Chronic rhinosinusitis with nasal polyps (CRSwNP) is an inflammatory condition characterized by outgrowth of sinonasal tissue [1]. According to the degree of eosinophilic infiltration, CRSwNP can be further divided into eosinophilic CRSwNP (eCRSwNP) and non-eosinophilic CRSwNP (neCRSwNP) [2]. The eCRSwNP phenotype is characterized by predominant Th2 immune responses, while neCRSwNP displays a heterogeneous inflammatory profile involving Th1, Th2, and Th17 pathways [3]. Regardless of these distinct immunological patterns, both subtypes demonstrate substantial subepithelial fibrin accumulation [4]. Fibrin degradation is facilitated by plasmin, which is generated from plasminogen upon cleavage by either urokinase plasminogen activator (uPA) or tissue plasminogen activator (tPA) [5]. Previous reports have shown significant downregulation of tPA, but not uPA, in both eCRSwNP and neCRSwNP patients [6]. Exposure to both Th1 and Th2 inflammatory mediators has been shown to downregulate tPA in cultured nasal epithelial cells [7]. However, the mechanisms underlying their regulation in CRSwNP remain poorly understood. Elucidating the mechanisms driving fibrin accumulation in CRSwNP could lead to improved therapeutic strategies.

SERPIN family B member 2 (SERPINB2), also known as plasminogen activator inhibitor type 2 (PAI-2), is a coagulation factor that inactivates uPA and tPA [8]. Previous studies have shown that SERPINB2 is dramatically elevated in bronchial epithelial cells of asthmatic patients and is positively associated with pivotal clinical markers, such as FEV1, FeNO, eosinophil counts, and asthma severity [9]. Single-cell RNA sequencing analysis has identified SERPINB2 as one of the core upregulated genes in human bronchial epithelial cells after segmental allergen challenge [10]. We have previously shown that SERPINB2 upregulated by IL-13 promotes the expression of 15-lipoxygenase-1 (15LO1), CCL26 and iNOS through activation of the STAT6 pathway in nasal polyp epithelial cells [11]. We also reported that elevated SERPINB2 in allergic rhinitis patients regulates MUC5AC expression via STAT6 signaling in nasal epithelial cells [12]. Based on these findings, we hypothesize that elevated SERPINB2 in nasal epithelial cells may suppress the expression and activity of tPA, thereby contributing to fibrin deposition in nasal polyps.

To test this hypothesis, SERPINB2 and tPA expression were analyzed in nasal polyp (NP) tissues from CRSwNP patients and turbinate tissues from healthy controls. Their colocalization was examined by immunofluorescence. Using air-liquid interface (ALI) cultures of primary nasal epithelial cells, the regulatory effects of SERPINB2 on tPA were examined through cytokine stimulation, recombinant protein treatment, and RNA interference. tPA enzymatic activity and fibrinolytic function were measured by fluorometric assays and D-dimer ELISA. Our results demonstrated that increased SERPINB2 in nasal polyp epithelial cells suppresses tPA expression and reduces tPA activity in CRSwNP.

In this study, we identified a novel SERPINB2-tPA regulatory axis in NECs that contributes to fibrin deposition in CRSwNP. Our findings demonstrate that SERPINB2 expression is significantly elevated in NP tissues and inversely correlates with tPA levels, with epithelial cells serving as the primary cellular source of this dysregulation. Importantly, we uncovered a dual inhibitory mechanism whereby SERPINB2 not only directly inhibits tPA enzymatic activity through classic SERPIN-mediated protease inhibition, but also suppresses tPA expression at the transcriptional level. This regulation is driven by both Th1 (IFN-$\gamma$) and Th2 (IL-13) cytokines, suggesting that irrespective of inflammatory endotype, CRSwNP patients may experience convergent impairment of local fibrinolytic capacity. The functional consequence of this SERPINB2-tPA imbalance is severely compromised fibrin degradation, which may create a permissive microenvironment for persistent tissue remodeling and polyp formation. These findings provide a mechanistic explanation for the fibrin accumulation observed in nasal polyps and identify SERPINB2 as a potential therapeutic target for the prevention of polyp development and recurrence.

The inverse relationship between SERPINB2 and tPA observed in nasal polyp tissues parallels findings in other fibroproliferative disorders [13, 14], suggesting a conserved pathogenic mechanism. Similar dysregulation has been reported in infectious pulmonary disease, chronic obstructive pulmonary disease, and allergic asthma [15, 16, 17], where impaired fibrinolysis contributes to excessive extracellular matrix deposition and progressive fibrosis. In the tumor microenvironment, elevated SERPINB2 has been associated with fibrin-rich matrices that promote cancer cell invasion and metastasis [18, 19]. However, our study reveals features unique to upper airway disease. Unlike PAI-1, SERPINB2 is highly expressed in nasal polyp epithelium, highlighting the central role of the airway epithelium in local hemostatic balance. The predominant epithelial localization also suggests that environmental triggers (allergens, pathogens, and pollutants) that directly stimulate epithelial cells may initiate the fibrinolytic dysfunction cascade [20]. Furthermore, the convergent regulation by both Th1 and Th2 cytokines distinguishes CRSwNP from other conditions where fibrinolytic impairment is typically associated with a specific inflammatory profile. This finding may explain why nasal polyps develop across different CRSwNP endotypes and why anti-inflammatory therapies targeting a single cytokine pathway show variable efficacy in preventing polyp recurrence.

Perhaps the most significant finding of our study is the discovery that SERPINB2 employs a dual mechanism to drive fibrinolytic dysfunction in CRSwNP. Excessive fibrin deposition is a pathological hallmark of CRSwNP, contributing to tissue remodeling and polyp recurrence [21, 22, 23]. The fibrinolytic system, primarily controlled by tPA and its inhibitors, normally maintains tissue homeostasis by clearing fibrin matrices [24]. However, the mechanisms underlying fibrinolytic impairment in nasal polyps have remained poorly understood. Our findings demonstrate that SERPINB2 disrupts this balance through two complementary pathways. Beyond its known function as a direct tPA inhibitor via SERPIN-mediated enzymatic inactivation, our data reveal a novel transcriptional regulatory role. Recombinant SERPINB2 dose-dependently suppressed tPA expression, while SERPINB2 knockdown rescued cytokine-induced tPA downregulation, providing strong evidence for this previously unrecognized mechanism. This dual inhibitory action—suppressing tPA gene expression while simultaneously inactivating residual tPA enzymatic activity—creates a synergistic anti-fibrinolytic effect that severely compromises local fibrin clearance in polyp tissues. Whether these two mechanisms interact through feedback loops remains an open question. SERPINB2-mediated inhibition of tPA activity may secondarily alter the local cytokine milieu and reinforce SERPINB2 expression in a feed-forward manner; conversely, tPA may reciprocally regulate SERPINB2 transcription through plasmin-dependent signaling. Elucidating the temporal hierarchy and potential interdependence of these two mechanisms represents an important direction for future investigation.

The molecular mechanisms underlying SERPINB2-mediated transcriptional suppression of tPA warrant further investigation. Potential mechanisms include: (i) SERPINB2 acting through cell surface receptor-mediated transduction signaling to modulate downstream transcriptional activity, consistent with the demonstrated GPER/SERPINB2 signaling axis [25]; (ii) intracellular SERPINB2 interacting with transcription factors at the PLAT promoter, supported by evidence that IRF1 functions as an upstream transcriptional regulator in type 1 inflammatory environments [26]; and (iii) epigenetic modifications of the tPA gene locus, which remain speculative and require dedicated investigation. These possibilities align with the emerging recognition that SERPINs exert functions beyond classical protease inhibition [27, 28]. The convergent suppression of tPA by IFN-$\gamma$ and IL-13 is particularly noteworthy: despite engaging distinct upstream pathways (STAT1 vs. STAT6) [11, 29], both cytokines coordinately upregulate SERPINB2 and downregulate tPA, implying the existence of shared downstream transcriptional repressors at the PLAT locus, potentially within the JAK/STAT signaling network that operates in CRSwNP [30]. Identifying these convergence points may reveal broadly applicable therapeutic targets across CRSwNP endotypes.

Our findings have important translational implications for the management of CRSwNP. First, the epithelial expression of SERPINB2 makes it potentially accessible as a disease biomarker through minimally invasive nasal brushing or lavage fluid sampling. Elevated SERPINB2 levels could serve as a surrogate marker of impaired fibrinolytic capacity and identify patients at high risk for polyp recurrence. Second, our knockdown experiments provide proof-of-concept that targeting SERPINB2 can restore tPA expression and fibrinolytic function. While therapeutic development would require extensive validation, potential strategies include neutralizing antibodies or small molecule inhibitors targeting the SERPINB2-tPA interaction. Ultimately, defective fibrin clearance and subsequent fibrin overload represent a shared pathogenic pathway underlying edematous changes across CRSwNP endotypes. Interventions targeting fibrinolytic dysfunction could provide treatment benefits independent of endotype by disrupting the self-reinforcing cascade linking fibrin deposition, tissue remodeling, and polyp formation.

Several limitations of this study should be acknowledged. First, this study relies primarily on in vitro ALI cultures and cross-sectional clinical tissue samples, and lacks in vivo functional validation. Future studies employing animal models with SERPINB2 knockout or epithelial-specific overexpression to assess fibrinolytic activity, fibrin deposition, and polyp formation would provide stronger causal evidence. Second, our in vitrostudies utilized healthy control-derived ALI cultures, which may not fully reflect the altered responsiveness of the polyp epithelium or the complex multicellular interactions in nasal polyp tissues. Third, our fibrinolysis assay utilized a serum-based system, which measures tPA-mediated plasminogen activation but does not fully recapitulate the tissue fibrin microenvironment of nasal polyps; future studies incorporating 3D fibrin gel models or ex vivo tissue explants would provide additional spatial and contextual information. Fourth, the clinical findings of this study are based on cross-sectional observations; longitudinal studies correlating SERPINB2 and tPA dynamics with disease progression and treatment response are needed to establish their clinical translational relevance.

This study identifies a novel SERPINB2-tPA regulatory axis that drives fibrinolytic dysfunction in CRSwNP. We demonstrate that SERPINB2 employs a dual inhibitory mechanism—suppressing tPA expression at the transcriptional level while directly inhibiting its enzymatic activity—resulting in severely impaired local fibrinolysis. The convergent upregulation of SERPINB2 by both the Th1 cytokine IFN-$\gamma$ and the Th2 cytokine IL-13 suggests that fibrinolytic impairment occurs across different inflammatory endotypes in CRSwNP. These findings provide mechanistic insights into the fibrin accumulation and persistent tissue remodeling in nasal polyps, and identify SERPINB2 as a promising therapeutic target for the prevention of polyp formation and recurrence.

The data presented in this study are available on request from the corresponding author.

Conceptualization, WZ, and ZL; methodology, YZ, JZ, SZ, and ZL; validation, YZ, JZ, and SZ; formal analysis, YZ, and ZL; investigation, YZ, JZ, SZ, and ZL; resources, WZ, and ZL; data curation, YZ, JZ, and SZ; writing—original draft preparation, YZ, and ZL; writing—review and editing, YZ, WZ, and ZL; visualization, YZ; supervision, WZ, and ZL; funding acquisition, WZ, and ZL. All authors contributed to editorial changes in the manuscript. All authors read and approved the final manuscript. All authors have participated sufficiently in the work and agreed to be accountable for all aspects of the work.

This study was approved by the Ethical Committee of Shanghai Sixth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, with written informed consent obtained from all participants. Approval Number: 2022-069. The study was carried out in accordance with the guidelines of the Declaration of Helsinki.

We would like to express our sincere gratitude to all patients and healthy volunteers who contributed samples to this study. We also thank all peer reviewers for their constructive opinions and suggestions.

This work was supported by the National Natural Science Foundation of China [No. 82271137 and No. 82000951]; Shanghai Municipal Health Commission [SHWSRS(2023)62].

The authors declare no conflicts of interest.

During the preparation of this work, the authors used Claude to improve the grammar, readability, and organization of the manuscript text. After using this tool, the authors reviewed and edited the content as needed and took full responsibility for the content of the publication.

Supplementary material associated with this article can be found, in the online version, at https://doi.org/10.31083/FBL50406.