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Neutrophil extracellular traps aggravate periodontitis by disturbing regulatory T-cell differentiation

Genes & Immunity volume 26, pages 475–485 (2025)Cite this article

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Abstract

Excessive neutrophil extracellular traps (NETs) induce an intense inflammatory response in periodontitis. Recently, Tregs were shown to be essential for attenuating inflammation-driven bone resorption. However, the regulation of Tregs differentiation by NETs in periodontitis is still unclear and needs further investigation. In this study, a murine experimental periodontitis model was established either without or with NETs depletion via DNase I. Firstly, we revealed that NETs accumulated significantly in both periodontal tissues and sera of mice models with periodontitis, while the depletion of NETs alleviated alveolar bone resorption. Moreover, RNA sequencing and bioinformatics analysis revealed that NETs depletion regulated the immune response of gingival tissue, especially affecting T-cell differentiation, and identified potential regulatory pathways. Subsequently, we verified that inhibition of NETs promoted the infiltration of Tregs and increased expression levels of IL-10 and TGF-β in periodontal tissue. Furthermore, in vitro studies demonstrated that NETs produced by P. g-LPS-stimulated neutrophils impeded the differentiation of co-cultured naive CD4+ T cells into Tregs, which could be restored by DNase I-mediated digestion of NETs. In conclusion, excessive NETs could exacerbate alveolar bone resorption in periodontitis by interfering with the differentiation of Tregs, and DNase I provides a novel targeted strategy for the immunotherapy of periodontitis.

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Fig. 1: Evidence of NETs in lesions of experimental periodontitis and DNase I treatment inhibits NETs formation.
Fig. 2: DNase I treatment reduced bone loss in experimental periodontitis.
Fig. 3: RNA-seq of gingival tissues in experimental periodontitis.
Fig. 4: DNase I treatment increases the number of Tregs in periodontal lesions.
Fig. 5: LPS stimulated neutrophils to form NETs.
Fig. 6

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Data availability

All data analyzed during this study are included in the published article as well as the supplemental files online. Further enquiries are available by contacting the corresponding author upon reasonable request.

References

  1. Hajishengallis G, Chavakis T. Local and systemic mechanisms linking periodontal disease and inflammatory comorbidities. Nat Rev Immunol. 2021;21:426–40.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Silva LM, Brenchley L, Moutsopoulos NM. Primary immunodeficiencies reveal the essential role of tissue neutrophils in periodontitis. Immunol Rev. 2019;287:226–35.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Khoury W, Glogauer J, Tenenbaum HC, Glogauer M. Oral inflammatory load: Neutrophils as oral health biomarkers. J Periodontal Res. 2020;55:594–601.

    Article  CAS  PubMed  Google Scholar 

  4. Hajishengallis G. New developments in neutrophil biology and periodontitis. Periodontology. 2020;82:78–92.

    Article  Google Scholar 

  5. Thiam HR, Wong SL, Wagner DD, Waterman CM. Cellular mechanisms of NETosis. Annu Rev Cell Dev Biol. 2020;36:191–218.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Brinkmann V, Reichard U, Goosmann C, Fauler B, Uhlemann Y, Weiss DS, et al. Neutrophil extracellular traps kill bacteria. Science. 2004;303:1532–5.

    Article  CAS  PubMed  Google Scholar 

  7. Souza FW, Miao EA. Neutrophils only die twice. Sci Adv. 2023;9:eadm8715.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Ebersole JL, Dawson D 3rd, Emecen-Huja P, Nagarajan R, Howard K, Grady ME, et al. The periodontal war: microbes and immunity. Periodontology. 2017;75:52–115.

    Article  Google Scholar 

  9. Chapple ILC, Hirschfeld J, Kantarci A, Wilensky A, Shapira L. The role of the host-neutrophil biology. Periodontology. 2023;00:1–47.

  10. Silva LM, Doyle AD, Greenwell-Wild T, Dutzan N, Tran CL, Abusleme L, et al. Fibrin is a critical regulator of neutrophil effector function at the oral mucosal barrier. Science. 2021;374:eabl5450.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Bonilha CS, Veras FP, de Queiroz Cunha F. NET-targeted therapy: effects, limitations, and potential strategies to enhance treatment efficacy. Trends Pharm Sci. 2023;44:622–34.

    Article  CAS  PubMed  Google Scholar 

  12. O’Neil LJ, Barrera-Vargas A, Sandoval-Heglund D, Merayo-Chalico J, Aguirre-Aguilar E, Aponte AM, et al. Neutrophil-mediated carbamylation promotes articular damage in rheumatoid arthritis. Sci Adv. 2020;6:eabd2688.

  13. Xia Y, Cheng T, Zhang C, Zhou M, Hu Z, Kang F, et al. Human bone marrow mesenchymal stem cell-derived extracellular vesicles restore Th17/Treg homeostasis in periodontitis via miR-1246. FASEB J. 2023;37:e23226.

    Article  CAS  PubMed  Google Scholar 

  14. Deng J, Lu C, Zhao Q, Chen K, Ma S, Li Z. The Th17/Treg cell balance: crosstalk among the immune system, bone and microbes in periodontitis. J Periodontal Res. 2022;57:246–55.

    Article  CAS  PubMed  Google Scholar 

  15. Miron RJ, Bohner M, Zhang Y, Bosshardt DD. Osteoinduction and osteoimmunology: emerging concepts. Periodontology. 2024;94:9–26.

  16. Kim TS, Silva LM, Theofilou VI, Greenwell-Wild T, Li L, Williams DW, et al. Neutrophil extracellular traps and extracellular histones potentiate IL-17 inflammation in periodontitis. J Exp Med. 2023;220:1–15.

    Article  Google Scholar 

  17. Wang H, Zhang H, Wang Y, Brown ZJ, Xia Y, Huang Z, et al. Regulatory T-cell and neutrophil extracellular trap interaction contributes to carcinogenesis in non-alcoholic steatohepatitis. J Hepatol. 2021;75:1271–83.

    Article  CAS  PubMed  Google Scholar 

  18. Lambert S, Hambro CA, Johnston A, Stuart PE, Tsoi LC, Nair RP, et al. Neutrophil extracellular traps induce human Th17 cells: effect of psoriasis-associated TRAF3IP2 genotype. J Investig Dermatol Symp Proc. 2019;139:1245–53.

    Article  CAS  Google Scholar 

  19. Mikami N, Kawakami R, Chen KY, Sugimoto A, Ohkura N, Sakaguchi S. Epigenetic conversion of conventional T cells into regulatory T cells by CD28 signal deprivation. Proc Natl Acad Sci USA. 2020;117:12258–68.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Marchesan J, Girnary MS, Jing L, Miao MZ, Zhang S, Sun L, et al. An experimental murine model to study periodontitis. Nat Protoc. 2018;13:2247–67.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Zhou B, Zhang M, Ma H, Wang Y, Qiu J, Liu Y, et al. Distinct palmitoylation of Foxp3 regulates the function of regulatory T cells via palmitoyltransferases. Cell Mol Immunol. 2024;21:787–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Li MO, Rudensky AY. T cell receptor signalling in the control of regulatory T cell differentiation and function. Nat Rev Immunol. 2016;16:220–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Wan MC, Jiao K, Zhu YN, Wan QQ, Zhang YP, Niu LZ. et al. Bacteria-mediated resistance of neutrophil extracellular traps to enzymatic degradation drives the formation of dental calculi. Nat Biomed Eng. 2024;8:1503

    Article  PubMed  Google Scholar 

  24. Liu B, Deng Y, Duan Z, Chu C, Wang X, Yang C, et al. Neutrophil extracellular traps promote intestinal barrier dysfunction by regulating macrophage polarization during trauma/hemorrhagic shock via the TGF-β signaling pathway. Cell Signal. 2024;113:110941.

    Article  CAS  PubMed  Google Scholar 

  25. Chen W, Chen H, Yang ZT, Mao EQ, Chen Y, Chen EZ. Free fatty acids-induced neutrophil extracellular traps lead to dendritic cells activation and T cell differentiation in acute lung injury. Aging. 2021;13:26148–60.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Hajishengallis G. Illuminating the oral microbiome and its host interactions: animal models of disease. FEMS Microbiol Rev. 2023;47:fuad018

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Xiao E, Mattos M, Vieira GHA, Chen S, Corrêa JD, Wu Y, et al. Diabetes enhances IL-17 expression and alters the oral microbiome to increase its pathogenicity. Cell Host Microbe. 2017;22:120–8.e4.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Kimura S, Nagai A, Onitsuka T, Koga T, Fujiwara T, Kaya H, et al. Induction of experimental periodontitis in mice with Porphyromonas gingivalis-adhered ligatures. J Periodontol. 2000;71:1167–73.

    Article  CAS  PubMed  Google Scholar 

  29. Tsukasaki M, Takayanagi H. Osteoimmunology: evolving concepts in bone–immune interactions in health and disease. Nat Rev Immunol. 2019;19:626–42.

    Article  CAS  PubMed  Google Scholar 

  30. Jia L, Jiang Y, Wu L, Fu J, Du J, Luo Z, et al. Porphyromonas gingivalis aggravates colitis via a gut microbiota-linoleic acid metabolism-Th17/Treg cell balance axis. Nat Commun. 2024;15:1617.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Oliveira SR, de Arruda JAA, Schneider AH, Bemquerer LM, de Souza RMS, Barbim P, et al. Neutrophil extracellular traps in rheumatoid arthritis and periodontitis: Contribution of PADI4 gene polymorphisms. J Clin Periodontol. 2023;51:e13921.

  32. Onuora S. NETs implicated in periodontitis-associated bone loss. Nat Rev Rheumatol. 2023;19:463.

    Article  CAS  PubMed  Google Scholar 

  33. Schneider AH, Taira TM, Públio GA, da Silva Prado D, Donate Yabuta PB, Dos Santos JC, et al. Neutrophil extracellular traps mediate bone erosion in rheumatoid arthritis by enhancing RANKL-induced osteoclastogenesis. Br J Pharm. 2024;181:429–46.

    Article  CAS  Google Scholar 

  34. Guilherme Neto JL, Rodrigues Venturini LG, Schneider AH, Taira TM, Duffles Rodrigues LF, Veras FP, et al. Neutrophil extracellular traps aggravate apical periodontitis by stimulating osteoclast formation. J Endod. 2023;49:1514–21.

    Article  PubMed  Google Scholar 

  35. Zou H, Zhou N, Huang Y, Luo A, Sun J. Phenotypes, roles, and modulation of regulatory lymphocytes in periodontitis and its associated systemic diseases. J Leukoc Biol. 2022;111:451–67.

    Article  CAS  PubMed  Google Scholar 

  36. Hascoët E, Blanchard F, Blin-Wakkach C, Guicheux J, Lesclous P, Cloitre A. New insights into inflammatory osteoclast precursors as therapeutic targets for rheumatoid arthritis and periodontitis. Bone Res. 2023;11:26.

    Article  PubMed  PubMed Central  Google Scholar 

  37. Liu Z, Chen X, Zhang Z, Zhang X, Saunders L, Zhou Y, et al. Nanofibrous spongy microspheres to distinctly release miRNA and growth factors to enrich regulatory T cells and rescue periodontal bone loss. ACS Nano. 2018;12:9785–99.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Ha DY, Jung JS, Choi GH, Ji S. Polarization of human gingival fibroblasts by Th1-, Th2-, Th17-, and Treg-derived cytokines. J Periodontal Res. 2022;57:487–501.

    Article  CAS  PubMed  Google Scholar 

  39. Tsukasaki M, Komatsu N, Nagashima K, Nitta T, Pluemsakunthai W, Shukunami C, et al. Host defense against oral microbiota by bone-damaging T cells. Nat Commun. 2018;9:701.

    Article  PubMed  PubMed Central  Google Scholar 

  40. Liu, Hayashi M, Ohsugi Y, Katagiri S, Akira S, Iwata T, et al. The IL-33/ST2 axis is protective against acute inflammation during the course of periodontitis. Nat Commun. 2024;15:2707.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Qin X, Liu JY, Wang T, Pashley DH, Al-Hashim AH, Abdelsayed R, et al. Role of indoleamine 2,3-dioxygenase in an inflammatory model of murine gingiva. J Periodontal Res. 2017;52:107–13.

    Article  CAS  PubMed  Google Scholar 

  42. Breedveld A, Groot Kormelink T, van Egmond M, de Jong EC. Granulocytes as modulators of dendritic cell function. J Leukoc Biol. 2017;102:1003–16.

    Article  CAS  PubMed  Google Scholar 

  43. Wilson AS, Randall KL, Pettitt JA, Ellyard JI, Blumenthal A, Enders A, et al. Neutrophil extracellular traps and their histones promote Th17 cell differentiation directly via TLR2. Nat Commun. 2022;13:1–12.

    Article  Google Scholar 

  44. Esberg A, Kindstedt E, Isehed C, Lindquist S, Holmlund A, Lundberg P. LIGHT protein: A novel gingival crevicular fluid biomarker associated with increased probing depth after periodontal surgery. J Clin Periodontol. 2024;51:852–62.

    Article  CAS  PubMed  Google Scholar 

  45. Minns D, Smith KJ, Alessandrini V, Hardisty G, Melrose L, Jackson-Jones L, et al. The neutrophil antimicrobial peptide cathelicidin promotes Th17 differentiation. Nat Commun. 2021;12:1285.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Apolinário Vieira GH, Aparecida Rivas AC, Figueiredo Costa K, Ferreira Oliveira LF, Tanaka Suzuki K, Reis Messora M, et al. Specific inhibition of IL-6 receptor attenuates inflammatory bone loss in experimental periodontitis. J Periodontol. 2021;92:1460–9.

    Article  PubMed  Google Scholar 

  47. Piao W, Lee ZL, Zapas G, Wu L, Jewell CM, Abdi R. et al. Regulatory T cell and endothelial cell crosstalk. Nat Rev Immunol. 2025;25:588–607.

    Article  CAS  PubMed  Google Scholar 

  48. Ohl L, Mohaupt M, Czeloth N, Hintzen G, Kiafard Z, Zwirner J, et al. CCR7 governs skin dendritic cell migration under inflammatory and steady-state conditions. Immunity. 2004;21:279–88.

    Article  CAS  PubMed  Google Scholar 

  49. Davalos-Misslitz AC, Rieckenberg J, Willenzon S, Worbs T, Kremmer E, Bernhardt G, et al. Generalized multi-organ autoimmunity in CCR7-deficient mice. Eur J Immunol. 2007;37:613–22.

    Article  CAS  PubMed  Google Scholar 

  50. Chen Y, Liang R, Shi X, Shen R, Liu L, Liu Y, et al. Targeting kinase ITK treats autoimmune arthritis via orchestrating T cell differentiation and function. Biomed Pharmacother. 2023;169:115886.

    Article  CAS  PubMed  Google Scholar 

  51. Stakos D, Skendros P, Konstantinides S, Ritis K. Traps N’ Clots: NET-mediated thrombosis and related diseases. Thromb Haemost. 2020;120:373–83.

    Article  PubMed  Google Scholar 

  52. Hu Y, Wang H, Liu Y. NETosis: sculpting tumor metastasis and immunotherapy. Immunol Rev. 2024;321:263–79.

    Article  CAS  PubMed  Google Scholar 

  53. Jain S, Darveau RP. Contribution of Porphyromonas gingivalis lipopolysaccharide to periodontitis. Periodontology. 2010;54:53–70.

    Article  Google Scholar 

  54. Herath TD, Darveau RP, Seneviratne CJ, Wang CY, Wang Y, Jin L. Tetra- and penta-acylated lipid A structures of Porphyromonas gingivalis LPS differentially activate TLR4-mediated NF-κB signal transduction cascade and immuno-inflammatory response in human gingival fibroblasts. PloS One. 2013;8:e58496.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. de Bont CM, Koopman WJH, Boelens WC, Pruijn GJM. Stimulus-dependent chromatin dynamics, citrullination, calcium signalling and ROS production during NET formation. Biochim Biophys Acta Mol Cell Res. 2018;1865:1621–9.

    Article  PubMed  Google Scholar 

  56. Tamura K, Miyato H, Kanamaru R, Sadatomo A, Takahashi K, Ohzawa H, et al. Activated neutrophils inhibit chemotactic migration of activated T lymphocytes to CXCL11 by multiple mechanisms. Cell Immunol. 2023;384:104663.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

Figures 1A and 6 were illustrated using Biorender.com.

Funding

This study was supported by National Natural Science Foundation of China (82470956, 82001008); Shanghai Municipal Health Commission (JKKPZX-2023-A07); Chinese Stomatological Association (Young Talent Project of Orthodontics, COS-B202105); China Oral Health Foundation (A2023-029); Fundamental Research Funds for the Central Universities (22120240282) to CL.

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Author notes

  1. These authors contributed equally: Beibei Chen, Danni Song.

Authors and Affiliations

  1. Shanghai Engineering Research Center of Tooth Restoration and Regeneration & Tongji Research Institute of Stomatology & Department of Orthodontics, Stomatological Hospital and Dental School, Tongji University, Shanghai, China

    Beibei Chen, Danni Song, Yongming Li & Chongshan Liao

  2. Division of Periodontology & Implant Dentistry, Faculty of Dentistry, the University of Hong Kong, Hong Kong SAR, China

    Tianfan Cheng & Lijian Jin

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  1. Beibei Chen
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  6. Chongshan Liao
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Contributions

BBC and CSL conceived the study. BBC, DNS, CSL and YML designed the project and experiment. BBC and DNS collected samples and performed the experiment. BBC wrote the manuscript and prepared the figures. BBC, CSL, YML, TFC and LJJ revised the manuscript. All authors agreed to publish the final version.

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Correspondence to Yongming Li or Chongshan Liao.

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The studies involving all animal experiments were reviewed and approved from the Ethics Committee of Stomatology Hospital, Tongji University (Approval No. 2022-DW-013). All methods were performed in accordance with the relevant guidelines and regulations.

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Chen, B., Song, D., Cheng, T. et al. Neutrophil extracellular traps aggravate periodontitis by disturbing regulatory T-cell differentiation. Genes Immun 26, 475–485 (2025). https://doi.org/10.1038/s41435-025-00350-6

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