Neutrophilic Chronic Rhinosinusitis Research Articles

Effects of Neutrophil and Eosinophil Extracellular Trap Formation on Refractoriness in Chronic Rhinosinusitis With Nasal Polyps

This study investigated the clinical implications of neutrophil extracellular trap (NET) formation (NETosis) and eosinophil extracellular trap (EET) formation (EETosis) regarding refractoriness in chronic rhinosinusitis (CRS) with nasal polyps (CRSwNP). Nasal polyp specimens were obtained from 117 patients with CRSwNP who received endoscopic sinus surgery. Disease control status at postoperative 1 year was assessed. Refractory cases were defined as partly controlled or uncontrolled cases according to the EPOS 2020 guidelines. NETosis and EETosis were evaluated through immunofluorescence staining (citrullinated histone H3-human neutrophil elastase and citrullinated histone-galectin-10, respectively) followed by manual counting. The z-score of NET and EET counts was used to define the following four groups: low extracellular trap formation (ETosis), NETosis-predominant, EETosis-predominant, and high-ETosis. The refractory and non-refractory groups showed significant differences in the tissue eosinophil count (P = 0.005) and EET count (P = 0.029). The tissue neutrophil count and the NET/neutrophil ratio were significantly different between the refractory and non-refractory groups of patients with neutrophilic CRS (P = 0.045, 0.031, respectively). Refractoriness significantly differed among the low-ETosis (30.77%), NETosis-predominant (47.83%), EETosis-predominant (56.67%), and high-ETosis (83.33%) groups (P = 0.005). The results of this study suggest that tissue Eosinophilia and EETosis may play a prognostic role, primarily in CRSwNP and thattissue neutrophilia and NETosis can play as prognostic biomarkers in neutrophilic CRSwNP.

TNF-α Regulates the Glucocorticoid Receptor Alpha Expression in Human Nasal Epithelial Cells Via p65-NF-κb and p38-MAPK Signaling Pathways.

Tumor necrosis factor (TNF)-α induces changes in the glucocorticoid receptor (GR) isoforms' expression in human nasal epithelial cells (HNECs) in chronic rhinosinusitis (CRS). However, the underlying mechanism of TNF-α induced GR isoforms' expression in HNECs remains unclear. Here, we explored changes in inflammatory cytokines and glucocorticoid receptor alpha isoform (GRα) expression in HNECs. To explore the expression of TNF-α in nasal polyps and nasal mucosa of CRS, fluorescence immunohistochemical analysis was employed. To investigate changes in inflammatory cytokines and GRα expression in HNECs, RT-PCR and western blotting were performed following the cells' incubation with TNF-α. Cells were pretreated with the nuclear factor-κB gene binding (NF-κB) inhibitor QNZ, the p38 inhibitor SB203580, and dexamethasone for one hour, then a TNF-α. Western blotting, RT-PCR, and immunofluorescence had been utilized for the cells' analysis and the ANOVA for the data analysis. The TNF-α fluorescence intensity was mainly distributed in nasal epithelial cells of nasal tissues. TNF-α prominently inhibited the expression of GRα mRNA from 6 to 24 h in HNECs. GRα protein was decreased from 12 to 24 h. Treatment with QNZ, SB203580, or dexamethasone inhibited the TNF-α and interleukin (IL)-6 mRNA expression and increased the GRα levels. TNF-α induced changes in the GR isoforms' expression in HNECs, and it was mediated through p65-NF-κB and p38-MAPK signal transduction pathways, which could be considered a promising neutrophilic CRS treatment.

Effect of Local Corticosteroid Administration on CD8+CD25+Foxp3+ Tregs in Neutrophilic CRSwNP

Introduction: CD8⁺CD25⁺Foxp3⁺ regulatory T cells (Tregs) play an important role in human’s immune tolerance. The study was aimed to assess the influence of budesonide nasal spray on CD8⁺CD25⁺Foxp3⁺ Tregs and to evaluate their cellular functions in neutrophilic chronic rhinosinusitis with nasal polyps (CRSwNPs). Methods: Fifteen patients with neutrophilic CRSwNPs were enrolled and received physiological saline or budesonide nasal spray treatment (Saline or Budesonide group) for 3 months. Nasal tissue samples were obtained from normal subjects or those patients and cultured in vitro. CD8⁺CD25⁺Foxp3⁺ Tregs were separated from normal or NP tissues and also cultured in vitro. Then interleukin (IL)-10 and its mRNA were evaluated in the above cell cultures. The cells were applied into NP cultures. Finally, myeloperoxidase (MPO), interferon (IFN)-γ, IL-1β, and tumor necrosis factor (TNF)-α were assessed in the tissue cultures. Results: CD8⁺CD25⁺Foxp3⁺ Tregs decreased in NP tissues. Budesonide administration did not enhance the percentage of these cells in polypoid tissues. IL-10 and its mRNA were increased in the above cell cultures from NPs. However, there were no statistical differences between the two treatments in the IL-10 expression. Additionally, levels of MPO, IFN-γ, IL-1β, and TNF-α were totally elevated in NP tissue cultures and reduced after the administration of CD8⁺CD25⁺Foxp3⁺ Tregs. However, there were no significant differences in concentrations of these mediators between these two groups of the CD8⁺CD25⁺Foxp3⁺ Tregs treatment in vitro. Conclusion: The findings indicate that CD8⁺CD25⁺Foxp3⁺ Tregs might regulate the neutrophilic inflammation, and budesonide nasal spray therapy could not ameliorate the inflammation in neutrophilic CRSwNPs.

Mucosal Eosinophilia and Neutrophilia Are Not Associated With QOL or Olfactory Function in Chronic Rhinosinusitis.

Chronic rhinosinusitis (CRS) is often differentiated by histopathologic phenotypes (eosinophilic versus neutrophilic), which may impact disease severity measures and outcomes. As such, it has been suggested that counts of cellular elements be included as part of a standard pathological report following endoscopic sinus surgery (ESS). This cross-sectional study evaluated associations of mucosal eosinophilia and neutrophilia with measures of quality-of-life (QoL) and olfactory function. Patients with medically refractory CRS completed the SNOT-22 survey and Brief Smell Identification Test (BSIT) at enrollment. In addition, baseline Lund-Mackay computed tomography (CT) and Lund-Kennedy endoscopy scores were collected. Ethmoid mucosa was biopsied during ESS and reviewed using microscopy to quantify densest infiltrate of eosinophils or neutrophils per high-powered-field (HPF). Eosinophilic CRS (eCRS) and neutrophilic CRS (nCRS), both with and without nasal polyposis (NP), were compared across SNOT-22 and BSIT scores. 77/168 patients demonstrated mucosal eosinophilia (eCRS) while a total of 42/168 patients demonstrated mucosal neutrophilia (nCRS). After adjusting for polyp status, 35/168 had eCRSsNP, 42/168 eCRSwNP, 75/168 non-eCRSsNP, 16/168 non-eCRSwNP. Additionally, 22/161 were noted to have nCRSsNP, 20/161 nCRSwNP, 84/161 non-nCRSwNP, and 35/161 non-nCRSsNP. A small subset of patients demonstrated both eosinophilia and neutrophilia: 14 CRSwNP and 7 CRSsNP. When evaluating average Lund-Mackay Scores (LMS), significant differences existed between non-eCRSsNP and eCRSsNP (p = 0.006). However, after controlling for nasal polyps, eosinophilia did not significantly associate with differences in the Lund-Kennedy Score. Neutrophilia did not significantly associate with any changes in LMS or LKS after controlling for NP. Eosinophilic and neutrophilic histopathologic subtypes did not significantly associate with differences in baseline SNOT-22 or BSIT measures after controlling for NP. Neither the presence of mucosal eosinophilia nor mucosal neutrophilia demonstrated significant associations with SNOT-22 quality-of-life or BSIT olfactory function scores when controlling for comorbid nasal polyposis.

Epithelial-to-mesenchymal transition in neutrophilic chronic rhinosinusitis.

Barrier dysfunction, tissue fibrosis, and remodeling are essential processes of the pathophysiology of chronic rhinosinusitis (CRS). The role of epithelial-to-mesenchymal transition (EMT) has been assessed in various studies in CRS. In this review, we summarized the pathophysiologic mechanisms of EMT related to CRS, particularly neutrophilic CRS. Loss of epithelial characteristics due to EMT makes leaky epithelium, and transformed mesenchymal cells cause fibrosis and remodeling. Hypoxia, allergens (house dust mites), infections, and air pollutants were related to the pathogenesis of neutrophilic CRS, and these factors are known to induce barrier dysfunction and EMT in sinonasal epithelia. Some molecular pathways related to EMT have been recognized in CRS, including interferon-γ/p38/extracellular signal-regulated kinase, high-mobility group box 1/receptor of advanced glycosylation end-products, TGF-β1/SMAD, and Wnt/β-catenin-signaling pathways. Apart from, several microRNAs (miR-21, miR-761, and miR-30a-5p) have been identified to regulate EMT in CRS. EMT is considered to be an important pathogenesis mechanism for CRS. The factors cause EMT in CRS, and the associated molecular mechanisms are related to neutrophilic inflammation. Further studies on CRS endotype and/or phenotype are needed to clarify the implication of EMT on CRS pathogenesis.

Patient-reported sleep outcomes lack association with mucosal eosinophilia or neutrophilia in patients with chronic rhinosinusitis undergoing functional endoscopic sinus surgery.

Chronic rhinosinusitis (CRS) is associated with sleep dysfunction, but the underlying pathophysiology is poorly understood. The purpose of this study was to determine if mucosal eosinophilia or neutrophilia were associated with sleep dysfunction severity or altered the improvement in sleep dysfunction following functional endoscopic sinus surgery (FESS). A total of 104 patients with medically refractory CRS with nasal polyposis (CRSwNP) and CRS without nasal polyposis (CRSsNP), completed the Pittsburgh Sleep Quality Index (PSQI) before and after FESS. Anterior ethmoid mucosa was collected during FESS and densest infiltrates of eosinophilia and neutrophilia per high-power field (HPF) were determined by microscopy. Eosinophilic (>10 eosinophils/HPF) and neutrophilic (>4 neutrophils/HPF) CRS were then compared to preoperative and postoperative PSQI measures. Of 104 study participants, 88 (85%) reported preoperative PSQI scores consistent with "poor sleep," (PSQI total > 5). The cohort overall demonstrated significant improvement in poor sleep (65%; χ2 = 12.03; p < 0.001) 16.8 ± 5.0 months after FESS. Regardless of nasal polyposis, neither eosinophilic nor neutrophilic CRS was associated with differences in mean postoperative PSQI improvement. However, in patients with neutrophilic CRSsNP, there was a significant relationship between severity of neutrophilia and improvement in sleep latency (R = -0.798, p = 0.003) and sleep efficacy (R = -0.777, p = 0.005). Chronic inflammation has been hypothesized to play a pathophysiologic role in sleep dysfunction associated with CRS. This study suggests that in patients with medically refractory CRS, evidence of mucosal eosinophilia and neutrophilia lack strong associations with patient-reported sleep dysfunction or improvements in sleep quality after FESS, overall. However, neutrophilia may impact sleep latency and efficacy in patients with CRSsNP.

Efficacy of Budesonide Nasal Spray on Neutrophilic Chronic Rhinosinusitis with Nasal Polyps: A Combined Clinical and Experimental Study

Background: Neutrophilic chronic rhinosinusitis with nasal polyps (CRSwNP) occur predominantly in Asian subjects. Appropriate treatments for this endotype have not been elucidated. This study aimed to evaluate the efficacy of budesonide nasal spray on neutrophilic CRSwNP. Materials and Methods: Fifteen neutrophilic CRSwNP patients were included, and then they received budesonide nasal spray treatment for 3 months. Biopsies of nasal polyps (NPs) were obtained from these subjects. Their clinical indexes were scored using Visual Analog Scale (VAS), Sino-Nasal Outcome Test (SNOT)-22, and Endoscopic Appearances (EAs). Histological analyses were used to assess numbers of neutrophils, goblet cells, and submucosal gland cells in NPs. Percentages of CD8⁺ T cells and CD4⁺CD25⁺Foxp3⁺ regulatory T cells (Tregs) were evaluated using flow cytometry. Mucin 5AC (MUC5AC), MUC5B, myeloperoxidase (MPO), interferon (IFN)-γ, and interleukin (IL)-1β and their mRNAs were also examined. After that, we cultured NP tissues in vitro and evaluated the abovementioned inflammatory parameters before and after the administration of budesonide. Results: Budesonide nasal spray did not improve clinical evaluations including VAS, SNOT-22, and EA scores. Numbers of neutrophils and goblet cells, the score of submucosal gland cells, percentages of CD8⁺ T cells and Tregs, MUC5AC, MUC5B, MPO, IFN-γ, and IL-1β and their mRNAs were not decreased in NPs after the budesonide treatment. Furthermore, the administration of budesonide into NP cultures also did not reduce their levels in comparison with those before the treatment. Conclusion: These findings demonstrate that budesonide treatment may not alleviate the inflammatory condition in neutrophilic CRSwNP.

Activated pyrin domain containing 3 (NLRP3) inflammasome in neutrophilic chronic rhinosinusitis with nasal polyps (CRSwNP)

NLRP3 inflammasome and IL-1β were significantly correlated and upregulated in high neutrophil infiltrated CRSwNP. The antigen and proinflammatory cytokines chemotaxis neutrophil infiltration by the NLRP3-IL-1β axis in polyp tissues, although glucocorticoid treatment alleviates it.

Elastase-Positive Neutrophils Are Associated With Refractoriness of Chronic Rhinosinusitis With Nasal Polyps in an Asian Population.

PurposeVarious 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.MethodsProtein 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 (Neuhigh) 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 (Neulow).ResultsIn terms of disease control status, NENP-Neulow patients showed the higher rate of disease control than NENP-Neuhigh and ENP-Neuhigh patients. Linear by linear association demonstrated the trend in refractoriness from NENP-Neulow to NENP-Neuhigh or ENP-Neulow to ENP-Neuhigh. 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.ConclusionsTissue neutrophilia had an influence on treatment outcomes in the Asian CRSwNP patients. HNE-positive cells and IL-36α may be biomarkers for predicting refractoriness in Asians with CRSwNP. Additionally, imbalances in HNE and α1 anti-trypsin may be associated with pathophysiology of neutrophilic chronic rhinosinusitis.

The IFN-\u03b3\u2013p38, ERK kinase axis exacerbates neutrophilic chronic rhinosinusitis by inducing the epithelial-to-mesenchymal transition

Chronic rhinosinusitis (CRS) is a heterogeneous and multifactorial inflammatory disease characterized by involvement of diverse types of inflammatory cells. Asian CRS patients frequently show infiltration of neutrophils and an elevated level of interferon (IFN)-γ; by contrast, western patients exhibit eosinophil infiltration and enhanced levels of Th2-related cytokines. Neutrophilia in tissues decreases sensitivity to corticosteroids, but the mechanisms underlying the progression of neutrophilic CRS are unclear. In this study, we investigated the role of IFN-γ in CRS patients with marked neutrophil infiltration. We report that the IFN-γ level is upregulated in the tissues of these patients, particularly those with non-eosinophilic nasal polyps. The level of IFN-γ was significantly correlated with markers of the epithelial-to-mesenchymal transition (EMT). We further demonstrated that IFN-γ induced the EMT via the p38 and extracellular signal-regulated kinase (ERK) pathways in a manner distinct from the hypoxia-inducible factor (HIF)-1α, SMAD, and NF-κB signaling pathways. In a murine nasal polyp (NP) model, blocking the p38 and ERK signaling pathways prevented NP formation and chemotactic cytokine secretion by neutrophils but not eosinophils. Taken together, our results suggest that IFN-γ can induce the EMT in nasal epithelial cells, and thus blocking the p38 and ERK pathways could be an effective therapeutic strategy against neutrophil-dominant CRS.

Activations of group 2 innate lymphoid cells depend on endotypes of chronic rhinosinusitis.

Chronic rhinosinusitis (CRS) is a complicated disease with several variants caused by different cellular and molecular mechanisms. The characterization of this heterogeneity supports the definition that the disease consists of many endotypes, such as eosinophilic and neutrophilic CRS, and so on. This study aimed to explore group 2 innate lymphoid cells (ILC2s) in neutrophilic CRS without nasal polyps (CRSsNP) and with nasal polyps (CRSwNP), and evaluate ILC2s across characteristics of the disease. Nasal biopsy samples were obtained from normal subjects or subjects with CRSsNP or CRSwNP during surgery. ILC2s were sorted and purified as CD45+Lin-CD127+CD4-CD8-CRTH2+CD161+ cells through flow cytometry, and were compared among three groups of subjects. Then, these samples were cultured in vitro, and inflammatory factors were assessed in tissue cultures. After that, human recombinant (rm) interleukin (IL)-33 or IL-17 were administered into the cultures, and we again examined relevant inflammatory substances. ILC2s were upregulated in neutrophilic CRSsNP and CRSwNP patients, and there were no statistical differences between them. Eosinophil cation protein (ECP), myeloperoxidase (MPO), IL-25, IL-33, IL-5, IL-13, interferon (IFN)-γ and IL-17 were increased in the cultures, however, only concentrations of MPO, IFN-γ and IL-17 were enhanced in CRSwNP tissues compared to CRSsNP ones. After administration of rmIL-33, ECP, IL-5 and IL-13 were all increased in tissues from CRSsNP and CRSwNP patients, however, there were no significant differences between them. Finally, we evaluated concentrations of several above inflammatory factors after the treatment of rmIL-17, and found that MPO and IFN-γ were enhanced in these two phenotypes of patients, and were elevated significantly in CRSwNP tissue cultures. These findings show that ILC2s might be inactivated in neutrophilic CRSsNP and CRSwNP based on this pilot study.

Zellzahl und Entzündungsparameter im peripheren Blut bei chronischer Rhinosinusitis

The aim of this study was to verify whether the following peripheral blood cell count and inflammation-based markers differ between various types of chronic rhinosinusitis (CRS): neutrophil-to-lymphocyte ratio (NLR), monocyte-to-lymphocyte ratio (MLR) and platelet-to-lymphocyte ratio (PLR), as well as erythrocyte sedimentation rate (ESR) and C‑reactive protein (CRP) levels. In all, 386 patients had complete peripheral blood count, ESR, CRP and nasal cytology. The severity of CRS symptoms was assessed using three-stage Lund-Mackay computed tomography (CT) scores. The participants were stratified based on the results of nasal cytology and by the presence of nasal polyps (NP). The inflammation-based markers were calculated by dividing the cell numbers of the different cell types by numbers of the other cell types. Blood leukocyte and neutrophil counts were higher in neutrophilic CRS. Differences between patients with CRS with nasal polyps (NP) and CRS without NP were significant for blood leukocytes, neutrophils, monocytes, eosinophils, CRP, NLR and MLR values. In CRS with NP, peripheral blood leukocyte, neutrophil, monocyte and eosinophil counts, as well as CRP, NLR and MLR values were higher than in CRS without NP. Our results show that all individuals with CRS could benefit from the analysis of blood counts and inflammation-based markers at the beginning of their evaluation. High levels of inflammation-based markers might guide the clinician towards planning more radical CRS therapy and use of systemic anti-inflammatory medication.

Comparison of bacterial examinations between eosinophilic and neutrophilic chronic rhinosinusitis with nasal polyps

Conclusion: We found no significant differences in the bacterial features of the maxillary sinuses between eosinophilic and neutrophilic chronic rhinosinusitis (CRS) with nasal polyps. Objectives: Since neutrophilic CRS is often influenced by a predisposition to bacterial infection, and eosinophilic CRS is likely to be developed by allergic antigens, differences in the microbiology between the two pathologies of CRS can be expected. The present study was designed to investigate the bacterial findings from the maxillary sinus in eosinophilic and neutrophilic CRS. Methods: Seventy patients with CRS with nasal polyps were divided into eosinophilic and neutrophilic types based on histopathological observations of the nasal polyps. The specimens for bacterial culture were obtained from the maxillary sinus during endoscopic sinus surgery. Results: In all, 29 and 41 patients were classified as having eosinophilic and neutrophilic CRS with nasal polyps, respectively. The isolation rate of bacteria showed no significant difference between eosinophilic (90%) and neutrophilic CRS (98%). Aerobic bacteria were found in 25 patients (86%) with eosinophilic CRS, which was not significantly different from that in neutrophilic CRS (40 patients, 98%). The isolation rate for aerobic and anaerobic bacteria showed no significant differences.