Our results suggest that IL-25 secreted from the sinonasal epithelia and infiltrating mast cells plays a crucial role in the pathogenesis of CRS with NPs in Asian patients. In addition, our results suggest the novel possibility of treating nasal polyposis with anti-IL-25 therapy.

Our data suggest a role for IL-33 in the pathogenesis of CRSwNP via neutrophil recruitment. Therefore, anti-IL-33 may provide a new treatment strategy to target infiltrating neutrophils in CRSwNP.

Purpose Endotype in chronic rhinosinusitis (CRS) has been established in the last decade. However, the exact immunologic profile of CRS still has controversy because it has a considerable immunologic heterogeneity. Therefore, we investigated various inflammatory mediators according to different nasal tissues in chronic rhinosinusitis and compared them within the same subject. Methods We collected uncinate process mucosa (UP) and nasal polyp (NP) tissues from controls, CRS without NP (CRSsNP) and CRS with NP (CRSwNP). Expression levels of 28 inflammatory mediators including T helper (Th) 1, Th2, Th17, proinflammatory cytokines and remodeling markers were determined by multiplex immunoassay and were analyzed using paired tests as well as principal component analysis (PCA) to investigate endotype in each subtype of CRS. Results Signature inflammatory mediators are interleukin (IL)-5, C-C motif chemokine ligand (CCL)-24, monocyte chemoattractant protein (MCP)-4, and vascular cell adhesion molecule (VCAM)-1 in eosinophilic NP, whereas IL-17A, IL-1β, and matrix metallopeptidase (MMP)-9 were detected as signature inflammatory markers in non-eosinophilic NP. Despite differences in inflammatory cytokine profile between eosinophilic and non-eosinophilic NP, the common upregulation of IL-5, CCL-11, IL-23, IL-2Rα, VCAM-1, MMP-3 and MMP-9 were shown in NP compared to UP within the same subject. In the PCA, we observed that Th2 immune response was helpful in discriminating between nasal tissues in subtypes of CRS and that there was a partial overlap between non-eosinophilic NP and eosinophilic NP in terms of Th2 mediators. Conclusions Commonly upregulated mediators in NP were Th2-associated, compared with UP regardless of CRS subtypes, whereas signature markers were distinct in each NP subtype. These findings imply that Th2 inflammatory responses may play a role in the development of NP regardless of CRSwNP subtypes.

Purpose: Various immune cells, including eosinophils and neutrophils, are known to contribute to the development of chronic rhinosinusitis with nasal polyps (CRSwNP). However, the current understanding of the role of neutrophils in the development of CRSwNP still remains unclear. Therefore, we investigated risk factors for refractoriness of CRSwNP in an Asian population. Methods: Protein levels of 17 neutrophil-related mediators in nasal polyps (NPs) were determined by multiplex immunoassay, and exploratory factor analysis using principal component analysis was performed. Immunofluorescence analysis was conducted to detect human neutrophil elastase (HNE) or myeloperoxidase (MPO)-positive cells. Tissue eosinophilic nasal polyp (ENP) and tissue neutrophilia (Neu high) were defined as greater than 70 eosinophils and 20 HNE-positive cells, otherwise was classified into non-eosinophilic nasal polyp (NENP) and absence of tissue neutrophilia (Neu low). Results: In terms of disease control status, NENP-Neu low patients showed the higher rate of disease control than NENP-Neu high and ENP-Neu high patients. Linear by linear association demonstrated the trend in refractoriness from NENP-Neu low to NENP-Neu high or ENP-Neu low to ENP-Neu high. When multiple logistic regression was performed, tissue neutrophilia (hazard ratio, 4.38; 95% confidence interval, 1.76-10.85) was found as the strongest risk factor for CRSwNP refractoriness. Additionally, exploratory factor analysis revealed that interleukin (IL)-18, interferon-γ, IL-1Ra, tumor necrosis factor-α, oncostatin M, and MPO were associated with good disease control status, whereas IL-36α and IL-1α were associated with refractory disease control status. In subgroup analysis, HNE-positive cells and IL-36α were significantly upregulated in the refractory group (P = 0.0132 and P = 0.0395, respectively), whereas MPO and IL-18 showed higher expression in the controlled group (P = 0.0002 and P = 0.0009, respectively). Moreover, immunofluorescence analysis revealed that IL-36R + HNE +-double positive cells were significantly increased in the refractory group compared to the control group. We also found that the ratio of HNE-positive cells to α1 anti-trypsin was increased in the refractory group.

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BACKGROUND: Neutrophils present as major inflammatory cells in refractory chronic rhinosinusitis with nasal polyps (CRSwNP), regardless of the endotype. However, their role in the pathophysiology of CRSwNP remains poorly understood. We investigated factors predicting the surgical outcomes of CRSwNP patients with focus on neutrophilic localization. METHODS: We employed machine-learning methods such as the decision tree and random forest models to predict the surgical outcomes of CRSwNP. Immunofluorescence analysis was conducted to detect human neutrophil elastase (HNE), Bcl-2, and Ki-67 in NP tissues. We counted the immunofluorescence-positive cells and divided them into three groups based on the infiltrated area, namely, epithelial, subepithelial, and perivascular groups. RESULTS: On machine learning, the decision tree algorithm demonstrated that the number of subepithelial HNE-positive cells, Lund-Mackay (LM) scores, and endotype (eosinophilic or non-eosinophilic) were the most important predictors of surgical outcomes in CRSwNP patients. Additionally, the random forest algorithm showed that, after ranking the mean decrease in the Gini index or the accuracy of each factor, the top three ranking factors associated with surgical outcomes were the LM score, age, and number of subepithelial HNE-positive cells. In terms of cellular proliferation, immunofluorescence analysis revealed that Ki-67/HNE-double positive and Bcl-2/HNE-double positive cells were significantly increased in the subepithelial area in refractory CRSwNP. CONCLUSION: Our machine-learning approach and immunofluorescence analysis demonstrated that subepithelial neutrophils in NP tissues had a high expression of Ki-67 and could serve as a cellular biomarker for predicting surgical outcomes in CRSwNP patients.

BackgroundNon-eosinophilic nasal polyps (NPs) show less inflammatory changes and are less commonly associated with lower airway inflammatory disorders such as asthma, compared with eosinophilic NPs. However, the development of non-eosinophilic NPs which is a predominant subtype in Asian population still remains unclear.MethodsA total of 81 patients (45 with non-eosinophilic NPs and 36 with eosinophilic NPs) were enrolled. Clinical information and computed tomography (CT), endoscopic, and histological findings were investigated. Tissue samples were analyzed for total IgE levels and for mRNA expression levels of interleukin (IL)-4, IL–5, IL–13, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-17A, IL–22, IL-23p19, transforming growth factor (TGF)-β1, TGF-β2, TGF-β3, and periostin. Immunostaining assessment of Ki–67 as a proliferation marker was performed.ResultsWe found that epithelial in-growing patterns such as pseudocysts were more frequently observed in histological and endoscopic evaluations of non-eosinophilic NPs, which was linked to increase epithelial staining of Ki–67, a proliferating marker. Eosinophilic NPs were characterized by high infiltration of inflammatory cells, compared with non-eosinophilic NPs. To investigate the developmental course of each subtype, CT was analyzed according to CT scores and subtypes. Non-eosinophilic NPs showed more localized pattern and maxillary sinus involvement, but lesser olfactory involvement in early stage whereas eosinophilic NPs were characterized by diffuse ethmoidal and olfactory involvement. In addition, high ethmoidal/maxillary (E/M) CT scores, indicating ethmoidal dominant involvement, were one of surrogate markers for eosinophilic NP. E/M CT scores was positively correlated with levels of TH2 inflammatory markers, including IL–4, IL–5, periostin mRNA expression and total IgE levels in NPs, whereas levels of the TH1 cytokine, IFN- γ were inversely correlated. Moreover, if the combinatorial algorithm meet the three of the four markers, including IL–5 (<2.379), periostin (<3.889), IFN-γ (>0.316), and E/M ratio (<2.167), non-eosinophilic CRSwNP are diagnosed with a sensitivity of 84.4% and a specificity of 84.8%.ConclusionHistologic, immunologic and clinical data suggest that non-eosinophilic NPs showed enhanced epithelial alteration and more localized maxillary involvement. Combination of cutoff value on IL–5, periostin, IFN-γ, and E/M scores may be one of surrogate markers for non-eosinophil NP subtype.

Purpose Despite medical and surgical treatments, some cases of nasal polyps (NP) exhibit recidivism. However, the endotype of refractory chronic rhinosinusitis with NP (CRSwNP) remains unclear. Therefore, the objective of this study was to characterize the immunological profile of refractory CRSwNP. Methods The control (n =23), primary NP group (pNP, n =70) and refractory NP group (rNP, n =86) were enrolled in this study. Patients who underwent revision surgeries due to failed maximal medical treatment after primary surgery were defined as the rNP group. A total of 18 inflammatory markers were investigated in nasal tissues using multiplex cytokine assay or enzyme-linked immunosorbent assay. Results The clinical characteristics of rNP included more extensive disease and worse clinical course after surgery. Additionally, rNP subjects showed higher infection rate (mucopurulence and culture-positive rate), more frequent use of antibiotics and suffered from symptomatic bacterial infection, increased asthma morbidity compared to pNP. Cytokine profile analysis showed that levels of Th17-associated mediators (myeloperoxidase, interleukin (IL)-8, IL-17A and IL-23), B-cell activating factor (BAFF) and Th1 cytokine (interferon-γ) were up-regulated in rNP compared to controls and pNP. Human neutrophil elastase-positive cells were also enhanced in rNP compared with pNP. Upregulation of Th17/Th1mediators and BAFF were observed in rNP, regardless of tissue eosinophilia or asthmatic comorbidity. Interestingly, eosinophilic markers, such as eosinophil cationic protein and C-C motif chemokine ligand 24, were up-regulated in asthmatic rNP compared to pNP and controls. Levels of anti-dsDNA immunoglobulin (Ig) G and IgA were up-regulated in rNP and highest in asthmatic eosinophilic rNP among subtypes of rNP. Conclusions Our results suggest that Th17/Th1-associated mediators and BAFF may play a role and be a potential therapeutic target in refractory CRSwNP. Additionally, eosinophilic markers and autoantibodies may contribute to refractoriness in asthmatic rNP.