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Beyond Toll-Like Receptors: Porphyromonas gingivalis Induces IL-6, IL-8, and VCAM-1 Expression Through NOD-Mediated NF-κB and ERK Signaling Pathways in Periodontal Fibroblasts

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Abstract

To investigate whether oligomerization domains (NODs) are involved in Porphyromonas gingivalis-induced interleukin (IL)-6, IL-8, and vascular cell adhesion molecule (VCAM)-1 expression beyond Toll-like receptors (TLRs), we investigated the role of NOD1/2 in P. gingivalis-induced IL-6, IL-8, and VCAM-1 expression in human gingival fibroblasts (hGFs) and periodontal ligament cells (hPDLCs). The mechanism was explored by activation and silence of NODs, electrophoretic mobility shift assay (EMSA), and pathway blockade assays. Results showed that P. gingivalis could induce NOD1, NOD2, IL-6, IL-8, and VCAM-1 expression in hGFs and hPDLs at mRNA and protein levels. Activation of NOD1/2 by agonists could clearly upregulate the expression of these genes, while silence of NOD1/2 could remarkably attenuate them. EMSA and blockade of NF-κB and extracellular-signal-regulated kinase (ERK)1/2 pathway assays also verified that the two pathways were involved in NOD1/2-mediated IL-6, IL-8, and VCAM-1 expression. In conclusion, our findings demonstrated that P. gingivalis induced IL-6, IL-8, and VCAM-1 expression in hGFs and hPDLCs through NOD1/2-mediated NF-κB and ERK1/2 signaling pathways beyond TLRs.

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Acknowledgments

We thank Prof. Chenxiong Lai from the Kaohsiung Medical University in Taiwan for generously providing P. gingivalis W83. This study is supported by National Natural Science Foundation of China (no. 181271150).

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The authors declare there is no conflict of interests.

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Authors and Affiliations

  1. Department of Periodontology, Peking University School and Hospital of Stomatology, 22 Zhongguancun South Avenue, Haidian District, Beijing, 100081, China

    Jianru Liu & Xiangying Ouyang

  2. Peking University School and Hospital of Stomatology, 22 Zhongguancun South Avenue, Haidian District, Beijing, 100081, China

    Yixiang Wang

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  1. Jianru Liu
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Correspondence to Yixiang Wang or Xiangying Ouyang.

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Supplementary figure 1

Characterization of hPDLCs and hGFs. The fibroblasts were identified by immunohistochemistry of mesenchymal marker vimentin and epithelial markers cytokeratin (CK)-14. Both of hPDLCs and hGFs were typically spindle-shaped and were positive for the mesenchymal marker vimentin (C & D) and negative for the epithelial marker cytokeratin 14, CK (A & B). (JPEG 237 kb)

High resolution image (TIFF 3411 kb)

Supplementary figure 2

mRNA expression of TLR4 in P. gingivalis-challenged hPDLCs and hGFs. Cells were treated with P. gingivalis at indicated MOI for indicated time. Real-time PCR was applied to detect TLR4 expression. No significant change of its expression level was revealed before and after bacterial challenge (A & B). (JPEG 23 kb)

High resolution image (TIFF 1994 kb)

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Liu, J., Wang, Y. & Ouyang, X. Beyond Toll-Like Receptors: Porphyromonas gingivalis Induces IL-6, IL-8, and VCAM-1 Expression Through NOD-Mediated NF-κB and ERK Signaling Pathways in Periodontal Fibroblasts. Inflammation 37, 522–533 (2014). https://doi.org/10.1007/s10753-013-9766-0

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