM)-1 and E-selectin. The purified native forms of major and minor fimbria induced chemokine and adhesion molecule expression similar to invasive P. Gingivalis, but failed to elicit IL-1beta production. In addition, the major and minor fimbria-mediated production of MCP-1 and IL-8 was inhibited in a dose-dependent manner by P. Gingivalis lipopolysaccharide (LPS). Both P. Gingivalis LPS and heat-killed organisms failed to stimulate HAEC. Treatment of endothelial cells with cytochalasin D abolished the observed pro-inflammatory MCP-1 and IL-8 response to invasive P. Gingivalis and both purified fimbria, but did not affect P. Gingivalis induction of IL-1beta. These results suggest that major and minor fimbria elicit chemokine production in HAEC through actin cytoskeletal rearrangements; however, induction of IL-1beta appears to occur via a separate mechanism. Collectively, these data support that invasive P. Gingivalis and fimbria stimulate endothelial cell activation, a necessary initial event in the development of atherogenesis.
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role for fimbriae and lysine specific cysteine proteinase gingipain k in expression of interleukin 8 and monocyte chemoattractant protein in porphyromonas Gingivalis infected endothelial cells
Infection and Immunity, 2002Co-Authors: Hamdy Nassar, Hsinhua Chou, Mary Khlgatian, Frank C Gibson, T E Van Dyke, Caroline A GencoAbstract:Recent cross-sectional and prospective epidemiological studies have demonstrated an association between periodontal disease and atherosclerosis and human coronary heart disease. Previously, we have established that the periodontal pathogen Porphyromonas Gingivalis is capable of invading aortic, heart, and human umbilical vein endothelial cells (HUVEC). Since atherosclerosis is a chronic inflammatory response initiated at the vascular wall, interactions of P. Gingivalis with endothelial cells and the subsequent host cell response to infection may be important in the pathogenesis of atherosclerosis. In this study we examined the consequences of P. Gingivalis infection of HUVEC on the expression of the chemokines interleukin-8 (IL-8) and monocyte chemotactic protein 1 (MCP-1). HUVEC were found to constitutively produce low levels of IL-8 and MCP-1. The addition of P. Gingivalis fimbrillin-specific peptides, lipopolysaccharides (LPS), or heat-killed whole cell preparations to HUVEC stimulated modest IL-8 and MCP-1 responses. In contrast, coculture of HUVEC with live P. Gingivalis strain A7436, 33277, or 381 abolished the IL-8 and MCP-1 responses. Inhibition of IL-8 and MCP-1 production was not dependent on bacterial adherence since similar results were obtained with the nonadherent P. Gingivalis fimA mutant DPG3 or when P. Gingivalis was preincubated with fimbrillin peptide antisera prior to the addition to HUVEC. Furthermore, treatment of P. Gingivalis-infected HUVEC with cytochalsin D, which prevented P. Gingivalis invasion, also abolished the constitutive IL-8 and MCP-1 responses. Treatment of HUVEC with E. coli LPS stimulated robust IL-8 and MCP-1 responses that were abolished when stimulated cells were cocultured with live P. Gingivalis. Analysis of P. Gingivalis-infected HUVEC cultures by an RNase protection assay revealed an increase in the IL-8 transcript relative to uninfected HUVEC. Pretreatment of P. Gingivalis with protease inhibitors prior to the addition to HUVEC prevented the inhibition of IL-8 and MCP-1 production in P. Gingivalis-infected HUVEC, indicating that the inhibition was proteolytically mediated. Coculture of HUVEC with a P. Gingivalis mutant deficient in lysine-specific cysteine proteinase (gingipain K [Kgp]) resulted in an increase in both IL-8 transcription and protein expression relative to that observed in HUVEC cocultured with the P. Gingivalis wild-type strain. These results indicate that P. Gingivalis can temporally modulate the chemokine response in endothelial cells through both fimbriae and gingipain-mediated mechanisms.
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fimbria dependent activation of cell adhesion molecule expression in porphyromonas Gingivalis infected endothelial cells
Infection and Immunity, 2002Co-Authors: Mary Khlgatian, Hamdy Nassar, Hsinhua Chou, Frank C Gibson, Caroline A GencoAbstract:Porphyromonas Gingivalis is an oral pathogen that has recently been associated with chronic inflammatory diseases such as atherosclerosis. The strength of the epidemiological associations of P. Gingivalis with atherosclerosis can be increased by the demonstration that P. Gingivalis can initiate and sustain growth in human vascular cells. We previously established that P. Gingivalis can invade aortic, heart, and human umbilical vein endothelial cells (HUVEC), that fimbriae are required for invasion of endothelial cells, and that fimbrillin peptides can induce the expression of the chemokines interleukin 8 and monocyte chemotactic protein. In this study, we examined the expression of surface-associated cell adhesion molecules on endothelial cells in response to P. Gingivalis infection by fluorescence-activated cell sorting FACS analysis and confocal microscopy. Coculture of HUVEC with P. Gingivalis strain 381 or A7436 resulted in the induction in the expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and P- and E-selectins, which was maximal at 48 h postinfection. In contrast, we did not observe induction of ICAM-1, VCAM-1, or P- or E-selectin expression in HUVEC cultured with the noninvasive P. Gingivalis fimA mutant DPG3 or when P. Gingivalis was incubated with fimbrillin peptide-specific anti-sera prior to the addition to HUVEC. Furthermore, the addition of a peptide corresponding to the N-terminal domain of fimbrillin to HUVEC resulted in an increase in ICAM-1, VCAM-1, and P- and E-selectins, which was maximal at 48 h and similar to that observed for live P. Gingivalis. Treatment of P. Gingivalis-infected HUVEC with cytochalsin D, which prevented P. Gingivalis invasion, also resulted in the inhibition of ICAM-1, VCAM-1, or P- and E-selectin expression. Taken together, these results indicate that active P. Gingivalis invasion of HUVEC mediated via the major fimbriae stimulates surface-associated cell adhesion molecule expression. Stimulation of adhesion molecules involved in the recruitment of leukocytes to sites of inflammation by P. Gingivalis may play a role in the pathogenesis of systemic inflammatory diseases associated with this microorganism, including atherosclerosis.
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invasion of aortic and heart endothelial cells by porphyromonas Gingivalis
Infection and Immunity, 1998Co-Authors: Rajashri G Deshpande, Mahfuz Khan, Caroline A GencoAbstract:Invasion of host cells is believed to be an important strategy utilized by a number of pathogens, which affords them protection from the host immune system. The connective tissues of the periodontium are extremely well vascularized, which allows invading microorganisms, such as the periodontal pathogen Porphyromonas Gingivalis, to readily enter the bloodstream. However, the ability of P. Gingivalis to actively invade endothelial cells has not been previously examined. In this study, we demonstrate that P. Gingivalis can invade bovine and human endothelial cells as assessed by an antibiotic protection assay and by transmission and scanning electron microscopy. P. Gingivalis A7436 was demonstrated to adhere to and to invade fetal bovine heart endothelial cells (FBHEC), bovine aortic endothelial cells (BAEC), and human umbilical vein endothelial cells (HUVEC). Invasion efficiencies of 0.1, 0.2, and 0.3% were obtained with BAEC, HUVEC, and FBHEC, respectively. Invasion of FBHEC and BAEC by P. Gingivalis A7436 assessed by electron microscopy revealed the formation of microvillus-like extensions around adherent bacteria followed by the engulfment of the pathogen within vacuoles. Invasion of BAEC by P. Gingivalis A7436 was inhibited by cytochalasin D, nocodazole, staurosporine, protease inhibitors, and sodium azide, indicating that cytoskeletal rearrangements, protein phosphorylation, energy metabolism, and P. Gingivalis proteases are essential for invasion. In contrast, addition of rifampin, nalidixic acid, and chloramphenicol had little effect on invasion, indicating that bacterial RNA, DNA, and de novo protein synthesis are not required for P. Gingivalis invasion of endothelial cells. Likewise de novo protein synthesis by endothelial cells was not required for invasion by P. Gingivalis. P. Gingivalis 381 was demonstrated to adhere to and to invade BAEC (0.11 and 0.1% efficiency, respectively). However, adherence and invasion of the corresponding fimA mutant DPG3, which lacks the major fimbriae, was not detected. These results indicate that P. Gingivalis can actively invade endothelial cells and that fimbriae are required for this process. P. Gingivalis invasion of endothelial cells may represent another strategy utilized by this pathogen to thwart the host immune response.
Poppy Mila Fadriani - One of the best experts on this subject based on the ideXlab platform.
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EFEKTIVITAS PHYTO RESPON EKSTRAK ETANOL DAUN KELOR (MORINGA OLEIFERA) TERHADAP VIRULENSI PORPHYROMONAS Gingivalis ATCC 33277 SECARA IN-VITRO
Fakultas Kedokteran Gigi Universitas Mahasaraswati Denpasar, 2020Co-Authors: Poppy Mila FadrianiAbstract:Porphyromonas Gingivalis ATCC 33277 merupakan bakteri biakan murni yang memiliki virulensi paling rendah dibandingkan dengan biakan murni Porphyromonas Gingivalis lainnya. Porphyromonas Gingivalis merupakan bakteri Gram-negatif anaerob obligat yang menjadi salah satu bakteri dominan penyebab periodontitis kronis.Terapi periodontitis kronis meliputi menghilangkan bakteri patogen seperti Porphyromonas Gingivalis. Penggunaan daun kelor(Moringa oleifera) dapat digunakan sebagai terapi untuk mengeliminasi bakteri patogen karena mengandung senyawa antibakteri. Penelitian ini bertujuan untuk mengetahui efektivitas phyto respon ekstrak etanol daun kelor (Moringa oleifera) terhadap virulensi Porphyromonas Gingivalis ATCC 33277. Uji potensi anti virulensi ekstrak etanol daun kelor (Moringa oleifera) terhadap Porphyromonas Gingivalis ATCC 33277 dilakukan dengan menggunakan metode difusi (sumuran) sebanyak 2 kali pengulangan pada media Tryptone Soy Agar (TSA). Konsentrasi ekstrak etanol 96% daun kelor (Moringa oleifera) yang digunakan pada penelitian ini yaitu 75%, 50%, 25%, 12,5%, dan 6,25%. Data hasil penelitian dianalisis menggunakan uji one way ANOVA. Berdasarkan hasil penelitian dapat disimpulkan bahwa ekstrak etanol daun kelor (Moringa oleifera) pada konsentrasi 50% dengan waktu inkubasi 24 jam memiliki efektivitas phyto respon paling kuat terhadap virulensi Porphyromonas Gingivalis ATCC 33277 secara in-vitro.Banda Ace
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EFEKTIVITAS PHYTO RESPON EKSTRAK ETANOL DAUN KELOR (MORINGA OLEIFERA) TERHADAP VIRULENSIPORPHYROMONASGingivalis ATCC 33277 SECARA IN-VITRO
Fakultas Kedokteran Gigi Universitas Mahasaraswati Denpasar, 2020Co-Authors: Poppy Mila FadrianiAbstract:Porphyromonas Gingivalis ATCC 33277 merupakan bakteri biakan murni yang memiliki virulensi paling rendah dibandingkan dengan biakan murni Porphyromonas Gingivalis lainnya. Porphyromonas Gingivalis merupakan bakteri Gram-negatif anaerob obligat yang menjadi salah satu bakteri dominan penyebab periodontitis kronis.Terapi periodontitis kronis meliputi menghilangkan bakteri patogen seperti Porphyromonas Gingivalis. Penggunaan daun kelor(Moringa oleifera) dapat digunakan sebagai terapi untuk mengeliminasi bakteri patogen karena mengandung senyawa antibakteri. Penelitian ini bertujuan untuk mengetahui efektivitas phyto respon ekstrak etanol daun kelor (Moringa oleifera) terhadap virulensi Porphyromonas Gingivalis ATCC 33277. Uji potensi anti virulensi ekstrak etanol daun kelor (Moringa oleifera) terhadap Porphyromonas Gingivalis ATCC 33277 dilakukan dengan menggunakan metode difusi (sumuran) sebanyak 2 kali pengulangan pada media Tryptone Soy Agar (TSA). Konsentrasi ekstrak etanol 96% daun kelor (Moringa oleifera) yang digunakan pada penelitian ini yaitu 75%, 50%, 25%, 12,5%, dan 6,25%. Data hasil penelitian dianalisis menggunakan uji one way ANOVA. Berdasarkan hasil penelitian dapat disimpulkan bahwa ekstrak etanol daun kelor (Moringa oleifera) pada konsentrasi 50% dengan waktu inkubasi 24 jam memiliki efektivitas phyto respon paling kuat terhadap virulensi Porphyromonas Gingivalis ATCC 33277 secara in-vitro.Banda Ace
Özlem Yilmaz - One of the best experts on this subject based on the ideXlab platform.
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Possible role of Porphyromonas Gingivalis in orodigestive cancers.
Journal of oral microbiology, 2019Co-Authors: Ingar Olsen, Özlem YilmazAbstract:There is increasing evidence for an association between periodontitis/tooth loss and oral, gastrointestinal, and pancreatic cancers. Periodontal disease, which is characterized by chronic inflammation and microbial dysbiosis, is a significant risk factor for orodigestive carcinogenesis. Porphyromonas Gingivalis is proposed as a keystone pathogen in chronic periodontitis causing both dysbiosis and discordant immune response. The present review focuses on the growing recognition of a relationship between P. Gingivalis and orodigestive cancers. Porphyromonas Gingivalis has been recovered in abundance from oral squamous cell carcinoma (OSCC). Recently established tumorigenesis models have indicated a direct relationship between P. Gingivalis and carcinogenesis. The bacterium upregulates specific receptors on OSCC cells and keratinocytes, induces epithelial-to-mesenchymal (EMT) transition of normal oral epithelial cells and activates metalloproteinase-9 and interleukin-8 in cultures of the carcinoma cells. In addition, P. Gingivalis accelerates cell cycling and suppresses apoptosis in cultures of primary oral epithelial cells. In oral cancer cells, the cell cycle is arrested and there is no effect on apoptosis, but macro autophagy is increased. Porphyromonas Gingivalis promotes distant metastasis and chemoresistance to anti-cancer agents and accelerates proliferation of oral tumor cells by affecting gene expression of defensins, by peptidyl-arginine deiminase and noncanonical activation of β-catenin. The pathogen also converts ethanol to the carcinogenic intermediate acetaldehyde. In addition, P. Gingivalis can be implicated in precancerous gastric and colon lesions, esophageal squamous cell carcinoma, head and neck (larynx, throat, lip, mouth and salivary glands) carcinoma, and pancreatic cancer. The fact that distant organs can be involved clearly emphasizes that P. Gingivalis has systemic tumorigenic effects in addition to the local effects in its native territory, the oral cavity. Although coinfection with other bacteria, viruses, and fungi occurs in periodontitis, P. Gingivalis relates to cancer even in absence of periodontitis. Thus, there may be a direct relationship between P. Gingivalis and orodigestive cancers.
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a novel kinase function of a nucleoside diphosphate kinase homologue in porphyromonas Gingivalis is critical in subversion of host cell apoptosis by targeting heat shock protein 27
Cellular Microbiology, 2018Co-Authors: Jungnam Lee, Joann S Roberts, Kalina R Atanasova, Nityananda Chowdhury, Özlem YilmazAbstract:We have previously shown that a homologue of a conserved nucleoside-diphosphate-kinase (Ndk) family of multifunctional enzymes and secreted molecule in Porphyromonas Gingivalis can modulate select host molecular pathways including downregulation of reactive-oxygen-species generation to promote bacterial survival in human gingival epithelial cells (GECs). In this study, we describe a novel kinase function for bacterial effector, P. Gingivalis-Ndk, in abrogating epithelial cell death by phosphorylating heat-shock protein 27 (HSP27) in GECs. Infection by P. Gingivalis was recently suggested to increase phosphorylation of HSP27 in cancer-epithelial cells; however, the mechanism and biological significance of antiapoptotic phospho-HSP27 during infection has never been characterised. Interestingly, using glutathione S-transferase-rNdk pull-down analysed by mass spectrometry, we identified HSP27 in GECs as a strong binder of P. Gingivalis-Ndk and further verified using confocal microscopy and ELISA. Therefore, we hypothesised P. Gingivalis-Ndk can phosphorylate HSP27 for inhibition of apoptosis in GECs. We further employed P. Gingivalis-Ndk protein constructs and an isogenic P. Gingivalis-ndk-deficient-mutant strain for functional examination. P. Gingivalis-infected GECs displayed significantly increased phospho-HSP27 compared with ndk-deficient-strain during 24 hr infection. Phospho-HSP27 was significantly increased by transfection of GFP-tagged-Ndk into uninfected-GECs, and in vitro phosphorylation assays revealed direct phosphorylation of HSP27 at serines 78 and 82 by P. Gingivalis-Ndk. Depletion of HSP27 via siRNA significantly reversed resistance against staurosporine-mediated-apoptosis during infection. Transfection of recombinant P. Gingivalis-Ndk protein into GECs substantially decreased staurosporine-induced-apoptosis. Finally, ndk-deficient-mutant strain was unable to inhibit staurosporine-induced Cytochrome C release/Caspase-9 activation. Thus, we show for the first time the phosphorylation of HSP27 by a bacterial effector-P. Gingivalis-Ndk-and a novel function of Ndks that is directly involved in inhibition of host cell apoptosis and the subsequent bacterial survival.
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human primary epithelial cells acquire an epithelial mesenchymal transition phenotype during long term infection by the oral opportunistic pathogen porphyromonas Gingivalis
Frontiers in Cellular and Infection Microbiology, 2017Co-Authors: Jungnam Lee, Joann S Roberts, Kalina R Atanasova, Nityananda Chowdhury, Kyudong Han, Özlem YilmazAbstract:Porphyromonas Gingivalis is a host-adapted oral pathogen associated with chronic periodontitis that successfully survives and persists in the oral epithelium. Recent studies have positively correlated periodontitis with increased risk and severity of oral squamous cell carcinoma (OSCC). Intriguingly, the presence of P. Gingivalis enhances tumorigenic properties independently of periodontitis and has therefore been proposed as a potential etiological agent for OSCC. However, the initial host molecular changes induced by P. Gingivalis infection which promote predisposition to cancerous transformation through EMT (epithelial-mesenchymal-transition), has never been studied in human primary cells which more closely mimic the physiological state of cells in vivo. In this study, we examine for the first time in primary oral epithelial cells (OECs) the expression and activation of key EMT mediators during long-term P. Gingivalis infection in vitro. We examined the inactive phosphorylated state of glycogen synthase kinase-3 beta (p-GSK3β) over 120 h P. Gingivalis infection and found p-GSK3β, an important EMT regulator, significantly increases over the course of infection (p < 0.01). Furthermore, we examined the expression of EMT-associated transcription factors, Slug, Snail, and Zeb1 and found significant increases (p < 0.01) over long-term P. Gingivalis infection in protein and mRNA expression. Additionally, the protein expression of mesenchymal intermediate filament, Vimentin, was substantially increased over 120 h of P. Gingivalis infection. Analysis of adhesion molecule E-cadherin showed a significant decrease (p < 0.05) in expression and a loss of membrane localization along with β-catenin in OECs. Matrix metalloproteinases (MMPs) 2, 7, and 9 are all markedly increased with long-term P. Gingivalis infection. Finally, migration of P. Gingivalis infected cells was evaluated using scratch assay in which primary OEC monolayers were wounded and treated with proliferation inhibitor, Mitomycin C. The cellular movement was determined by microscopy. Results displayed P. Gingivalis infection promoted cell migration which was slightly enhanced by co-infection with Fusobacterium nucleatum, another oral opportunistic pathogen. Therefore, this study demonstrates human primary OECs acquire initial molecular/cellular changes that are consistent with EMT induction during long-term infection by P. Gingivalis and provides a critically novel framework for future mechanistic studies.
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intrinsic apoptotic pathways of gingival epithelial cells modulated by porphyromonas Gingivalis
Cellular Microbiology, 2007Co-Authors: Song Mao, Özlem Yilmaz, Yoonsuk Park, Yoshiaki Hasegawa, Gena D Tribble, Chloe E James, Martin Handfield, Franci M Stavropoulos, Richard J LamontAbstract:Porphyromonas Gingivalis can inhibit chemically induced apoptosis in primary cultures of gingival epithelial cells through blocking activation of the effector caspase-3. The anti-apoptotic phenotype of P. Gingivalis is conserved across strains and does not depend on the presence of fimbriae, as fimbriae-deficient mutants and a naturally occurring non-fimbriated strain were able to impede apoptosis. To dissect the survival pathways modulated by P. Gingivalis, protein and gene expression of a number of components of apoptotic death pathways were investigated. P. Gingivalis infection of epithelial cells resulted in the phosphorylation of JAK1 and Stat3. Quantitative real-time reverse transcription polymerase chain reaction showed that expression of Survivin and Stat3 itself, targets of activated Stat3, were elevated in P. Gingivalis-infected cells. siRNA knockdown of JAK1, in combination with knockdown of Akt, abrogated the ability of P. Gingivalis to block apoptosis. In contrast, cIAP-1 and cIAP-2 were not differentially regulated at either the protein or mRNA levels by P. Gingivalis. One mechanism by which P. Gingivalis can block apoptotic pathways in gingival epithelial cells therefore is through manipulation of the JAK/Stat pathway that controls the intrinsic mitochondrial cell death pathways. Induction of a pro-survival phenotype may prevent programmed host cell death and aid survival of P. Gingivalis within gingival epithelial cells.
Eugene J Leys - One of the best experts on this subject based on the ideXlab platform.
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quantitative real time pcr for porphyromonas Gingivalis and total bacteria
Journal of Clinical Microbiology, 2000Co-Authors: Sharon R Lyons, Ann L Griffen, Eugene J LeysAbstract:Accurate quantitation of the number of cells of individual bacterial species in dental plaque samples is needed for understanding the bacterial etiology of periodontitis. Real-time PCR offers a sensitive, efficient, and reliable approach to quantitation. Using the TaqMan system we were able to determine both the amount of Porphyromonas Gingivalis and the total number of bacterial cells present in plaque samples. Using species-specific primers and a fluorescent probe, detection of DNA from serial dilutions of P. Gingivalis cells was linear over a large range of DNA concentrations (correlation coefficient = 0.96). No difference was observed between P. Gingivalis DNA alone and the same DNA mixed with DNA isolated from dental plaque, indicating that P. Gingivalis levels can be determined accurately from clinical samples. The total number of cells of all bacterial species was determined using universal primers and a fluorescent probe. Standard curves using four different bacterial species gave similar results (correlation coefficient = 0.86). Levels of both P. Gingivalis and total bacteria were determined from a series of human plaque samples. High levels of P. Gingivalis were observed in several of the samples from subjects with periodontitis and none of those from healthy subjects. Real-time quantitative PCR provided a sensitive and reliable method for quantitating P. Gingivalis. In addition, it allowed the determination of the total number of bacterial cells present in a complex sample so that the percentage of P. Gingivalis cells could be determined.
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prevalence of porphyromonas Gingivalis and periodontal health status
Journal of Clinical Microbiology, 1998Co-Authors: Ann L Griffen, Mitzi R Becker, Sharon R Lyons, Melvin L Moeschberger, Eugene J LeysAbstract:Periodontitis is a common, progressive disease that eventually affects the majority of the population. The local destruction of periodontitis is believed to result from a bacterial infection of the gingival sulcus, and several clinical studies have provided evidence to implicate Porphyromonas Gingivalis. If P. Gingivalis is a periodontal pathogen, it would be expected to be present in most subjects with disease and rarely detected in subjects with good periodontal health. However, in most previous studies, P. Gingivalis has not been detected in the majority of subjects with disease, and age-matched, periodontally healthy controls were not included for comparison. The purpose of the study reported here was to compare the prevalence of P. Gingivalis in a group with periodontitis to that of a group that is periodontally healthy. A comprehensive sampling strategy and a sensitive PCR assay were used to maximize the likelihood of detection. The target sequence for P. Gingivalis-specific amplification was the transcribed spacer region within the ribosomal operon. P. Gingivalis was detected in only 25% (46 of 181) of the healthy subjects but was detected in 79% (103 of 130) of the periodontitis group (P < 0.0001). The odds ratio for being infected with P. Gingivalis was 11.2 times greater in the periodontitis group than in the healthy group (95% confidence interval, 6.5 to 19.2). These data implicate P. Gingivalis in the pathogenesis of periodontitis and suggest that P. Gingivalis may not be a normal inhabitant of a periodontally healthy dentition.
Eric C Reynolds - One of the best experts on this subject based on the ideXlab platform.
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macrophage depletion abates porphyromonas Gingivalis induced alveolar bone resorption in mice
Journal of Immunology, 2014Co-Authors: Roselind S Lam, Neil M Obriensimpson, Jason C Lenzo, James A Holden, Gail C Brammar, Katrina A Walsh, J Mcnaughtan, Dennis K Rowler, Nico Van Rooijen, Eric C ReynoldsAbstract:The role of the macrophage in the immunopathology of periodontitis has not been well defined. In this study, we show that intraoral inoculation of mice with Porphyromonas Gingivalis resulted in infection, alveolar bone resorption, and a significant increase in F4/80(+) macrophages in gingival and submandibular lymph node tissues. Macrophage depletion using clodronate-liposomes resulted in a significant reduction in F4/80(+) macrophage infiltration of gingival and submandibular lymph node tissues and significantly (p < 0.01) less P. Gingivalis-induced bone resorption compared with controls in BALB/c and C57BL/6 mice. In both mouse strains, the P. Gingivalis-specific IgG Ab subclass and serum cytokine [IL-4, IL-10, IFN-γ, and IL-12 (p70)] responses were significantly (p < 0.01) lower in the macrophage-depleted groups. Macrophage depletion resulted in a significant reduction in the level of P. Gingivalis infection, and the level of P. Gingivalis infection was significantly correlated with the level of alveolar bone resorption. M1 macrophages (CD86(+)), rather than M2 macrophages (CD206(+)), were the dominant macrophage phenotype of the gingival infiltrate in response to P. Gingivalis infection. P. Gingivalis induced a significant (p < 0.01) increase in NO production and a small increase in urea concentration, as well as a significant increase in the secretion of IL-1β, IL-6, IL-10, IL-12 (p70), eotaxin, G-CSF, GM-CSF, macrophage chemoattractant protein-1, macrophage inflammatory protein-α and -β, and TNF-α in isolated murine macrophages. In conclusion, P. Gingivalis infection induced infiltration of functional/inflammatory M1 macrophages into gingival tissue and alveolar bone resorption. Macrophage depletion reduced P. Gingivalis infection and alveolar bone resorption by modulating the host immune response.