Marie-Jeanne Staquet

University of Lyon, Lyons, Rhône-Alpes, France

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Publications (38)36.24 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Previous studies have suggested that odontoblasts sense Gram-positive bacteria components through Toll-like receptor 2 (TLR2) and trigger dental pulp immunity by producing specific pro-inflammatory cytokines and chemokines. However, most of the determinants of the inflammatory odontoblast response to these pathogens remain unknown. Objectives: The aim of this study was to identify inflammation-related pathways activated in vivo in bacteria-challenged inflamed human dental pulps and in vitro in odontoblast-like cells activated by two Gram-positive bacteria-derived TLR2 specific agonists. Methods: Gene expression of cyclooxygenase-2 (COX2), the NADPH oxidase subunits NOX1, NOX2, NOX3, NOX4, NOX5, DUOX1 and DUOX2, nitric oxide synthases 1, 2 and 3 (NOS1, NOS2 and NOS3) and heme oxygenase-1 (HMOX-1) was assessed by real-time PCR in healthy and bacteria-challenged inflamed dental pulps and in odontoblast-like cells stimulated with lipoteichoic acid (LTA), a cell wall component, or Pam2CSK4, a synthetic diacylated lipopeptide. NOS2 protein was localised in pulp samples by immunohistochemistry. NOS2 (inducible NOS) production was determined in Pam2CSK4-stimulated cells by western-blot. NOS activity and NO production were measured with Griess colorimetric assay in cells and culture supernatants, respectively. Results: Gene expression of COX2, the NADPH oxidase subunits NOX1, NOX2 and NOX4, NOS1, NOS2, NOS3, and HMOX1 were significantly increased in inflamed pulps compared to healthy ones. NOS2 was the most up-regulated gene. NOS2 protein was immunolocalized in odontoblasts beneath caries lesions but not in healthy samples. LTA stimulation of human odontoblast-like cells induced a low but significant increase of COX2, NOX1, NOX2, NOX5, DUOX1, NOS1, NOS2, NOS3 gene expression. Pam2CSK4 induced a strong increase of the same genes and of HMOX1. NOS2 protein production, NOS activity and NO release were clearly increased by Pam2CSK4. Conclusion: These results suggest that odontoblasts might contribute to the inflammatory response triggered in the human dental pulp by Gram-positive bacteria entering dentin during the caries process.
    Annual Meeting of the IADR Continental European Division 2013; 09/2013
  • [Show abstract] [Hide abstract]
    ABSTRACT: Previous studies have suggested that odontoblasts sense gram-positive bacteria components through Toll-like receptor 2 (TLR2) and trigger dental pulp immunity by producing proinflammatory cytokines. Currently, the factors that modulate odontoblast TLR2 activation are unknown. Our aim was to investigate lipopolysaccharide-binding protein (LBP) effects on the TLR2-mediated odontoblast response. Human odontoblast-like cells were stimulated with lipoteichoic acid (LTA) (a TLR2 ligand), LBP, CD14 (a TLR2 cofactor), or various combinations of LTA/LBP, LTA/CD14, or LTA/CD14/LBP. CXCL8, IL6, and TLR2 gene expression was assessed by real-time polymerase chain reaction. CXCL8 and interleukin (IL)-6 production was determined by enzyme-linked immunosorbent assay in culture supernatants of cells stimulated with LTA, LTA/CD14, or LTA/CD14/LBP. LBP effects on nuclear factor kappa B (NF-κB), p38, JNK, ERK, STAT3, and p70S6 signaling pathways were determined in LTA-stimulated odontoblast-like cells with a multiplex biometric immunoassay. LBP effects were compared with specific inhibitors of these signaling pathways. LBP transcript and protein were investigated in vivo in healthy and inflamed dental pulps by real-time polymerase chain reaction and immunohistochemistry. Activation of CXCL8, IL6, and TLR2 gene expression and CXCL8 and IL-6 secretion in LTA- and LTA/CD14-stimulated odontoblast-like cells was significantly decreased by LBP. LBP inhibited NF-κB and p38 signaling pathways in LTA-stimulated cells in a similar way to NF-κB and p38 inhibitors. LBP transcript and protein were detected in vivo in inflamed dental pulps but not in healthy ones. These results demonstrate that LBP reduces TLR2-dependent production of inflammatory cytokines by odontoblast-like cells. We suggest that in this way it could modulate host defense in human dental pulp.
    Journal of endodontics 08/2013; 39(8):1008-14. · 2.95 Impact Factor
  • 11th International Conference on Tooth Morphogenesis and Differentiation, La Londe-les-Maures, France; 05/2013
  • [Show abstract] [Hide abstract]
    ABSTRACT: Previous studies have suggested that odontoblasts sense components from the Gram-positive bacteria cell wall through Toll-like receptor 2 (TLR2) and trigger dental pulp innate immunity by producing specific pro-inflammatory cytokines and chemokines. Whether they synthesize antimicrobial agents to directly combat intradentinal pathogens remains poorly understood. Objectives: The purpose of this study was to determine whether odontoblasts are able to produce nitric oxide (NO) upon TLR2 engagement. Methods: Odontoblast-like cells differentiated from human dental pulp explants were stimulated with the TLR2 agonists lipoteichoic acid (LTA), a cell wall component of Gram-positive bacteria, and Pam2CSK4, a diacylated lipoprotein synthetic analog. Expression of genes coding for nitric oxide synthases 1, 2 and 3 (NOS1, NOS2 and NOS3) was assessed by real-time polymerase chain reaction (PCR) analysis in LTA- and Pam2CSK4-stimulated odontoblast-like cells. Production of NOS2 (inducible NOS) was determined in Pam2CSK4-stimulated cells by western-blot. Cellular NOS activity and NO production in culture supernatants were measured with Griess colorimetric assay. NOS2 expression was analyzed by PCR in healthy and bacteria-challenged inflamed dental pulps. NOS2 protein was localised in pulp samples by immunohistochemistry. Results: NOS1, NOS2 and NOS3 expression was increased in odontoblast-like cells stimulated with LTA or Pam2CSK4. NOS2 was the most up-regulated gene. Pam2CSK4 was a strongest stimulator than LTA. NOS2 protein production, NOS activity and NO release were strongly increased by Pam2CSK4. In vivo, NOS2 and, to a lesser extent, NOS1 and NOS3 were up-regulated in inflamed dental pulps compared with healthy ones. NOS2 protein was immunolocalized in odontoblasts beneath carious lesion but not at some distance from the lesion or in healthy samples. Conclusion: These results demonstrate that human odontoblast-like cells are able to produce NO upon TLR2 engagement and suggest that these cells might in this way be involved in the bactericidal response to dentin-invading Gram-positive bacteria.
    IADR/AADR/CADR General Session and Exhibition 2013; 03/2013
  • 91st General Session of the International Association for Dental Research, Seattle, USA; 03/2013
  • [Show abstract] [Hide abstract]
    ABSTRACT: Previous studies have suggested that human odontoblasts are involved in the dental pulp immune and inflammatory response to oral pathogens that invade dentin during the caries process. However, how odontoblasts regulate the early pulp response to Gram-positive bacteria, which predominate in shallow and moderate dentin caries, is not completely understood. We previously showed that activation of Toll-like receptor (TLR)2, a pattern recognition molecule activated by lipoteichoic acid (LTA) from Gram-positive bacteria, triggers TLR2 up-regulation, NF-κB nuclear translocation, and production of pro-inflammatory chemokines and cytokines, including CXCL8 and IL-6, by odontoblasts in vitro. Objective: To determine whether odontoblast activation by LTA is modulated by the lipopolysaccharide-binding protein (LBP), an acute-phase protein over-expressed in inflamed pulps which has been shown to interact with LTA. Methods: Human teeth were collected with the informed consent of the patients, in accordance with the Declaration of Helsinki and following a protocol approved by the local ethics committee. Odontoblast-like cells were differentiated from pulp explant cultures derived from non-erupted healthy third molars. Cells were stimulated for 4 h with various concentrations of LTA, alone or in association with LBP. Gene expression of the pro-inflammatory chemokine CXCL8, the cytokine interleukin-6 (IL-6), and the LTA receptor TLR2 was assessed using real-time PCR. Results were expressed as mean values SD and statistical analysis was determined with Student's t test. Results: CXCL8, IL-6 and TLR2 genes were significantly up-regulated by LTA, but this increase was reduced when LBP was added together with LTA in the culture medium. Conclusion: These data demonstrate that LBP negatively regulates TLR2 signalling induced by LTA in human odontoblasts and might play a role in protecting human dental pulp from excessive immune response to cariogenic Gram-positive bacteria. Supported by University Lyon 1, CNRS, IFRO and Rgion Rhone-Alpes.
    45th Meeting of the Continental European Division of the International Association of Dental Research (CED-IADR) with the Scandinavian Division (NOF) 2011; 09/2011
  • 10th World Congress on Inflammation, Paris, France; 06/2011
  • TOLL2011: Decoding Innate Immunity, Riva del Garda, Italy; 05/2011
  • [Show abstract] [Hide abstract]
    ABSTRACT: Recent studies have suggested that odontoblasts are involved in the dental pulp immune response to oral pathogens that invade human dentin during the caries process. How odontoblasts regulate the early inflammatory and immune pulp response to Gram-positive bacteria, which predominate in shallow and moderate dentin caries, is still poorly understood. In this study, we investigated the production of pro- and anti-inflammatory cytokines by odontoblast-like cells upon engagement of Toll-like receptor (TLR) 2, a pattern recognition molecule activated by Gram-positive bacteria components. We used a highly sensitive Milliplex(®) kit for detecting cytokines released by cells stimulated with lipoteichoic acid (LTA), a cell wall component of Gram-positive bacteria, or with the potent TLR2 synthetic agonist Pam2CSK4. We found that odontoblasts produce the pro-inflammatory cytokines interleukin (IL)-6 and CXCL8, as well as the immunosuppressive cytokine IL-10 in response to TLR2 agonists. GM-CSF, IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-7, IL-12(p70), IL-13 and TNF-α were not detected. These data indicate that TLR2 activation in human odontoblasts selectively induces production of mediators known to influence positively or negatively inflammatory and immune responses in pathogen-challenged tissues. We suggest that these molecules might be important in regulating the fine tuning of the pulp response to Gram-positive bacteria which enter dentin during the caries process.
    Immunobiology 04/2011; 216(4):513-7. · 2.81 Impact Factor
  • Conference Paper: TLRs in pulp defence
    Tissue Injury and Pulp Regeneration Meeting, Geneva, Switzerland; 07/2010
  • 88th General Session of the International Association for Dental Research, Barcelona, Spain; 07/2010
  • 88th General Session of the International Association for Dental Research, Barcelona, Spain; 07/2010
  • Keystone Symposia Conference – Innate Immunity: mechanisms linking with adaptive immunity, Dublin, Ireland; 06/2010
  • Les Cahiers de l’Association Dentaire Française. 01/2010; 26:46-52.
  • Integrative Post-Genomics Meeting 2009, Lyon, France; 11/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Human odontoblasts trigger immune response s to oral bacteria that invade dental tissues during the caries process. To date, their ability to regulate the expression of the nucleotide-binding domain leucine-rich repeat containing receptor NOD2 when challenged by Gram-positive bacteria is unknown. In this study, we investigated NOD2 expression in healthy and inflamed human dental pulps challenged by bacteria, and in cultured odontoblast-like cells stimulated with lipoteichoic acid (LTA), a Toll-like receptor (TLR) 2 agonist which is specific for Gram-positive bacteria. We found that NOD2 gene expression was significantly up-regulated in pulps with acute inflammation compared to healthy ones. In vitro, LTA augmented NOD2 gene expression and protein level in odontoblast-like cells. The increase was more pronounced in odontoblast-like cells compared to dental pulp fibroblasts. Blocking experiments in odontoblast-like cells with anti-TLR2 antibody strongly reduced the NOD2 gene expression increase, whereas stimulation with the synthetic TLR2 ligand Pam(2)CSK(4) confirmed NOD2 gene up-regulation following TLR2 engagement. These data suggest that NOD2 up-regulation is part of the odontoblast immune response to Gram-positive bacteria and might be important in protecting human dental pulp from the deleterious effects of cariogenic pathogens.
    Innate Immunity 10/2009; 17(1):29-34. · 2.46 Impact Factor
  • 44th Annual Meeting of the International Association for Dental Research, Continental European Division, Munich, Germany; 09/2009
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    ABSTRACT: Map-1B belongs to the family of proteins that govern the dynamic state and organization of microtubules within cells. MAP-1B is a microtubule-associated protein highly expressed during the development of the nervous system. Its expression, regulated by the fragile X mental retardation protein (FMRP), is essential to stabilize microtubules during the elongation of dendrites and neurites. Other microtubules-associated molecules such as tau or MAP2 seem to act similarly. The aim of this work was to identify the MAP-1B expression in in vitro and in vivo human odontoblasts during development and carious processes. The expression of MAP2 and tau was also studied. In cultured cells, MAP-1B expression was analyzed by real-time polymerase chain reaction, flow cytometry, and Western blot. Its distribution was visualized by in situ hybridization and immunochemistry both in vitro and in vivo. The expression of FMRP, MAP2, and tau was identified by real-time polymerase chain reaction and immunochemistry. MAP-1B is specifically expressed in odontoblasts from adult third molars as well as incisor germs from human embryos. In adult carious teeth, it is also expressed in newly differentiated dentin-forming cells. In vitro, MAP-1B expression is related to the differentiation state of odontoblasts. MAP-1B clearly underlines the cellular architecture of cell bodies and processes of differentiated cells. FMRP, MAP2, and tau are also detected in vivo. On the basis of these data, MAP-1B could be considered as a new protein involved in the terminal differentiation of odontoblasts.
    Journal of endodontics 08/2009; 35(7):992-6. · 2.95 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Odontoblasts, dental pulp fibroblasts and immature dendritic cells (DCs) have been involved in the human dental pulp immune response to oral pathogens that invade dentine during the caries process. How they regulate the inflammatory response to Gram-positive bacteria remains nevertheless largely unknown. In this study we investigated the production of the pro-inflammatory cytokines tumour necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and interleukin-8 (CXCL8) in these three cell types upon stimulation with lipoteichoic acid (LTA), a cell wall component of Gram-positive bacteria that activates the pattern recognition molecule Toll-like receptor 2 (TLR2). We observed that TNF-alpha gene expression was up-regulated in all LTA-stimulated cell types. IL-1beta gene expression was not or barely detectable in odontoblast-like cells and pulp fibroblasts when stimulated or not, but was expressed in immature DCs and increased upon stimulation. TNF-alpha and IL-1beta proteins were detected in DC culture supernatants but not in odontoblast-like cell and pulp fibroblast ones. CXCL8 gene and protein were clearly expressed and increased in the three cell types upon LTA stimulation. These data indicate that LTA-dependent TLR2 activation in odontoblasts and pulp fibroblasts, in contrast to immature DCs, does not lead to significant TNF-alpha and IL-1beta production, but that all three cell types influence the pulp inflammatory/immune response through CXCL8 synthesis and secretion.
    Immunobiology 03/2009; 215(1):53-9. · 2.81 Impact Factor
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    ABSTRACT: Recent studies have demonstrated that human dental pulp cells sense pathogens and elicit innate and/or adaptive immunity. Particular attention has been paid to odontoblasts that are situated at the pulp-dentin interface and constitute the first line of defense to cariogenic bacteria entering dentin after enamel disruption. In this review, recent in vitro and in vivo data suggesting that odontoblasts initiate immune/inflammatory events within the dental pulp in response to cariogenic bacteria are discussed. These data include sensing of pathogens by Toll-like receptors (TLRs), production of chemokines upon cell stimulation with microbial by-products and induction of dendritic cell migration. Additional results presented here reveal that all TLR genes are expressed in the healthy human dental pulp that is thus well equipped to combat pathogens entering the tissue. Seventeen chemokine genes including CXCL12, CCL2, CXCL9, CX3CL1, CCL8, CXCL10, CCL16, CCL5, CXCL2, CCL4, CXCL11 and CCL3, and 9 chemokine receptor genes including CXCR4, CCR1, CCR5, CX3CR1, CCR10 and CXCR3, are also expressed in pulp. TLR2, CCL2 and CXCL1 are upregulated in odontoblasts both under caries lesions and upon stimulation with pathogen by-products. These molecules thus appear as preferential targets for the design of therapeutic agents able to reduce the immune/inflammatory response to cariogenic bacteria and favor pulp healing.
    Journal of Experimental Zoology Part B Molecular and Developmental Evolution 01/2009; 312B(5):425-36. · 2.12 Impact Factor

Publication Stats

137 Citations
36.24 Total Impact Points

Institutions

  • 2007–2013
    • University of Lyon
      Lyons, Rhône-Alpes, France
  • 2011
    • Ecole normale supérieure de Lyon
      Lyons, Rhône-Alpes, France
  • 2009–2011
    • Institut de Génomique Fonctionnelle de Lyon
      Lyons, Rhône-Alpes, France
    • French National Institute for Agricultural Research
      • Institut de Génomique Fonctionnelle de Lyon (IGFL)
      Paris, Ile-de-France, France
    • Hospices Civils de Lyon
      Lyons, Rhône-Alpes, France
  • 2006
    • French Institute of Health and Medical Research
      Lutetia Parisorum, Île-de-France, France