Spatially resolved single cell atlas deciphers SAA1 inflammatory epithelial cells

Abstract

Disruption of epithelial integrity is a pivotal event in inflammation, disease pathogenesis, and tissue homeostasis. To investigate these processes in chronic inflammatory disease, we performed spatial transcriptomics integrated with single-cell RNA sequencing (scRNA-seq) on human gingival tissue, coupled with metagenomic analysis of matched subgingival plaque. This approach allowed in situ characterization of epithelial heterogeneity and microbiome-epithelium-connective tissue crosstalk. We identified a distinct inflammatory epithelial subpopulation (SAA1+Epi), situated within the junctional epithelium, that becomes activated through the TLR2-PITX2 axis by Porphyromonas gingivalis lipopolysaccharide. These SAA1+Epi cells secrete TGFβ, which induces an inflammatory program in the connective tissue by driving the differentiation of inflammation-associated fibroblasts (C3+FB) via the PI3K/Akt pathway. Concurrently, SAA1+Epi cells express chemotactic factors such as CXCL6 to recruit NK cells, thereby sustaining the inflammatory niche. The transcription factor PITX2 emerged as a critical regulator of SAA1+Epi differentiation; targeting PITX2 suppressed   C3+FB induction and natural killer (NK) cells recruitment, ultimately attenuating periodontitis progression. Our findings position SAA1+Epi as a frontline responder to dysbiotic bacteria at the inflammatory interface and underscore its essential role in regulating epithelial-connective tissue homeostasis during inflammation.

References

[1] Shannon JP, et al. Group 1 innate lymphoid-cell-derived interferon-gamma maintains anti-viral vigilance in the mucosal epithelium. Immunity, 2021, 54: 276-290.e275.

[2] Roncero-Carol J, et al. Epithelial cells provide immunocompetence to the early embryo for bacterial clearance. Cell Host Microbe, 2025, 33: 1106-1120.e1108.

[3] Sumbal J, et al. Contractile fibroblasts form a transient niche for the branching mammary epithelium. Nat. Commun., 2025, 16. 8576

[4] Bachtel ND, et al. Intestinal mast cell-derived leukotrienes mediate the anaphylactic response to ingested antigens. Science, 2025, 389. eadp0246

[5] Azulay N, et al. A spatial atlas of human gastrointestinal acute GVHD reveals epithelial and immune dynamics underlying disease pathophysiology. Sci. Transl. Med., 2025, 17. eadu6032

[6] Garrett S, Asada MC, Sun J. Axin1’s mystique in manipulating microbiome amidst colitis. Gut Microbes, 2023, 15: 2286674.

[7] Quintero M, Samuelson LC. Paneth cells: dispensable yet irreplaceable for the intestinal stem cell niche. Cell Mol. Gastroenterol. Hepatol., 2025, 19: 101443.

[8] Kang GJ, et al. Quercetagetin inhibits macrophage-derived chemokine in HaCaT human keratinocytes via the regulation of signal transducer and activator of transcription 1, suppressor of cytokine signalling 1 and transforming growth factor-beta1. Br. J. Dermatol., 2014, 171: 512-523.

[9] Sumiyoshi K, et al. TGF-beta/Smad signaling inhibits IFN-gamma and TNF-alpha-induced TARC (CCL17) production in HaCaT cells. J. Dermatol. Sci., 2003, 31: 53-58.

[10] Taebnia N, Romling U, Lauschke VM. In vitro and ex vivo modeling of enteric bacterial infections. Gut Microbes, 2023, 15: 2158034.

[11] Disease, G. B. D., Injury & Risk Factor, C. Burden of 375 diseases and injuries, risk-attributable burden of 88 risk factors, and healthy life expectancy in 204 countries and territories, including 660 subnational locations, 1990-2023: a systematic analysis for the Global Burden of Disease Study 2023. Lancet406, 1873–1922 (2025).

[12] Xu T, et al. Oral virome metagenomic catalog links Porphyromonas gingivalis phages to obesity and type 2 diabetes. Cell Rep. Med., 2025, 6. 102325

[13] Bashir NZ, Woolf BAR, Burgess S, Bernabé E. Periodontitis and Systemic Disease: The Impact of Covariate Selection. J. Dent. Res. 105, 59–66 (2026).

[14] Zhao X, et al. Administration of Porphyromonas gingivalis in pregnant mice enhances glycolysis and histone lactylation/ADAM17 leading to cleft palate in offspring. Int J. Oral. Sci., 2025, 17: 18.

[15] Chang AM, Kantrong N, Darveau RP. Maintaining homeostatic control of periodontal epithelial tissue. Periodontology, 2021, 86: 188-200.

[16] Abusleme L, Hoare A, Hong BY, Diaz PI. Microbial signatures of health, gingivitis, and periodontitis. Periodontology, 2021, 86: 57-78.

[17] Abusleme L, et al. The subgingival microbiome in health and periodontitis and its relationship with community biomass and inflammation. ISME J., 2013, 7: 1016-1025.

[18] Ji S, Choi Y. Microbial and host factors that affect bacterial invasion of the gingiva. J. Dent. Res., 2020, 99: 1013-1020.

[19] Kim TS, et al. Epithelial-derived interleukin-23 promotes oral mucosal immunopathology. Immunity, 2024, 57: 859-875.e811.

[20] Song B, et al. The role of Toll-like receptors in periodontitis. Oral. Dis., 2017, 23: 168-180.

[21] Zhang, H. et al. Interfacial coordination-engineered emulgel modulates the osteo-immune microenvironment in periodontitis treatment Via HIF1 signaling pathway-induced metabolic reprogramming. Adv Sci (Weinh). 12, e09998 (2025).

[22] Zhao YY, et al. Injectable cellular vesicle-based bone meal for inflammatory bone defect repair through restoring immune homeostasis. Theranostics, 2025, 15: 4465-4480.

[23] Cai XY, et al. Inflammation-triggered Gli1(+) stem cells engage with extracellular vesicles to prime aberrant neutrophils to exacerbate periodontal immunopathology. Cell Mol. Immunol., 2025, 22: 371-389.

[24] Ji X, et al. DnaK of Parvimonas micra extracellular vesicles interacts with the host fibroblasts BAG3-IKK-gamma axis to accelerate TNF-alpha secretion in oral lichen planus. Microbiome, 2025, 13. 164

[25] Li L, et al. Multi-omics data reveal that SAA1 + fibroblasts exacerbate periodontitis by regulating macrophage inflammation and chemotaxis. J. Transl. Med., 2025, 23. 882

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

[27] Chen Y, et al. Single-cell RNA landscape of the osteoimmunology microenvironment in periodontitis. Theranostics, 2022, 12: 1074-1096.

[28] Ren F, et al. Fibroblast derived C3 promotes the progression of experimental periodontitis through macrophage M1 polarization and osteoclast differentiation. Int. J. Oral. Sci., 2025, 17: 30.

[29] Easter QT, et al. Single-cell and spatially resolved interactomics of tooth-associated keratinocytes in periodontitis. Nat. Commun., 2024, 15. 5016

[30] Zhang Y, et al. Single-cell spatial atlas of smoking-induced changes in human gingival tissues. Int. J. Oral. Sci., 2025, 17: 60.

[31] Shi B, et al. The subgingival microbiome associated with periodontitis in type 2 diabetes mellitus. ISME J., 2020, 14: 519-530.

[32] Xu H, Qian Y, Jia S, Shi Z, Zhong Q. Comparative analysis of subgingival microbiota in patients with mild, moderate, and severe chronic periodontitis. Oral. Dis., 2023, 29: 2865-2877.

[33] Yost S, Duran-Pinedo AE, Teles R, Krishnan K, Frias-Lopez J. Functional signatures of oral dysbiosis during periodontitis progression revealed by microbial metatranscriptome analysis. Genome Med., 2015, 7. 27

[34] Kashiwagi I, et al. Smad2 and Smad3 inversely regulate TGF-beta autoinduction in Clostridium butyricum-activated dendritic cells. Immunity, 2015, 43: 65-79.

[35] Wang L, et al. Gut microbial bile acid metabolite skews macrophage polarization and contributes to high-fat diet-induced colonic inflammation. Gut Microbes, 2020, 12: 1-20.

[36] Ji Y, et al. Toll-like receptors TLR2 and TLR4 block the replication of pancreatic beta cells in diet-induced obesity. Nat. Immunol., 2019, 20: 677-686.

[37] Basu M, et al. Invasion of ovarian cancer cells is induced byPITX2-mediated activation of TGF-beta and Activin-A. Mol. Cancer, 2015, 14. 162

[38] Drouin J. 60 years of POMC: transcriptional and epigenetic regulation of POMC gene expression. J. Mol. Endocrinol., 2016, 56: T99T112.

[39] Citi S. Intestinal barriers protect against disease. Science, 2018, 359: 1097-1098.

[40] Gerassy-Vainberg S, et al. Radiation induces proinflammatory dysbiosis: transmission of inflammatory susceptibility by host cytokine induction. Gut, 2018, 67: 97-107.

[41] Feret N, et al. Decoding epithelial regeneration in the cornea: multi-omic analysis reveals cellular plasticity as central mechanism. Cell Mol. Biol. Lett., 2025, 30: 131.

[42] Sakthivel, K. et al. Integrin-Piezo1 axis drives ECM remodeling and invasion of 3D breast epithelium. Adv. Sci (Weinh).13, e09932 (2026).

[43] Shen Z, et al. The spatial transcriptomic landscape of human gingiva in health and periodontitis. Sci. China Life Sci., 2024, 67: 720-732.

[44] Caetano, A. J. et al. Defining human mesenchymal and epithelial heterogeneity in response to oral inflammatory disease. eLife10, e62810 (2021).

[45] Takahashi N, et al. Gingival epithelial barrier: regulation by beneficial and harmful microbes. Tissue Barriers, 2019, 7: e1651158.

[46] Yamamoto M, Aizawa R. Maintaining a protective state for human periodontal tissue. Periodontology, 2021, 86: 142-156.

[47] Leonard R, et al. Single-nuclei multiomics analysis identifies abnormal cardiomyocytes in a murine model of cardiac development. Nat. Commun., 2025, 16. 6947

[48] Reis LM, Seese SE, Costakos D, Semina EV. Congenital anterior segment ocular disorders: genotype-phenotype correlations and emerging novel mechanisms. Prog. Retin. Eye Res., 2024, 102: 101288.

[49] Tamura R, et al. PITX2 expression in non-functional pituitary neuroendocrine tumor with cavernous sinus invasion. Endocr. Pathol., 2019, 30: 81-89.

[50] Shao F, et al. Transcriptional programs of Pitx2 and Tfap2a/Tfap2b controlling lineage specification of mandibular epithelium during tooth initiation. PLoS Genet., 2024, 20: e1011364.

[51] Li Q, et al. PITX2 expression and Neanderthal introgression in HS3ST3A1 contribute to variation in tooth dimensions in modern humans. Curr. Biol., 2025, 35: 131-144.e136.

[52] Brozek, R., Lorenz, A., Dorocka-Bobkowska, B. & Kurpisz, M. Immunoinflammatory response in periodontal diseases. J. Physiol. Pharmacol.76, 10.26402/jpp.2025.3.02 (2025).

[53] Liu Y, et al. Single-cell analyses of the oral mucosa reveal immune cell signatures. J. Dent. Res., 2023, 102: 514-524.

[54] Wang Z, et al. The impact of aging on neutrophil functions and the contribution to periodontitis. Int. J. Oral. Sci., 2025, 17: 10.

[55] Kim TS, Moutsopoulos NM. Neutrophils and neutrophil extracellular traps in oral health and disease. Exp. Mol. Med., 2024, 56: 1055-1065.

[56] Nieto MA, Cano A. The epithelial-mesenchymal transition under control: global programs to regulate epithelial plasticity. Semin. Cancer Biol., 2012, 22: 361-368.

[57] Saliem SS, et al. Pathogenesis of periodontitis - A potential role for epithelial-mesenchymal transition. Jpn. Dent. Sci. Rev., 2022, 58: 268-278.

[58] Nieto MA. The ins and outs of the epithelial to mesenchymal transition in health and disease. Annu. Rev. Cell Dev. Biol., 2011, 27: 347-376.

[59] Lee J, et al. Human primary epithelial cells acquire an epithelial-mesenchymal-transition phenotype during long-term infection by the oral opportunistic pathogen, Porphyromonas gingivalis. Front. Cell Infect. Microbiol., 2017, 7: 493.

[60] Sisto, M., Ribatti, D. & Lisi, S. Organ fibrosis and autoimmunity: the role of inflammation in TGFbeta-dependent EMT. Biomolecules11, 310 (2021).

[61] Wu YR, Hsing CH, Chiu CJ, Chiang BL, Hsu YH. Eosinophil-derived interleukin-24 compromises epithelial integrity and aggravates airway remodeling in mouse models of allergic asthma. Nat. Commun., 2025, 16. 9248

[62] Fujita T, et al. Interleukin-8 induces DNA synthesis, migration and down-regulation of cleaved caspase-3 in cultured human gingival epithelial cells. J. Periodontal Res, 2015, 50: 479-485.

[63] Sztukowska MN, et al. Porphyromonas gingivalis initiates a mesenchymal-like transition through ZEB1 in gingival epithelial cells. Cell Microbiol., 2016, 18: 844-858.

[64] Marchesan J, et al. An experimental murine model to study periodontitis. Nat. Protoc., 2018, 13: 2247-2267.

[65] Zheng X, et al. Fusobacterium nucleatum extracellular vesicles are enriched in colorectal cancer and facilitate bacterial adhesion. Sci. Adv., 2024, 10. eado0016

[66] Haghverdi L, Lun ATL, Morgan MD, Marioni JC. Batch effects in single-cell RNA-sequencing data are corrected by matching mutual nearest neighbors. Nat. Biotechnol., 2018, 36: 421-427.

Funding

National Natural Science Foundation of China (National Science Foundation of China)(81991501)

Department of Science and Technology of Sichuan Province (Sichuan Provincial Department of Science and Technology)(2024NSFSC1584)

Research and Development Program, West China Hospital of Stomatology Sichuan University (RD-02-202510)

Rights & permissions

The Author(s)