scholarly journals Porphyromonas gingivalis Induction of MicroRNA-203 Expression Controls Suppressor of Cytokine Signaling 3 in Gingival Epithelial Cells

2011 ◽  
Vol 79 (7) ◽  
pp. 2632-2637 ◽  
Author(s):  
Catherine E. Moffatt ◽  
Richard J. Lamont
Keyword(s):  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Luciferase Assay ◽  
Cytokine Signaling ◽  
Mrna Levels ◽  
Suppressor Of Cytokine Signaling ◽  
Content Type ◽  
Downstream Target ◽  
Qrt Pcr ◽  
Gingival Epithelial Cells

ABSTRACTPorphyromonas gingivalisis a pathogen in severe periodontal disease. Able to exploit an intracellular lifestyle within primary gingival epithelial cells (GECs), a reservoir ofP. gingivaliscan persist within the gingival epithelia. This process is facilitated by manipulation of the host cell signal transduction cascades which can impact cell cycle, cell death, and cytokine responses. Using microarrays, we investigated the ability ofP. gingivalis33277 to regulate microRNA (miRNA) expression in GECs. One of several miRNAs differentially regulated by GECs in the presence ofP. gingivaliswas miRNA-203 (miR-203), which was upregulated 4-fold compared to uninfected controls. Differential regulation of miR-203 was confirmed by quantitative reverse transcription-PCR (qRT-PCR). Putative targets of miR-203, suppressor of cytokine signaling 3 (SOCS3) and SOCS6, were evaluated by qRT-PCR. SOCS3 and SOCS6 mRNA levels were reduced >5-fold and >2-fold, respectively, inP. gingivalis-infected GECs compared to controls. Silencing of miR-203 using a small interfering RNA construct reversed the inhibition of SOCS3 expression. A dual luciferase assay confirmed binding of miR-203 to the putative target binding site of the SOCS3 3′ untranslated region. Western blot analysis demonstrated that activation of signal transducer and activator of transcription 3 (Stat3), a downstream target of SOCS, was diminished following miR-203 silencing. This study shows that induction of miRNAs byP. gingivaliscan modulate important host signaling responses.

scholarly journals Noncanonical Activation of β-Catenin by Porphyromonas gingivalis

2015 ◽  
Vol 83 (8) ◽  
pp. 3195-3203 ◽  
Author(s):  
Yun Zhou ◽  
Maryta Sztukowska ◽  
Qian Wang ◽  
Hiroaki Inaba ◽  
Jan Potempa ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Immunofluorescence Microscopy ◽  
Proteolytic Processing ◽  
Luciferase Reporter ◽  
Cell Fractionation ◽  
Mrna Levels ◽  
Major Pathway ◽  
Content Type ◽  
Gingival Epithelial Cells

Porphyromonas gingivalisis an established pathogen in periodontal disease and an emerging pathogen in serious systemic conditions, including some forms of cancer. We investigated the effect ofP. gingivalison β-catenin signaling, a major pathway in the control of cell proliferation and tumorigenesis. Infection of gingival epithelial cells withP. gingivalisdid not influence the phosphorylation status of β-catenin but resulted in proteolytic processing. The use of mutants deficient in gingipain production, along with gingipain-specific inhibitors, revealed that gingipain proteolytic activity was required for β-catenin processing. The β-catenin destruction complex components Axin1, adenomatous polyposis coli (APC), and GSK3β were also proteolytically processed byP. gingivalisgingipains. Cell fractionation and Western blotting demonstrated that β-catenin fragments were translocated to the nucleus. The accumulation of β-catenin in the nucleus followingP. gingivalisinfection was confirmed by immunofluorescence microscopy. A luciferase reporter assay showed thatP. gingivalisincreased the activity of the β-catenin-dependent TCF/LEF promoter.P. gingivalisdid not increase Wnt3a mRNA levels, a finding consistent withP. gingivalis-induced proteolytic processing causing the increase in TCF/LEF promoter activity. Thus, our data indicate thatP. gingivaliscan induce the noncanonical activation of β-catenin and disassociation of the β-catenin destruction complex by gingipain-dependent proteolytic processing. β-Catenin activation in epithelial cells byP. gingivalismay contribute to a proliferative phenotype.

scholarly journals Interleukin-8 and Intercellular Adhesion Molecule 1 Regulation in Oral Epithelial Cells by Selected Periodontal Bacteria: Multiple Effects of Porphyromonas gingivalis via Antagonistic Mechanisms

2001 ◽  
Vol 69 (3) ◽  
pp. 1364-1372 ◽  
Author(s):  
George T.-J. Huang ◽  
Daniel Kim ◽  
Jonathan K.-H. Lee ◽  
Howard K. Kuramitsu ◽  
Susan Kinder Haake
Keyword(s):  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Adhesion Molecule ◽  
Intercellular Adhesion ◽  
Interleukin 8 ◽  
Intercellular Adhesion Molecule ◽  
Mrna Levels ◽  
Periodontal Pathogens ◽  
Intercellular Adhesion Molecule 1 ◽  
Gingival Epithelial Cells

ABSTRACT Interaction of bacteria with mucosal surfaces can modulate the production of proinflammatory cytokines and adhesion molecules produced by epithelial cells. Previously, we showed that expression of interleukin-8 (IL-8) and intercellular adhesion molecule 1 (ICAM-1) by gingival epithelial cells increases following interaction with several putative periodontal pathogens. In contrast, expression of IL-8 and ICAM-1 is reduced after Porphyromonas gingivalis ATCC 33277 challenge. In the present study, we investigated the mechanisms that govern the regulation of these two molecules in bacterially infected gingival epithelial cells. Experimental approaches included bacterial stimulation of gingival epithelial cells by either a brief challenge (1.5 to 2 h) or a continuous coculture throughout the incubation period. The kinetics of IL-8 and ICAM-1 expression following brief challenge were such that (i) secretion of IL-8 by gingival epithelial cells reached its peak 2 h following Fusobacterium nucleatum infection whereas it rapidly decreased within 2 h after P. gingivalis infection and remained decreased up to 30 h and (ii) IL-8 and ICAM-1 mRNA levels were up-regulated rapidly 2 to 4 h postinfection and then decreased to basal levels 8 to 20 h after infection with either Actinobacillus actinomycetemcomitans, F. nucleatum, or P. gingivalis. Attenuation of IL-8 secretion was facilitated by adherent P. gingivalis strains. The IL-8 secreted from epithelial cells after F. nucleatum stimulation could be down-regulated by subsequent infection with P. gingivalisor its culture supernatant. Although these results suggested that IL-8 attenuation at the protein level might be associated with P. gingivalis proteases, the Arg- and Lys-gingipain proteases did not appear to be solely responsible for IL-8 attenuation. In addition, while P. gingivalis up-regulated IL-8 mRNA expression, this effect was overridden when the bacteria were continuously cocultured with the epithelial cells. The IL-8 mRNA levels in epithelial cells following sequential challenge with P. gingivalis andF. nucleatum and vice versa were approximately identical and were lower than those following F. nucleatum challenge alone and higher than control levels or those following P. gingivalis challenge alone. Thus, together with the protease effect, P. gingivalis possesses a powerful strategy to ensure the down-regulation of IL-8 and ICAM-1.

scholarly journals Metronidazole-Treated Porphyromonas gingivalis Persisters Invade Human Gingival Epithelial Cells and Perturb Innate Responses

2020 ◽  
Vol 64 (6) ◽  
Author(s):  
Chuan Wang ◽  
Tianfan Cheng ◽  
Xuan Li ◽  
Lijian Jin
Keyword(s):  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Present Report ◽  
Well Being ◽  
Host Responses ◽  
Periodontal Health ◽  
Content Type ◽  
Gingival Epithelial Cells ◽  
Human Gingival Epithelial Cells ◽  
Lethal Dosage

ABSTRACT Periodontitis as a biofilm-associated inflammatory disease is highly prevalent worldwide. It severely affects oral health and yet closely links to systemic diseases like diabetes and cardiovascular disease. Porphyromonas gingivalis as a “keystone” periodontopathogen drives the shift of microbe-host symbiosis to dysbiosis and critically contributes to the pathogenesis of periodontitis. Persisters represent a tiny subset of biofilm-associated microbes highly tolerant to lethal treatment of antimicrobials, and, notably, metronidazole-tolerant P. gingivalis persisters have recently been identified by our group. This study further explored the interactive profiles of metronidazole-treated P. gingivalis persisters (M-PgPs) with human gingival epithelial cells (HGECs). P. gingivalis cells (ATCC 33277) at stationary phase were treated with a lethal dosage of metronidazole (100 μg/ml, 6 h) for generating M-PgPs. The interaction of M-PgPs with HGECs was assessed by microscopy, flow cytometry, cytokine profiling, and quantitative PCR (qPCR). We demonstrated that the overall morphology and ultracellular structure of M-PgPs remained unchanged. Importantly, M-PgPs maintained the capabilities to adhere to and invade HGECs. Moreover, M-PgPs significantly suppressed proinflammatory cytokine expression in HGECs at a level comparable to that seen with the untreated P. gingivalis cells, through the thermosensitive components. The present report reveals that P. gingivalis persisters induced by lethal treatment of antibiotics were able to maintain their capabilities to adhere to and invade human gingival epithelial cells and to perturb the innate host responses. Novel strategies and approaches need to be developed for tackling P. gingivalis and favorably modulating the dysregulated immunoinflammatory responses for oral/periodontal health and general well-being.

scholarly journals Porphyromonas gingivalis enhances FasL expression via up-regulation of NFκB-mediated gene transcription and induces apoptotic cell death in human gingival epithelial cells

Microbiology ◽  
2006 ◽  
Vol 152 (3) ◽  
pp. 797-806 ◽  
Author(s):  
Suzana Brozovic ◽  
Rashmita Sahoo ◽  
Shirish Barve ◽  
Hideki Shiba ◽  
Silvia Uriarte ◽  
...  
Keyword(s):  
Cell Death ◽  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Caspase 3 ◽  
Apoptotic Cell ◽  
Apoptotic Cell Death ◽  
Luciferase Assay ◽  
Caspase 8 ◽  
Gingival Epithelial Cells ◽  
Human Gingival Epithelial Cells

The interaction between epithelial cells and micro-organisms is often a crucial initiating event in infectious diseases. Infection with Porphyromonas gingivalis, a Gram-negative anaerobe, is strongly associated with severe periodontal disease. This bacterium possesses an array of virulence factors, some of which can induce apoptosis. The tumour necrosis factor (TNF) receptor family is involved in the regulation of cellular homeostasis, cell surface molecules involved in phagocytosis, Fas ligand (L) expression and activation of the caspase cascade resulting in DNA fragmentation and cell blebbing. The current study examined the role of nuclear factor-κB (NFκB) in FasL-mediated apoptotic cell death in primary human gingival epithelial cells (HGEC) induced by heat-killed P. gingivalis, probably through TLR signalling pathways. A marked up-regulation of TLR2 and Fas–FasL was detected in HGEC stimulated with P. gingivalis. Activation of NFκB by P. gingivalis in HGEC was demonstrated by an NFκB promoter luciferase assay as well as by phosphorylation of p65 as detected by Western blotting. Activation of cleaved caspase-3 and caspase-8 resulted in apoptotic cell death of HGEC. The survival proteins c-IAP-1/c-IAP-2 were decreased in HGEC exposed to P. gingivalis. HGEC apoptosis induced by P. gingivalis was inhibited by an anti-human FasL monoclonal antibody. Blockade of NFκB by helenalin resulted in down-regulation of FasL whereas a caspase-8 inhibitor did not decrease FasL. Taken together, these studies show that P. gingivalis can induce epithelial cell apoptosis through Fas–FasL up-regulation and activation of caspase-3 and caspase-8.

scholarly journals Identification of Porphyromonas gingivalis Genes Specifically Expressed in Human Gingival Epithelial Cells by Using Differential Display Reverse Transcription-PCR

2004 ◽  
Vol 72 (7) ◽  
pp. 3752-3758 ◽  
Author(s):  
Yoonsuk Park ◽  
Özlem Yilmaz ◽  
Il-Young Jung ◽  
Richard J. Lamont
Keyword(s):  
Epithelial Cells ◽  
Porphyromonas Gingivalis ◽  
Abc Transporter ◽  
Differential Display ◽  
Molecular Mechanisms ◽  
Gene Products ◽  
Reverse Transcription Pcr ◽  
Gingival Epithelial Cells ◽  
Insertional Inactivation ◽  
Human Gingival Epithelial Cells

ABSTRACT Porphyromonas gingivalis, one of the causative agents of adult periodontitis, can invade and survive within host epithelial cells. The molecular mechanisms by which P. gingivalis induces uptake and adapts to an intracellular environment are not fully understood. In this study, we have investigated the genetic responses of P. gingivalis internalized within human gingival epithelial cells (GECs) in order to identify factors involved in invasion and survival. We compared the differential display of arbitrarily PCR-amplified gene transcripts in P. gingivalis recovered from GECs with the display of transcripts in P. gingivalis control cultures. Over 20 potential differentially expressed transcripts were identified. Among these, pepO, encoding an endopeptidase, and genes encoding an ATP-binding cassette (ABC) transporter and a cation-transporting ATPase were upregulated in GECs. To investigate the functionality of these gene products, mutants were generated by insertional inactivation. Compared to the parental strain, mutants of each gene showed a significant reduction in their invasion capabilities. In addition, GEC cytoskeletal responses to the mutants were distinct from those induced by the parent. In contrast, adhesion of the mutant strains to GECs was not affected by lack of expression of the gene products. These results suggest that PepO, a cation-transporting ATPase, and an ABC transporter are required for the intracellular lifestyle of P. gingivalis.

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