Fibroblast derived C3 promotes the progression of experimental periodontitis through macrophage M1 polarization and osteoclast differentiation

Abstract

Complement C3 plays a critical role in periodontitis. However, its source, role and underlying mechanisms remain unclear. In our study, by analyzing single-cell sequencing data from mouse model of periodontitis, we identified that C3 is primarily derived from periodontal fibroblasts. Subsequently, we demonstrated that C3a has a detrimental effect in ligature-induced periodontitis. C3ar ^−/− mice exhibited significantly less destruction of periodontal support tissues compared to wild-type mice, characterized by mild gingival tissue damage and reduced alveolar bone loss. This reduction was associated with decreased production of pro-inflammatory mediators and reduced osteoclast infiltration in the periodontal tissues. Mechanistic studies suggested that C3a could promote macrophage polarization and osteoclast differentiation. Finally, by analyzing single-cell sequencing data from the periodontal tissues of patients with periodontitis, we found that the results observed in mice were consistent with human data. Therefore, our findings clearly demonstrate the destructive role of fibroblast-derived C3 in ligature-induced periodontitis, driven by macrophage M1 polarization and osteoclast differentiation. These data strongly support the feasibility of C3a-targeted interventions for the treatment of human periodontitis.

References

[1] HajishengallisG. Periodontitis: from microbial immune subversion to systemic inflammation. Nat. Rev. Immunol., 2014, 15: 30-44

[2] EkePI, BorgnakkeWS, GencoRJ. Recent epidemiologic trends in periodontitis in the USA. Periodontology 2000, 2019, 82: 257-267

[3] TrindadeD, et al.. Prevalence of periodontitis in dentate people between 2011 and 2020: A systematic review and meta-analysis of epidemiological studies. J. Clin. Periodontol., 2023, 50: 604-626

[4] HienzSA, PaliwalS, IvanovskiS. Mechanisms of Bone Resorption in Periodontitis. J. Immunol. Res., 2015, 2015: 1-10

[5] ParsegianK, RandallD, CurtisM, IoannidouE. Association between periodontitis and chronic kidney disease. Periodontology 2000, 2022, 89: 114-124

[6] PaiSI, MatheusHR, GuastaldiFPS. Effects of periodontitis on cancer outcomes in the era of immunotherapy. Lancet Healthy Longev., 2023, 4: e166-e175

[7] Krutyhołowa, A. et al. Host and bacterial factors linking periodontitis and rheumatoid arthritis. Front. Immunol. https://doi.org/10.3389/fimmu.2022.980805 (2022).

[8] Lee, H. et al. Immunological link between periodontitis and type 2 diabetes deciphered by single-cell RNA analysis. Clin. Transl. Med. https://doi.org/10.1002/ctm2.1503 (2023).

[9] Muñoz AguileraE, et al.. Periodontitis is associated with hypertension: a systematic review and meta-analysis. Cardiovasc. Res., 2020, 116: 28-39

[10] Meng, X. et al. Periodontitis exacerbates pulmonary hypertension by promoting IFNγ+ T cell infiltration in mice. Int. J. Oral Sci. https://doi.org/10.1038/s41368-024-00291-2 (2024).

[11] SanzM, et al.. Periodontitis and cardiovascular diseases: consensus report. J. Clin. Periodontol., 2020, 47: 268-288

[12] Kim, T.-H. et al. A literature review of bioactive substances for the treatment of periodontitis: In vitro, in vivo and clinical studies. Heliyon https://doi.org/10.1016/j.heliyon.2024.e24216 (2024).

[13] HanJ, MenicaninD, GronthosS, BartoldPM. Stem cells, tissue engineering and periodontal regeneration. Aust. Dent. J., 2013, 59: 117-130

[14] RanS, et al.. Recent advances in injectable hydrogel therapies for periodontitis. J. Mater. Chem. B, 2024, 12: 6005-6032

[15] HajishengallisG. Immunomicrobial pathogenesis of periodontitis: keystones, pathobionts, and host response. Trends Immunol., 2014, 35: 3-11

[16] HawksworthOA, LiXX, CoulthardLG, WolvetangEJ, WoodruffTM. New concepts on the therapeutic control of complement anaphylatoxin receptors. Mol. Immunol., 2017, 89: 36-43

[17] Hajishengallis, G. et al. Complement-dependent mechanisms and interventions in periodontal disease. Front. Immunol. https://doi.org/10.3389/fimmu.2019.00406 (2019).

[18] FriščićJ, et al.. The complement system drives local inflammatory tissue priming by metabolic reprogramming of synovial fibroblasts. Immunity, 2021, 54: 1002-1021.e1010

[19] CuiJ, WuX, SongY, ChenY, WanJ. Complement C3 exacerbates renal interstitial fibrosis by facilitating the M1 macrophage phenotype in a mouse model of unilateral ureteral obstruction. Am. J. Physiol., 2019, 317: F1171-F1182

[20] CoulthardLG, WoodruffTM. Is the complement activation product C3a a proinflammatory molecule? Re-evaluating the evidence and the myth. J. Immunol., 2015, 194: 3542-3548

[21] SimaC, ViniegraA, GlogauerM. Macrophage immunomodulation in chronic osteolytic diseases—the case of periodontitis. J. Leukoc. Biol., 2019, 105: 473-487

[22] Sun, X. et al. Polarized macrophages in periodontitis: characteristics, function, and molecular signaling. Front. Immunol. https://doi.org/10.3389/fimmu.2021.763334 (2021).

[23] YuanG, et al.. RGS12 Drives macrophage activation and osteoclastogenesis in periodontitis. J. Dent. Res., 2021, 101: 448-457

[24] Cui, Y. et al. Melatonin engineering M2 macrophage-derived exosomes mediate endoplasmic reticulum stress and immune reprogramming for periodontitis therapy. Adv. Sci. https://doi.org/10.1002/advs.202302029 (2023).

[25] WuX, ChenH, WangY, GuY. Akt2 affects periodontal inflammation via altering the M1/M2 Ratio. J. Dent. Res., 2020, 99: 577-587

[26] ZhuangZ, et al.. Induction of M2 macrophages prevents bone loss in murine periodontitis models. J. Dent. Res., 2018, 98: 200-208

[27] Jang, J. S. et al. PINK1 restrains periodontitis-induced bone loss by preventing osteoclast mitophagy impairment. Redox Biol. https://doi.org/10.1016/j.redox.2023.103023 (2024).

[28] ChenY, et al.. The effect of the Litcubanine A on the treatment of murine experimental periodontitis by inhibiting monocyte–macrophage chemotaxis and osteoclast differentiation. J. Periodontal Res., 2023, 58: 948-958

[29] Kitaura, H. et al. Osteocyte-related cytokines regulate osteoclast formation and bone resorption. Int. J. Mol. Sci. https://doi.org/10.3390/ijms21145169 (2020).

[30] Lin, P. et al. Application of ligature-induced periodontitis in mice to explore the molecular mechanism of periodontal disease. Int. J. Mol. Sci. https://doi.org/10.3390/ijms22168900 (2021).

[31] Pan, W., Wang, Q. & Chen, Q. The cytokine network involved in the host immune response to periodontitis. Int. J. Oral Sci. https://doi.org/10.1038/s41368-019-0064-z (2019).

[32] Didilescu, A. C., Chinthamani, S., Scannapieco, F. A. & Sharma, A. NLRP3 inflammasome activity and periodontal disease pathogenesis—a bidirectional relationship. Oral Dis. https://doi.org/10.1111/odi.15005 (2024).

[33] MonachPA. Complement. Arthritis Rheumatol., 2023, 76: 1-8

[34] LamsterIB, AhloJK. Analysis of gingival crevicular fluid as applied to the diagnosis of oral and systemic diseases. Ann. N. Y. Acad. Sci., 2007, 1098: 216-229

[35] PattersMR, NiekrashCE, LangNP. Assessment of complement cleavage in gingival fluid during experimental gingivitis in man. J. Clin. Periodontol., 2005, 16: 33-37

[36] NiekrashCE, PattersMR. Simultaneous assessment of complement components C3, C4, and B and their cleavage products in human gingival fluid. J. Periodontal Res., 2006, 20: 268-275

[37] ReisES, et al.. Safety profile after prolonged C3 inhibition. Clin. Immunol., 2018, 197: 96-106

[38] Dobó, J., Kocsis, A., Dani, R. & Gál, P. Proprotein convertases and the complement system. Front. Immunol. https://doi.org/10.3389/fimmu.2022.958121 (2022).

[39] WestEE, KemperC. Complosome—the intracellular complement system. Nat. Rev. Nephrol., 2023, 19: 426-439

[40] KolevM, et al.. Diapedesis-induced integrin signaling via LFA-1 facilitates tissue immunity by inducing intrinsic complement C3 expression in immune cells. Immunity, 2020, 52: 513-527.e518

[41] Santos-López, J., de la Paz, K., Fernández, F. J. & Vega, M. C. Structural biology of complement receptors. Front. Immunol. https://doi.org/10.3389/fimmu.2023.1239146 (2023).

[42] WuK-Y, et al.. The C3a/C3aR axis mediates anti-inflammatory activity and protects against uropathogenic E coli–induced kidney injury in mice. Kidney Int., 2019, 96: 612-627

[43] AsgariE, et al.. C3a modulates IL-1β secretion in human monocytes by regulating ATP efflux and subsequent NLRP3 inflammasome activation. Blood, 2013, 122: 3473-3481

[44] Xue, C., Guo, Q., Subramanian, H., Gupta, K. & Ali, H. Regulation of C3a receptor signaling in human mast cells by G protein coupled receptor kinases. PLoS ONE https://doi.org/10.1371/journal.pone.0022559 (2011).

[45] Zhang, C. et al. Complement C3a signaling facilitates skeletal muscle regeneration by regulating monocyte function and trafficking. Nat. Commun. https://doi.org/10.1038/s41467-017-01526-z (2017).

[46] Liu, X. et al. Osteoclast differentiation and formation induced by titanium implantation through complement C3a. Mater. Sci. Eng. C https://doi.org/10.1016/j.msec.2021.111932 (2021).

[47] Van RaemdonckK, et al.. CCL21/CCR7 signaling in macrophages promotes joint inflammation and Th17-mediated osteoclast formation in rheumatoid arthritis. Cell. Mol. Life Sci., 2019, 77: 1387-1399

[48] IgnatiusA, et al.. Complement C3a and C5a modulate osteoclast formation and inflammatory response of osteoblasts in synergism with IL-1β. J. Cell. Biochem., 2011, 112: 2594-2605

[49] Kondo, T., Gleason, A., Okawa, H., Hokugo, A. & Nishimura, I. Mouse gingival single-cell transcriptomic atlas identified a novel fibroblast subpopulation activated to guide oral barrier immunity in periodontitis. eLife https://doi.org/10.7554/eLife.88183 (2023).

[50] WilliamsDW, et al.. Human oral mucosa cell atlas reveals a stromal-neutrophil axis regulating tissue immunity. Cell, 2021, 184: 4090-4104.e4015

Funding

National Natural Science Foundation of China (National Science Foundation of China)(2022YFC2504200)

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