{"title":"ZBP1-NLRP3 axis integrates PANoptosis and ferroptosis during inflammatory injury in human dental pulp fibroblasts","authors":"Ai-E. He , Xing Wang , Ni Xie, Yun-He Xiao","doi":"10.1016/j.archoralbio.2025.106398","DOIUrl":null,"url":null,"abstract":"<div><h3>Objective</h3><div>To define how Z-DNA binding protein 1 (ZBP1) and NOD-like receptor family pyrin domain-containing 3 (NLRP3) signaling regulate lipopolysaccharide (LPS)-induced inflammation, PANoptosis, and ferroptosis in human dental pulp fibroblasts (HDPFs).</div></div><div><h3>Design</h3><div>HDPFs were treated with LPS, and <em>ZBP1</em> and <em>NLRP3</em> were silenced using small interfering RNA (siRNA), individually or in combination. Inflammatory mediators and death-pathway markers were quantified by quantitative real-time PCR (qRT-PCR), Western blotting, enzyme-linked immunosorbent assay (ELISA), and biochemical assays; Annexin V/propidium iodide flow cytometry assessed cell-death distributions.</div></div><div><h3>Results:</h3><div>LPS significantly increased ZBP1 and NLRP3 expression and elevated cytokine/chemokine release; each was attenuated by <em>ZBP1</em> or <em>NLRP3</em> knockdown, with the greatest reduction after dual silencing. LPS triggered PANoptosis, as indicated by increased Annexin V⁺/PI⁺ cell populations and upregulation of caspase-1, cleaved caspase-8, RIPK3, GSDMD, and p-MLKL/MLKL, which were significantly reduced by inhibition of the ZBP1-NLRP3 axis. Ferroptosis features were also evident after LPS, including impaired iron homeostasis (downregulated ferritin heavy chain 1 [FTH1] and ferroportin [FPN1] with Fe²⁺ accumulation), enhanced lipid peroxidation (upregulated ALOX15, LPCAT3, PTGS2 with increased malondialdehyde and lipid reactive oxygen species), and weakened antioxidant defenses (reduced glutathione peroxidase-4 [GPX4], solute carrier family 7 member 11 [SLC7A11], glutathione, and GPX4 activity). These changes were mitigated by single-gene silencing and most effectively by dual knockdown.</div></div><div><h3>Conclusion</h3><div>The ZBP1-NLRP3 axis acts upstream to coordinate LPS-induced PANoptosis and ferroptosis in HDPFs. Targeting this axis dampens inflammatory cell death and oxidative-metabolic dysregulation, highlighting a potential therapeutic strategy for pulpitis-related tissue injury.</div></div>","PeriodicalId":8288,"journal":{"name":"Archives of oral biology","volume":"180 ","pages":"Article 106398"},"PeriodicalIF":2.1000,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Archives of oral biology","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0003996925002262","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"DENTISTRY, ORAL SURGERY & MEDICINE","Score":null,"Total":0}
引用次数: 0
Abstract
Objective
To define how Z-DNA binding protein 1 (ZBP1) and NOD-like receptor family pyrin domain-containing 3 (NLRP3) signaling regulate lipopolysaccharide (LPS)-induced inflammation, PANoptosis, and ferroptosis in human dental pulp fibroblasts (HDPFs).
Design
HDPFs were treated with LPS, and ZBP1 and NLRP3 were silenced using small interfering RNA (siRNA), individually or in combination. Inflammatory mediators and death-pathway markers were quantified by quantitative real-time PCR (qRT-PCR), Western blotting, enzyme-linked immunosorbent assay (ELISA), and biochemical assays; Annexin V/propidium iodide flow cytometry assessed cell-death distributions.
Results:
LPS significantly increased ZBP1 and NLRP3 expression and elevated cytokine/chemokine release; each was attenuated by ZBP1 or NLRP3 knockdown, with the greatest reduction after dual silencing. LPS triggered PANoptosis, as indicated by increased Annexin V⁺/PI⁺ cell populations and upregulation of caspase-1, cleaved caspase-8, RIPK3, GSDMD, and p-MLKL/MLKL, which were significantly reduced by inhibition of the ZBP1-NLRP3 axis. Ferroptosis features were also evident after LPS, including impaired iron homeostasis (downregulated ferritin heavy chain 1 [FTH1] and ferroportin [FPN1] with Fe²⁺ accumulation), enhanced lipid peroxidation (upregulated ALOX15, LPCAT3, PTGS2 with increased malondialdehyde and lipid reactive oxygen species), and weakened antioxidant defenses (reduced glutathione peroxidase-4 [GPX4], solute carrier family 7 member 11 [SLC7A11], glutathione, and GPX4 activity). These changes were mitigated by single-gene silencing and most effectively by dual knockdown.
Conclusion
The ZBP1-NLRP3 axis acts upstream to coordinate LPS-induced PANoptosis and ferroptosis in HDPFs. Targeting this axis dampens inflammatory cell death and oxidative-metabolic dysregulation, highlighting a potential therapeutic strategy for pulpitis-related tissue injury.
期刊介绍:
Archives of Oral Biology is an international journal which aims to publish papers of the highest scientific quality in the oral and craniofacial sciences. The journal is particularly interested in research which advances knowledge in the mechanisms of craniofacial development and disease, including:
Cell and molecular biology
Molecular genetics
Immunology
Pathogenesis
Cellular microbiology
Embryology
Syndromology
Forensic dentistry