Deep immune profiling of chronic rhinosinusitis in allergic and non-allergic cohorts using mass cytometry.

Chronic rhinosinusitis (CRS) is a persistent nasal and paranasal sinus mucosa inflammation comprising two phenotypes, namely CRS with nasal polyps (CRSwNP) and without (CRSsNP). CRSwNP can be associated with asthma and hypersensitivity to non-steroidal anti-inflammatory drug (NSAID) in a syndrome known as NSAID-exacerbated respiratory disease (N-ERD). Furthermore, CRS frequently intertwines with respiratory allergies. This study investigated levels of 33 different nasal and serum cytokines and phenotypic characteristics of peripheral blood mononuclear cells (PBMCs) within cohorts of CRS patients (n = 24), additionally examining the influence of comorbid respiratory allergies by mass cytometry. N-ERD patients showed heightened type 2 nasal cytokine levels. Mass cytometry revealed increased activated naive B cell levels in CRSwNP and N-ERD, while resting naive B cells were higher in CRSsNP. Th2a cell levels were significantly elevated in allergic subjects, but not in CRS groups. In conclusion, there are distinct immunological features in PBMCs of CRS phenotypes and allergy.

Nasal IL-13 production identifies patients with late-phase allergic responses.

BACKGROUND: There is limited knowledge on how local cytokine secretion patterns after nasal allergen challenge correlate with clinical symptoms especially with regard to the "late allergic response," which occurs in approximately 40% to 50% of patients with allergy. OBJECTIVE: We sought to characterize the immunologic and clinical nasal responses to birch pollen allergen challenge with a special focus on the late allergic response. METHODS: In this randomized, double-blind, placebo-controlled trial, birch pollen-allergic participants were challenged with birch pollen extract (n = 20) or placebo (n = 10) on 3 consecutive days. On days 1 and 3, nasal secretions were collected at selected time points over a 24-hour time course for the measurement of 33 inflammatory mediators. Clinical responses were determined through subjective symptom scores and objective nasal airflow measurements. RESULTS: Provoked participants had significantly greater clinical responses and showed significant increases in tryptase and the soluble IL-33 receptor serum stimulation 2 (sST2) in nasal secretions within minutes compared with the placebo group. Eight of 20 provoked participants displayed high IL-13 levels 2 to 8 hours after allergen provocation. This group also showed significant changes in clinical parameters, with a secondary drop in nasal airflow measured by peak nasal inspiratory flow and increased symptoms of nasal obstruction, which significantly differed from IL-13 nonresponders after 6 hours. CONCLUSIONS: IL-13 response status correlates with clinical responses and type 2 cytokine responses in the late phase after allergen provocation.

Dupilumab increases aspirin tolerance in NSAID-exacerbated respiratory disease.

BACKGROUND: Nonsteroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (N-ERD) comprises the triad of chronic rhinosinusitis with nasal polyps, asthma and intolerance to NSAIDs. Dupilumab treatment, targeting the interleukin-4 (IL-4) receptor α, significantly reduces polyp burden as well as asthma symptoms. Here we aimed to investigate the effect of dupilumab on aspirin intolerance, burden of disease and nasal cytokine profiles in patients with N-ERD. METHODS: In this open-label trial, adult patients with confirmed N-ERD were treated with dupilumab for 6 months. Clinical parameters (e.g. total polyp scores, quality of life questionnaires, smell test, spirometry), oral aspirin provocation testing and blood, nasal and urine sampling were monitored at regular intervals for up to 6 months after starting dupilumab therapy. RESULTS: Of the 31 patients included in the study, 30 completed both aspirin provocation tests. After 6 months of treatment with dupilumab, 23% of patients (n=7 of 30) developed complete aspirin tolerance and an additional 33% of patients (n=10 of 30) tolerated higher doses. Polyp burden was significantly reduced (total polyp score: -2.68±1.84, p<0.001), while pulmonary symptoms (asthma control test: +2.34±3.67, p<0.001) and olfactory performance improved (University of Pennsylvania Smell Identification Test: +11.16±9.54, p<0.001) in all patients after therapy. Patients with increased aspirin tolerance showed a significant decrease in urinary leukotriene E4 levels and their improvement in clinical parameters was associated with a reduction of eotaxin-1, C-C motif chemokine ligand 17, IL-5, IL-17A and IL-6. CONCLUSION: In this study, 57% of N-ERD patients tolerated higher doses of aspirin under dupilumab therapy.

Radiosensitizing effect of galunisertib, a TGF-ß receptor I inhibitor, on head and neck squamous cell carcinoma in vitro.

BACKGROUND: Resistance to radiation therapy poses a major clinical problem for patients suffering from head and neck squamous cell carcinoma (HNSCC). Transforming growth factor ß (TGF-ß) has emerged as a potential target. This study aimed to investigate the radiosensitizing effect of galunisertib, a small molecule TGF-ß receptor kinase I inhibitor, on HNSCC cells in vitro. METHODS: Three HNSCC cell lines were treated with galunisertib alone, or in combination with radiation. Of those three cell lines, one has a known inactivating mutation of the TGF-ß pathway (Cal27), one has a TGF-ß pathway deficiency (FaDu) and one has no known alteration (SCC-25). The effect on metabolic activity was evaluated by a resazurin-based reduction assay. Cell migration was evaluated by wound-healing assay, clonogenic survival by colony formation assay and cell cycle by FACS analysis. RESULTS: Galunisertib reduced metabolic activity in FaDu, increased in SCC-25 and had no effect on CAL27. Migration was significantly reduced by galunisertib in all three cell lines and showed additive effects in combination with radiation in CAL27 and SCC-25. Colony-forming capabilities were reduced in SCC-25 by galunisertib and also showed an additive effect with adjuvant radiation treatment. Cell cycle analysis showed a reduction of cells in G1 phase in response to galunisertib treatment. CONCLUSION: Our results indicate a potential antineoplastic effect of galunisertib in HNSCC with intact TGF-ß signaling in combination with radiation.

Differences in men and women suffering from CRSwNP and AERD in quality of life.

PURPOSE: While the overall impact of chronic rhinosinusitis (CRS) on patients' health is diverse, many affected individuals have a substantially impaired quality of life (QoL). The aim of this study was to evaluate the impact of sex-associated differences specifically in the subgroups of CRS with nasal polyps (CRSwNP) and aspirin-exacerbated respiratory disease (AERD) by assessing QoL parameters in women and men separately. METHODS: In a retrospective single-center study, 59 patients with CRSwNP (39 males and 20 females) and 46 patients with AERD (18 males and 28 females) were included. Patient-reported outcome measures (PROM) evaluating QoL via the Sino-Nasal Outcome Test-20 German Adapted Version (SNOT-20 GAV) as well as the total polyp score (TPS) were analysed. RESULTS: There was no significant difference in TPS (p = 0.5550) and total SNOT-20 GAV scores (p = 0.0726) between male or female patients with CRSwNP or AERD. Furthermore, no significant sex differences were found within disease groups regarding the subcategories of the SNOT-20 GAV items. CONCLUSION: Thus, quality of life is severely impaired in patients suffering from various forms of CRS regardless of their sex.

Allergen-specific IgE levels and the ability of IgE-allergen complexes to cross-link determine the extent of CD23-mediated T-cell activation.

BACKGROUND: CD23 mediates IgE-facilitated allergen presentation and subsequent allergen-specific T-cell activation in allergic patients. OBJECTIVE: We sought to investigate key factors regulating IgE-facilitated allergen presentation through CD23 and subsequent T-cell activation. METHODS: To study T-cell activation by free allergens and different types of IgE-Bet v 1 complexes, we used a molecular model based on monoclonal human Bet v 1-specific IgE, monomeric and oligomeric Bet v 1 allergen, an MHC-matched CD23-expressing B-cell line, and a T-cell line expressing a human Bet v 1-specific T-cell receptor. The ability to cross-link Fcε receptors of complexes consisting of either IgE and monomeric Bet v 1 or IgE and oligomeric Bet v 1 was studied in human FcεRI-expressing basophils. T-cell proliferation by monomeric or oligomeric Bet v 1, which cross-links Fcε receptors to a different extent, was studied in allergic patients' PBMCs with and without CD23-expressing B cells. RESULTS: In our model non-cross-linking IgE-Bet v 1 monomer complexes, as well as cross-linking IgE-Bet v 1 oligomer complexes, induced T-cell activation, which was dependent on the concentration of specific IgE. However, T-cell activation by cross-linking IgE-Bet v 1 oligomer complexes was approximately 125-fold more efficient. Relevant T-cell proliferation occurred in allergic patients' PBMCs only in the presence of B cells, and its magnitude depended on the ability of IgE-Bet v 1 complexes to cross-link CD23. CONCLUSION: The extent of CD23-mediated T-cell activation depends on the concentration of allergen-specific IgE and the cross-linking ability of IgE-allergen complexes.

Real-Life Study of Patient Preference for Dupilumab or Revision Surgery for Recurrent Chronic Rhinosinusitis with Nasal Polyps.

(1) Background: Chronic rhinosinusitis with nasal polyps (CRSwNP) has a high rate of recurrence in patients, despite therapy with local corticosteroids and functional endoscopic sinus surgery. Dupilumab, a recombinant monoclonal human IgG4 antibody directed against the IL-4 receptor α that inhibits both IL-4 and IL-13 signal transduction, is available for symptomatic therapy. Patient preference between repeated surgery and injection therapy with Dupilumab is not known. (2) Methods: Patients who had experienced at least one surgical intervention for nasal polyps and were treated with Dupilumab for at least 3 months completed a retrospective patient questionnaire. (3) Results: In a cohort of 75 previously operated CRSwNP patients, 91.5% preferred therapy with Dupilumab to repeated surgery for nasal polyps. Preference for Dupilumab in the subgroups of patients with concomitant Non-steroidal Anti-inflammatory Drugs Exacerbated Respiratory Disease (N-ERD) (n = 32), patients with concomitant asthma (n = 25), and patients without concomitant disease (n = 18) was 100%, 96%, and 72%, respectively. (4) Conclusions: Patient preference for Dupilumab over repeat surgery is strongest in previously operated CRSwNP patients with concomitant asthma or N-ERD, but remains very high in patients without concomitant disease.

Analysis of cosmetic results of metopic synostosis: concordance and interobserver variability.

Trigonocephaly is caused by premature fusion of the metopic suture of the calvarium. The resultant facial and skull deformities place a newborn at a higher risk of cerebral impairment. Frontal orbital advancement and remodeling is the accepted surgical management. An analysis was undertaken to assess the cosmetic results for nonsyndromic metopic synostosis following frontal orbital advancement and remodeling and to determine whether the observer technique used had validity.The photographs of 20 consecutive patients with metopic synostosis were assessed preoperatively, postoperatively, and at 3 and 5 years of age. Each photograph was analyzed by 2 pairs of trained rotating observers. The findings were compared and reviewed by the fifth member, who acted as a "referee." Where there were disagreements between the findings, the referee came to a final decision. The presence and severity of trigonocephaly, temporal hollowing, hypotelorism, orbital asymmetry, epicanthic folds, and scarring were assessed. κ Analysis was used to determine the validity of observer concordance when using photographic material.There was a significant overall trend in craniofacial aesthetic improvement with time. However, 12% had some persistence of trigonocephaly. Temporal hollowing was usually persistent. κ Statistics varied among attributes and time period assessed. The greatest concordance was for improvement of epicanthic folds and trigonocephaly, with observers agreeing 84.1% and 82.6% of the time, respectively. There was least concordance for severity of hypotelorism and scarring.This study suggests that observer analysis of photographic records is a valid outcome measure for cosmesis, although it yields variable interobserver concordance according to the feature assessed. Surgery corrected most but not all the features of trigonocephaly. The prognostic indicators for good cosmetic outcome are presented.

Platelet-to-lymphocyte ratio might predict the response to dupilumab treatment for patients with nasal polyposis.

BACKGROUND: Dupilumab is a monoclonal antibody against interleukin 4 receptor alpha and has proven to be clinically effective in treating patients with chronic rhinosinusitis with nasal polyps (CRSwNP). However, a certain number of patients are non- or partial responders. This study aims to investigate the relevance of inflammatory markers with regard to therapy response to dupilumab in CRSwNP patients. METHODS: All patients with CRSwNP treated with dupilumab at a tertiary healthcare center with available pretreatment inflammatory markers were included. The values of pretreatment neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) were associated with the outcome. Patients were stratified according to the respective median value (> median was considered high). The binary logistic regression was performed with regard to total treatment response (post-treatment total nasal polyp score (NPS) 0). RESULTS: A total of 65 CRSwNP patients with available pretreatment peripheral blood values were included in the study. The mean pre- and post-treatment total NPS values were 4.3 ± 1.9 and 1.2 ± 1.6, respectively. High PLR (> 131.2) was independently associated with a 3.9-fold higher probability of reaching the NPS value of 0 in the multivariable analysis. On the other hand, High NLR (> 1.9) did not significantly associate with the outcome. CONCLUSIONS: The current study provides insights into the potential positive predictive value of the high PLR (> 131.2) in CRSwNP patients regarding treatment with dupilumab. There is a need for further prospective studies for validation of these results, especially in cohorts of patients with severe CRSwNP.

Low free triiodothyronine and immune-related hyperthyroidism are associated with overall and progression-free survival in head and neck squamous cell carcinoma treated with pembrolizumab.

INTRODUCTION: Thyroid function is frequently impaired in recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC). In patients treated with pembrolizumab, immune-related adverse events (irAEs) of the thyroid are common. However, the prognostic significance of baseline and on-treatment thyroid dysfunction is currently unclear. METHODS: This study included 95 patients who received pembrolizumab for R/M HNSCC between 2016 and 2022. Baseline thyroid status, according to serum hormone levels, and irAEs were assessed. Univariable and multivariable Cox regression analyses were performed for overall survival (OS) and progression-free survival (PFS). Furthermore, the best overall response according to the prognostic groups was examined. RESULTS: Low fT3 (HR: 2.52, p = 0.006), immune-related hyperthyroidism (HR: 0.11, p = 0.038), ECOG performance status ≥2 (HR: 3.72, p = 0.002), and platinum-refractory disease (HR: 3.29, p = 0.020) were independently associated with OS. Furthermore, immune-related hyperthyroidism was associated with longer PFS (HR: 0.13, p = 0.007), a higher objective response rate (83% vs. 31%, p = 0.018), and a higher disease control rate (100% vs. 43%, p = 0.008). Thyroid-related autoantibodies were elevated in 40% of thyroid irAEs cases with available measurements. Out of 16 thyroid irAEs, 15 occurred in patients with fT3 above the lower limit of normal. CONCLUSION: Low fT3 was associated with worse OS. Immune-related hyperthyroidism was correlated with both improved OS and PFS. Baseline fT3 assessment and close on-treatment monitoring of serum thyroid levels may be valuable for risk stratification in R/M HNSCC patients receiving pembrolizumab.

The "real life" efficacy of dupilumab is independent of initial polyp size and concomitant steroids in CRSwNP.

BACKGROUND: Dupilumab significantly improves symptom control in chronic rhinosinusitis with nasal polyps (CRSwNP). Patients with large polyps at the initiation of treatment (total polyp score (TPS) ≥ 5) have been the focus in published studies. Patients with significant burden of disease but small polyps (TPS ≤ 4) have not yet been evaluated for clinical response. This study set out to evaluate the benefit of dupilumab treatment on cohorts of small (TPS ≤ 4) compared to large polyps (TPS ≥ 5). Furthermore, benefit of concomitant oral and/or nasal steroid therapy has been evaluated. METHODS: 97 patients with CRSwNP, who were begun on dupilumab between January 2020 and October 2021, were included. All patients were followed-up for 6 months. At each visit they underwent nasal endoscopy, smell identification tests and filled out validated patient questionnaires. RESULTS: Significant drops in TPS were seen in both patient groups after 6 months of therapy, dropping from a median score of 3 to 0 and from 6 to 2 in patients with small and large polyps respectively. Furthermore, a linear mixed model calculated a drop of 22% and 24% in TPS per month in patients with small and large polyps respectively with no significant difference in rate of decline. Finally the model showed that neither oral nor nasal steroids influenced the rate of response to dupilumab therapy. CONCLUSIONS: Polyp size at the initiation of dupilumab therapy and whether patients continue to take steroid therapy does not appear to influence effectiveness of dupilumab treatment.

The nasal microbiome in patients suffering from non-steroidal anti-inflammatory drugs-exacerbated respiratory disease in absence of corticosteroids.

Chronic rhinosinusitis (CRS) is a chronic inflammatory disease phenotypically classified by the absence (CRSsNP) or presence of nasal polyps (CRSwNP). The latter may also be associated with asthma and hypersensitivity towards non-steroidal anti-inflammatory drugs (NSAID) as a triad termed NSAID-exacerbated respiratory disease (N-ERD). The role of the microbiome in these different disease entities with regard to the underlying inflammatory process and disease burden is yet not fully understood. To address this question, we measured clinical parameters and collected nasal samples (nasal mucosal fluids, microbiome swabs from middle meatus and anterior naris) of patients suffering from CRSsNP (n=20), CRSwNP (n=20) or N-ERD (n=20) as well as from patients without CRS (=disease controls, n=20). Importantly, all subjects refrained from taking local or systemic corticosteroids or immunosuppressants for at least two weeks prior to sampling. The nasal microbiome was analyzed using 16S rRNA gene amplicon sequencing, and levels of 33 inflammatory cytokines were determined in nasal mucosal fluids using the MSD platform. Patients suffering from N-ERD and CRSwNP showed significantly worse smell perception and significantly higher levels of type 2 associated cytokines IL-5, IL-9, Eotaxin and CCL17. Across all 4 patient groups, Corynebacteria and Staphylococci showed the highest relative abundances. Although no significant difference in alpha and beta diversity was observed between the control and the CRS groups, pairwise testing revealed a higher relative abundance of Staphylococci in the middle meatus in N-ERD patients as compared to CRSwNP (p<0.001), CRSsNP (p<0.01) and disease controls (p<0.05) and of Lawsonella in patients suffering from CRSwNP in middle meatus and anterior naris in comparison to CRSsNP (p<0.0001 for both locations) and disease controls (p<0.01 and p<0.0001). Furthermore, we observed a positive correlation of Staphylococci with IL-5 (Pearson r=0.548) and a negative correlation for Corynebacteria and Eotaxin-3 (r=-0.540). Thus, in patients refraining from oral and nasal corticosteroid therapy for at least two weeks known to alter microbiome composition, we did not observe differences in microbiome alpha or beta diversity between various CRS entities and disease controls. However, our data suggest a close association between increased bacterial colonization with Staphylococci and decreased colonization by Corynebacteria as well as increased type 2 inflammation.

Comprehensive Analysis of Nasal Polyps Reveals a More Pronounced Type 2 Transcriptomic Profile of Epithelial Cells and Mast Cells in Aspirin-Exacerbated Respiratory Disease.

Chronic rhinosinusitis with nasal polyps is affecting up to 3% of Western populations. About 10% of patients with nasal polyps also suffer from asthma and intolerance to aspirin, a syndrome called aspirin-exacerbated respiratory disease. Although eosinophilic inflammation is predominant in polyps of both diseases, phenotypic differences in the tissue-derived microenvironment, elucidating disease-specific characteristics, have not yet been identified. We sought to obtain detailed information about phenotypic and transcriptional differences in epithelial and immune cells in polyps of aspirin-tolerant and intolerant patients. Cytokine profiles in nasal secretions and serum of patients suffering from aspirin-exacerbated respiratory disease (n = 10) or chronic rhinosinusitis with nasal polyps (n = 9) were assessed using a multiplex mesoscale discovery assay. After enrichment for immune cell subsets by flow cytometry, we performed transcriptomic profiling by employing single-cell RNA sequencing. Aspirin-intolerant patients displayed significantly elevated IL-5 and CCL17 levels in nasal secretions corresponding to a more pronounced eosinophilic type 2 inflammation. Transcriptomic profiling revealed that epithelial and mast cells not only complement one another in terms of gene expression associated with the 15-lipoxygenase pathway but also show a clear type 2-associated inflammatory phenotype as identified by the upregulation of POSTN, CCL26, and IL13 in patients with aspirin-exacerbated respiratory disease. Interestingly, we also observed cellular stress responses indicated by an increase of MTRNR2L12, MTRNR2L8, and NEAT1 across all immune cell subsets in this disease entity. In conclusion, our findings support the hypothesis that epithelial and mast cells act in concert as potential drivers of the pathogenesis of the aspirin-exacerbated respiratory disease.

Chronic rhinosinusitis symptoms differentially impact the likelihood of major depressive disorders.

OBJECTIVES: The extent to which sinonasal symptoms impact the likelihood of major depressive disorders in chronic rhinosinusitis patients with nasal polyposis (CRSwNP) remains incompletely characterized. In this study, we sought to determine whether individual symptom clusters differentially impact the likelihood of depression in a cohort of CRSwNP patients. METHODS: We retrospectively included 77 patients with CRSwNP. The severity of sinonasal symptoms was assessed using the 22-item Sino-Nasal Outcome Test (SNOT-22) and grouped according to a previously validated four-subdomain structure: nasal, otologic/facial pain, sleep, and emotional subdomains. The likelihood of major depressive disorders was assessed using the Patient Health Questionnaire-2 (PHQ-2). The clinical characteristic of symptom severity (nasal polyp size) and disease-specific information, such as the number of previous sinonasal surgeries, were also collected. RESULTS: The sleep subdomain was most strongly associated with the likelihood of major depressive disorders, followed by the otologic/facial pain subdomain, after controlling for demographics and clinical indicators of symptom severity (nasal polyp size). We found a SNOT-22 score ≥ 30.5 to be an accurate indicator of scoring higher than or equal to 2 on the PHQ-2 in CRSwNP patients. This had a sensitivity of 83.33% and a specificity of 75.47%. CONCLUSION: Distinct sinonasal symptom clusters differentially impact the likelihood of depression in CRSwNP patients. Raising awareness for those with severe sinonasal symptomatology might help identify more patients with a higher probability of comorbid depression.Level of Evidence: 4.

Changes in Acoustic Aspects of Vocal Function in Children After Adenotonsillectomy.

BACKGROUND: Adenotonsillectomy is one of the most common pediatric surgical procedures. Postoperative voice changes are a very common concern among patient's parents. Therefore, the aim of this study is to analyze acoustic voice parameters after adenotonsillectomy, tonsillectomy, and adenoidectomy in pediatric patients in a tertiary referral academic center. PATIENTS AND METHODS: All pediatric patients undergoing an adenotonsillectomy, tonsillectomy or adenoidectomy in a single center from 2002 to 2018 were included in the study. Change of fundamental frequency, jitter, shimmer, and harmonic-noise ratio at first, seventh and 30th postoperative day compared to preoperative values were the primary outcome parameters. Statistical analysis was performed using repeated measures analysis of variance model. RESULTS: A total of 1258 patients were included in the study. The mean age of patients at the time of surgery was 8.3 years (range 3.0-18.0 years). Around 698 were male (55.5%) and 560 female (44.5%). The values of fundamental frequency increased significantly after the first and seventh postoperative day (P = 0.001 both) but normalized 1 month after surgery (P = 0.962). At the first postoperative month, values of jitter and shimmer decreased significantly (P = 0.005 and P = 0.002, respectively). Measurements of harmonic-noise ratio revealed a significant increase 30 days after surgery (P = 0.004). CONCLUSION: Statistically significant differences in objective voice parameters within the first postoperative month after tonsillectomy, adenoidectomy, and adenotonsillectomy were observed. The fundamental frequency returned to normal 1 month after surgery. These findings can contribute in soothing the concerns of parents regarding postoperative voice changes.

Using mass cytometry for the analysis of samples of the human airways.

Mass cytometry (MC) is a powerful method for mapping complex cellular systems at single-cell levels, based on the detection of cellular proteins. Numerous studies have been performed using human blood, but there is a lack of protocols describing the processing and labeling of bronchoalveolar lavage fluid (BALF) and nasal polyps (NP) for acquisition by MC. These specimens are essential in the investigation of immune cell characteristics in airway diseases such as asthma and chronic rhinosinusitis with NP (CRSwNP). Here we optimized a workflow for processing, labeling, and acquisition of BALF and NP cells by MC. Among three methods tested for NP digestion, combined enzymatic/mechanical processing yielded maximum cell recovery, viability and labeling patterns compared to the other methods. Treatment with DNAse improved sample acquisition by MC. In a final step, we performed a comparison of blood, BALF and NP cell composition using a 31-marker MC antibody panel, revealing expected differences between the different tissue but also heterogeneity among the BALF and NP samples. We here introduce an optimized workflow for the MC analysis of human NP and BALF, which enables comparative analysis of different samples in larger cohorts. A deeper understanding of immune cell characteristics in these samples may guide future researchers and clinicians to a better disease management.

The molecular march of primary and recurrent nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) results from the aberrant and uncontrolled growth of the nasopharyngeal epithelium. It is highly associated with the Epstein-Barr virus, especially in regions where it is endemic. In the last decade, significant advances in genetic sequencing techniques have allowed the discovery of many new abnormal molecular processes that undoubtedly contribute to the establishment, growth and spread of this deadly disease. In this review, we consider NPC as EBV induced. We summarise the recent discoveries and how they add to our understanding of the pathophysiology of NPC in the context of genomics first in primary and then in recurrent disease. Overall, we find key early events lead to p16 inactivation and cyclin D1 expression, allowing latent viral infection. Host and viral factors work together to affect a variety of molecular pathways, the most fundamental being activation of NF-κB. Nonetheless, much still yearns to be discovered, especially in recurrent NPC.

Prevalence and Symptom Burden of Nasal Polyps in a Large Austrian Population.

BACKGROUND: Although chronic rhinosinusitis with nasal polyps is a common inflammatory condition with significant morbidity and financial cost, information regarding prevalence and disease burden of this condition is scarce. OBJECTIVE: In this study, we determined nasal polyp prevalence, polyp grade, concomitant disease, and symptom burden in more than 10,000 central European subjects. METHODS: In this retrospective, cross-sectional study, 10,259 patients who had undergone routine examination of their nose by nasal endoscopy during a visit at a publicly accessible ear, nose, throat outpatient facility in Vienna were included. Patient details including presenting complaint, nasal symptoms, polyp score, age, gender, treatment, asthma, and allergic status were extracted retrospectively. A detailed questionnaire including history of nasal symptoms, Sino-Nasal Outcome Test-20 German Adapted Version, and visual analog scale was available for 101 patients with nasal polyps. RESULTS: Nasal polyps were detected in 189 of the 10,259 (1.84%) patients. The calculated prevalence of polyps in Austria, adjusted for age and gender, was 1.95%. The average total polyp score (TPS) was 3.4, and 72.5% had a TPS of ≤4, with males and asthmatics having significantly larger polyps. Questionnaire analysis revealed that 67% suffered from a low symptom burden of ≤36. According to current European Position Paper on Rhinosinusitis and Nasal Polyps (EPOS) guidelines, 6% of patients with polyps met and another 8% potentially fulfilled the eligibility criteria for biological therapy. CONCLUSION: Nasal polyp prevalence was calculated to be 1.95% of the Austrian population. Large polyps (TPS >4) were found in 25%, 33% suffered from a high nasal symptom burden, and between 6% and 14% of patients with polyps would be eligible for biological therapy according to EPOS guidelines.

Tracing Human IgE B Cell Antigen Receptor-Bearing Cells With a Monoclonal Anti-Human IgE Antibody That Specifically Recognizes Non-Receptor-Bound IgE.

Up to 30% of the population suffers from immunoglobulin E (IgE)-mediated allergies. Despite current stepwise gating approaches, the unambiguous identification of human IgE-producing cells by flow cytometry and immunohistology remains challenging. This is mainly due to the scarcity of these cells and the fact that IgE is not only expressed in a membrane-bound form on the surface of IgE-producing cells in form of the B cell antigen receptor (BCR), but is more frequently found on various cell types bound to the low and high affinity receptors, CD23 and FcϵRI, respectively. Here we sought to develop a sequential gating strategy for unambiguous detection of cells bearing the IgE BCR on their surface. To that aim we first tested the monoclonal anti-IgE antibody omalizumab for its ability to discriminate between IgE BCR and receptor-bound IgE using cells producing IgE or bearing IgE bound to CD23 as well as basophils exhibiting FcϵRI receptor-bound IgE. Using flow cytometry, we demonstrated that omalizumab recognized IgE producing cells with a high sensitivity of up to 1 IgE+ cell in 1000 human peripheral blood mononuclear cells (PBMCs). These results were confirmed by confocal microscopy both in cell suspensions as well as in nasal polyp tissue sections. Finally, we established a consecutive gating strategy allowing the clear identification of class-switched, allergen-specific IgE+ memory B cells and plasmablasts/plasma cells in human PBMCs. Birch pollen specific IgE+ memory B cells represented on average 0.734% of total CD19+ B cells in allergic patients after allergen exposure. Thus, we developed a new protocol for exclusive staining of non-receptor bound allergen-specific IgE+ B cell subsets in human samples.