Postsurgical outcomes of nonfunctioning pituitary adenomas: a patient-level meta-analysis.

BACKGROUND: Surgical resection is the main treatment for symptomatic nonfunctioning pituitary adenomas (NFPA). We aimed to analyze the impact of surgical approach, completeness of resection, and postoperative radiotherapy on long-term progression-free survival (PFS) of NFPA, using individual patient data (IPD) meta-analysis. METHODS: An electronic literature searched was conducted on PubMed, EMBASE, and Web of Science from database inception to 6 November 2022. Studies describing the natural history of surgically resected NFPA, with provision of Kaplan-Meier curves, were included. These were digitized to obtain IPD, which was pooled in one-stage and two-stage meta-analysis to determine hazard ratios (HRs) and 95%CIs of gross total resection (GTR) versus subtotal resection (STR), and postoperative radiotherapy versus none. An indirect analysis of single-arm data between endoscopic endonasal (EES) and microscopic transsphenoidal (MTS) surgical technique was also performed. RESULTS: Altogether, eleven studies (3941 patients) were retrieved. PFS was significantly lower in STR than GTR (shared-frailty HR 0.32, 95%CI 0.27-0.39, p < 0.001). Postoperative radiotherapy significantly improved PFS compared to no radiotherapy (shared-frailty HR 0.20, 95%CI 0.15-0.26, p < 0.001), including in the subgroup of patients with STR (shared-frailty HR 0.12, 95%CI 0.08-0.18, p < 0.001). Similar PFS was observed between EES and MTS (indirect HR 1.09, 95%CI 0.92-1.30, p = 0.301). CONCLUSIONS: This systematic review and patient-level meta-analysis provides a robust prognostication of surgically treated NFPA. We reinforce current guidelines stating that GTR should be the standard of surgical resection. Postoperative radiotherapy is of considerable benefit, especially for patients with STR. Surgical approach does not significantly affect long-term prognosis. REGISTRATION: PROSPERO CRD42022374034.

Prolonged Primary Rhinovirus Infection of Human Nasal Epithelial Cells Diminishes the Viral Load of Secondary Influenza H3N2 Infection via the Antiviral State Mediated by RIG-I and Interferon-Stimulated Genes.

Our previous study revealed that prolonged human rhinovirus (HRV) infection rapidly induces antiviral interferons (IFNs) and chemokines during the acute stage of infection. It also showed that expression levels of RIG-I and interferon-stimulated genes (ISGs) were sustained in tandem with the persistent expression of HRV RNA and HRV proteins at the late stage of the 14-day infection period. Some studies have explored the protective effects of initial acute HRV infection on secondary influenza A virus (IAV) infection. However, the susceptibility of human nasal epithelial cells (hNECs) to re-infection by the same HRV serotype, and to secondary IAV infection following prolonged primary HRV infection, has not been studied in detail. Therefore, the aim of this study was to investigate the effects and underlying mechanisms of HRV persistence on the susceptibility of hNECs against HRV re-infection and secondary IAV infection. We analyzed the viral replication and innate immune responses of hNECs infected with the same HRV serotype A16 and IAV H3N2 at 14 days after initial HRV-A16 infection. Prolonged primary HRV infection significantly diminished the IAV load of secondary H3N2 infection, but not the HRV load of HRV-A16 re-infection. The reduced IAV load of secondary H3N2 infection may be explained by increased baseline expression levels of RIG-I and ISGs, specifically MX1 and IFITM1, which are induced by prolonged primary HRV infection. As is congruent with this finding, in those cells that received early and multi-dose pre-treatment with Rupintrivir (HRV 3C protease inhibitor) prior to secondary IAV infection, the reduction in IAV load was abolished compared to the group without pre-treatment with Rupintrivir. In conclusion, the antiviral state induced from prolonged primary HRV infection mediated by RIG-I and ISGs (including MX1 and IFITM1) can confer a protective innate immune defense mechanism against secondary influenza infection.

Deconstructive versus reconstructive approach to internal carotid artery blowout in radiated nasopharyngeal carcinoma: A tertiary center experience and systematic review.

The objective of this study was to compare the outcomes of parent artery occlusion (PAO) versus stent-assisted reconstruction in radiated nasopharyngeal carcinoma (NPC) patients with internal carotid artery (ICA) blowouts. A retrospective review from our institution (2011-2021) and systematic review of Pubmed and Embase (1995-2022) was performed. Twenty-eight eligible studies were identified. Eighty-six PAOs and 37 stent-assisted reconstructions were analyzed, including 11 PAOs and 5 stents from our institution. Stents were associated with significantly higher incidence of overall re-bleeding (16.2% [95% CI 7.4-31.9] vs. 4.6% [95% CI 1.3-13.5], p = 0.047), delayed stroke (5.4% [95% CI 1.3-19.4] vs. 0%, p = 0.034) and reduced median survival (7.1 [95% CI 3.8-14.0] months vs. 29.0 [95% CI 9.4-63.4] months, p = 0.017) compared to PAO. There were no significant differences in terms of overall stroke, infection, extruded/migrated foreign body, and peri-procedure death. PAO is preferred over reconstructive treatment in patients with adequate collateral circulation.

Light-sheet laser speckle imaging for cilia motility assessment.

Mucociliary clearance is an important innate defense mechanism predominantly mediated by ciliated cells in the upper respiratory tract. Ciliary motility on the respiratory epithelium surface and mucus pathogen trapping assist in maintaining healthy airways. Optical imaging methods have been used to obtain several indicators for assessing ciliary movement. Light-sheet laser speckle imaging (LSH-LSI) is a label-free and non-invasive optical technique for three-dimensional and quantitative mapping of velocities of microscopic scatterers. Here, we propose to use an inverted LSH-LSI platform to study cilia motility. We have experimentally confirmed that LSH-LSI can reliably measure the ciliary beating frequency and has the potential to provide many additional quantitative indicators for characterizing the ciliary beating pattern without labeling. For example, the asymmetry between the power stroke and the recovery stroke is apparent in the local velocity waveform. PIV (particle imaging velocimetry) analysis of laser speckle data could determine the cilia motion directions in different phases.

The Diagnostic and Prognostic Role of Biomarkers in Chronic Rhinosinusitis.

Chronic rhinosinusitis (CRS) refers to an inflammatory disease of the sinonasal mucosa, with a significant economic burden and impact on quality of life. The diagnosis of CRS is conventionally made on careful history and physical examination, including nasoendoscopic assessment which requires technical expertise. There has been increasing interest in using biomarkers in the non-invasive diagnosis and prognostication of CRS, tailored to the disease inflammatory endotype. Potential biomarkers currently being studied can be isolated from peripheral blood, exhaled nasal gases or nasal secretions, as well as sinonasal tissue. In particular, various biomarkers have revolutionized the way in which CRS is managed, revealing new inflammatory pathways where novel therapeutic drugs are employed to curb the inflammatory process, which may be different from one patient to the next. Biomarkers that have been extensively studied in CRS, such as eosinophil count, IgE, and IL-5, have been associated with a TH2 inflammatory endotype which correlates with an eosinophilic CRSwNP phenotype that predicts a poorer prognosis, tends to recur after conventional surgical treatment, but responds to glucocorticoid treatment. Newer biomarkers that demonstrate potential, such as nasal nitric oxide, can support a diagnosis of CRS with or without nasal polyps, especially when invasive tests such as nasoendoscopy are unavailable. Other biomarkers such as periostin can be used to monitor disease course after treatment of CRS. With a personalized treatment plan, the management of CRS can be individualized, optimizing treatment efficiency and reducing adverse outcomes. As such, this review aims to compile and summarize the existing literature regarding the utility of biomarkers in CRS in terms of diagnosis and prognostication, and also makes recommendations for further studies to fill current knowledge gaps.

Endovascular embolisation of external carotid artery system haemorrhage in radiated nasopharyngeal carcinoma.

PURPOSE: To review the effectiveness and safety of embolisation in managing haemorrhage from the external carotid artery (ECA) system in radiated nasopharyngeal carcinoma (NPC) patients. METHODS: Radiated NPC patients who presented with severe oronasal bleeding and underwent digital subtraction angiography that excluded blowouts from the internal carotid artery from 2011 to 2021 were reviewed. Those who subsequently underwent embolisation of the ECA system were analysed for technical success rate, post-embolisation re-bleeding rate and complications. RESULTS: Seventeen embolisations were performed in fifteen patients during the 10-year period. The technical success rate was 100%, however the early haemostatic rate (no re-bleed within 7 days of embolisation) was 70.6% (12/17) and the overall long-term haemostatic rate was 58.8% (10/17). The re-bleed rates of targeted and empiric embolisations were 33.3% (3/9) and 50.0% (4/8), respectively. The re-bleed rates with liquid agents, coils and particles were 0% (0/7), 33.3% (1/3) and 85.7% (6/7), respectively. Amongst the embolisations utilising liquid agents, 71.4% (5/7) were targeted, distal embolisations. All re-bleeds underwent surgical ligation or repeat embolisation; half of them further experienced recurrent bleeding. There were no significant complications with embolisation. CONCLUSION: Although embolisation of the ECA system in NPC has a high technical success rate and is safe, re-bleeding appears to be common. Targeted, distal embolisation with liquid embolics appear to have good haemostatic effect. Clinicians should be aware that patients may need repeated procedures to secure haemostasis.

Moderate Dose Irradiation Induces DNA Damage and Impairments of Barrier and Host Defense in Nasal Epithelial Cells in vitro.

Purpose: Radiotherapy (RT) is the mainstay treatment for head and neck cancers. However, chronic and recurrent upper respiratory tract infections and inflammation have been commonly reported in patients post-RT. The underlying mechanisms remain poorly understood. Method and Materials: We used a well-established model of human nasal epithelial cells (hNECs) that forms a pseudostratified layer in the air-liquid interface (ALI) and exposed it to single or repeated moderate dose γ-irradiation (1Gy). We assessed the DNA damage and evaluated the biological properties of hNECs at different time points post-RT. Further, we explored the host immunity alterations in irradiated hNECs with polyinosinic-polycytidylic acid sodium salt (poly [I:C]) and lipopolysaccharides (LPS). Results: IR induced DNA double strand breaks (DSBs) and triggered DNA damage response in hNECs. Repeated IR significantly reduced basal cell proliferation with low expression of p63/KRT5 and Ki67, induced cilia loss and inhibited mucus secretion. In addition, IR decreased ZO-1 expression and caused a significant decline in the transepithelial electrical resistance (TEER). Moreover, hyperreactive response against pathogen invasion and disrupted epithelial host defense can be observed in hNECs exposed to repeated IR. Conclusion: Our study suggests that IR induced prolonged structural and functional impairments of hNECs may contribute to patients post-RT with increased risk of developing chronic and recurrent upper respiratory tract infection and inflammation.

Transcriptomics of rhinovirus persistence reveals sustained expression of RIG-I and interferon-stimulated genes in nasal epithelial cells in vitro.

BACKGROUND: Human rhinoviruses (HRVs) are frequently associated with asthma exacerbations, and have been found in the airways of asthmatic patients. While HRV-induced acute infection is well-documented, it is less clear whether the nasal epithelium sustains prolonged HRV infections along with the associated activation of host immune responses. OBJECTIVE: To investigate sustainably regulated host responses of human nasal epithelial cells (hNECs) during HRV persistence. METHODS: Using a time-course study, HRV16 persistence and viral replication dynamics were established using an in vitro infection model of hNECs. RNA sequencing was performed on hNECs in the early and late stages of infection at 3 and 14 days post-infection (dpi), respectively. The functional enrichment of differentially expressed genes (DEGs) was evaluated using gene ontology (GO) and Ingenuity pathway analysis. RESULTS: HRV RNA and protein expression persisted throughout prolonged infections, even after decreased production of infectious virus progeny. GO analysis of unique DEGs indicated altered regulation of pathways related to ciliary function and airway remodeling at 3 dpi and serine-type endopeptidase activity at 14 dpi. The functional enrichment of shared DEGs between the two time-points was related to interferon (IFN) and cytoplasmic pattern recognition receptor (PRR) signaling pathways. Validation of the sustained regulation of candidate genes confirmed the persistent expression of RIG-I and revealed its close co-regulation with interferon-stimulated genes (ISGs) during HRV persistence. CONCLUSIONS: The persistence of HRV RNA does not necessarily indicate an active infection during prolonged infection. The sustained expression of RIG-I and ISGs in response to viral RNA persistence highlights the importance of assessing how immune-activating host factors can change during active HRV infection and the immune regulation that persists thereafter.

Human Nasal Epithelial Cells Sustain Persistent SARS-CoV-2 Infection In Vitro, despite Eliciting a Prolonged Antiviral Response.

The dynamics of SARS-CoV-2 infection in COVID-19 patients are highly variable, with a subset of patients demonstrating prolonged virus shedding, which poses a significant challenge for disease management and transmission control. In this study, the long-term dynamics of SARS-CoV-2 infection were investigated using a human well-differentiated nasal epithelial cell (NEC) model of infection. NECs were observed to release SARS-CoV-2 virus onto the apical surface for up to 28 days postinfection (dpi), further corroborated by viral antigen staining. Single-cell transcriptome sequencing (sc-seq) was utilized to explore the host response from infected NECs after short-term (3-dpi) and long-term (28-dpi) infection. We identified a unique population of cells harboring high viral loads present at both 3 and 28 dpi, characterized by expression of cell stress-related genes DDIT3 and ATF3 and enriched for genes involved in tumor necrosis factor alpha (TNF-α) signaling and apoptosis. Remarkably, this sc-seq analysis revealed an antiviral gene signature within all NEC cell types even at 28 dpi. We demonstrate increased replication of basal cells, absence of widespread cell death within the epithelial monolayer, and the ability of SARS-CoV-2 to replicate despite a continuous interferon response as factors likely contributing to SARS-CoV-2 persistence. This study provides a model system for development of therapeutics aimed at improving viral clearance in immunocompromised patients and implies a crucial role for immune cells in mediating viral clearance from infected epithelia. IMPORTANCE Increasing medical attention has been drawn to the persistence of symptoms (long-COVID syndrome) or live virus shedding from subsets of COVID-19 patients weeks to months after the initial onset of symptoms. In vitro approaches to model viral or symptom persistence are needed to fully dissect the complex and likely varied mechanisms underlying these clinical observations. We show that in vitro differentiated human NECs are persistently infected with SARS-CoV-2 for up to 28 dpi. This viral replication occurred despite the presence of an antiviral gene signature across all NEC cell types even at 28 dpi. This indicates that epithelial cell intrinsic antiviral responses are insufficient for the clearance of SARS-CoV-2, implying an essential role for tissue-resident and infiltrating immune cells for eventual viral clearance from infected airway tissue in COVID-19 patients.

Induction of IL-25 Expression in Human Nasal Polyp Epithelium by Influenza Virus Infection is Abated by Interferon-Alpha Pretreatment.

BACKGROUND: Epithelial cytokines including IL-25, IL-33 and thymic stromal lymphopoietin (TLSP) are recently established as drivers of type 2 chronic inflammatory diseases such as chronic rhinosinusitis with nasal polyps (CRSwNP). Here, we further confirmed the increased expression of IL-25 in CRSwNP and investigated potential contributors of IL-25 in CRSwNP epithelium. METHODS: Sixty CRSwNP, 25 CRSsNP and 15 healthy control tissues were examined for IL-25 expression and for the accompanying type 2 inflammatory cytokines. We then tested different respiratory virus infections on human nasal epithelial cells (hNECs) for their ability to trigger IL-25 expression. In addition, we subjected hNECs generated from CRSwNP tissues to pretreatment with recombinant interferon-alpha (IFN-α) prior to viral infection to evaluate IFN effects on IL-25 induction. RESULTS: We confirmed that significantly enhanced levels of IL-25 were observed in CRSwNP tissues, and that IL-25 expression correlated with type 2 inflammatory cytokine expression. In vitro, we observed significantly elevated IL-25 in hNECs infected with influenza A virus as early as 24 hours post-infection (hpi), regardless of tissue origin, and IL-25 correlated positively with viral load. While other respiratory viruses exhibited increasing trends of IL-25, these were not significant at the time-points tested. IFN-α treatment of CRSwNP epithelium was found to exert bimodal effects, ie IFN-α treatment alone induced moderate IL-25 expression, whereas IFN-α pretreatment of hNECs before influenza infection significantly diminished IL-25 induction by active influenza virus infection. CONCLUSION: We have authenticated the observation of elevated IL-25 in CRSwNP, which is correlated with type 2 inflammatory cytokines. Notably, we identified influenza virus infection as a potential contributor of IL-25 in both control and CRSwNP epithelium during active infection. This IL-25 induction can be abated by IFN-α pretreatment which ameliorated active influenza infection. TRIAL REGISTRATION: Chictr.org.cn ChiCTR-BON-16010179, Registered 18 December 2016, http://www.chictr.org.cn/showproj.aspx?proj=17331. The authors agree on the sharing of deidentified participant data where it pertains to request directly related to the data in this article when contacted (Haiyu Hong; honghy@mail.sysu.edu.cn).

Mucus composition abnormalities in sinonasal mucosa of chronic rhinosinusitis with and without nasal polyps.

Mucus secretion and its composition are vital in the maintenance of airway health, among which hypoxia-inducible factors (HIFs) are thought to be involved in the regulation of mucin synthesis and regulation. Nasal mucus composition difference between healthy individuals and chronic rhinosinusitis (CRS) patients may contribute to the pathology of chronic nasal diseases, but so far, their role has yet to be completely understood. Nasal biopsy specimens were obtained from 24 healthy subjects and 99 patients with CRS without (CRSsNP, n=36) or with (CRSwNP, n=63) nasal polyps. Immunohistochemical (IHC) and immunofluorescent (IF) staining, quantitative real-time PCR, and western blot were performed to compare the nasal mucus composition between the subjects. Areas of the serous gland and mucous gland were both significantly increased in CRSsNP patients. In CRSwNP patients, a decrease in submucosal gland density and a marked increase in goblet cells were observed. The major gel-forming mucins in the sinonasal mucosa of CRSsNP and CRSwNP are MUC5B and MUC5AC respectively. Mucous cells are found in a higher proportion in both CRSsNP and CRSwNP. The proportion of MUC5AC-positive goblet cells was increased in CRSwNP. The mRNA level of HIF-2α was significantly increased in CRS, and both HIF-1α and HIF-2α were expressed in serous cell but not mucous cell. Over secretion and altered composition of mucus are observed in sinonasal mucosa of CRS, which was mainly associated with glandular hyperplasia in CRSsNP and goblet cell hyperplasia in CRSwNP. Mucus abnormality compromised both non-specific and specific antimicrobial capabilities in the sinonasal mucosa. HIF expression may contribute to differences in mucin synthesis and serous gland regulation, which needs further investigation to understand the pathology of CRS.

Contact-Free Co-Culture Model for the Study of Innate Immune Cell Activation During Respiratory Virus Infection.

The early interactions between the nasal epithelial layer and the innate immune cells during viral infections remains an under-explored area. The significance of innate immunity signaling in viral infections has increased substantially as patients with respiratory infections who exhibit high innate T cell activation show a better disease outcome. Hence, dissecting these early innate immune interactions allows the elucidation of the processes that govern them and may facilitate the development of potential therapeutic targets and strategies for dampening or even preventing early progression of viral infections. This protocol details a versatile model that can be used to study early crosstalk, interactions, and activation of innate immune cells from factors secreted by virally infected airway epithelial cells. Using an H3N2 influenza virus (A/Aichi/2/1968) as the representative virus model, innate cell activation of co-cultured peripheral blood mononuclear cells (PBMCs) has been analyzed using flow cytometry to investigate the subsets of cells that are activated by the soluble factors released from the epithelium in response to the viral infection. The results demonstrate the gating strategy for differentiating the subsets of cells and reveal the clear differences between the activated populations of PBMCs and their crosstalk with the control and infected epithelium. The activated subsets can then be further analyzed to determine their functions as well as molecular changes specific to the cells. Findings from such a crosstalk investigation may uncover factors that are important for the activation of vital innate cell populations, which are beneficial in controlling and suppressing the progression of viral infection. Furthermore, these factors can be universally applied to different viral diseases, especially to newly emerging viruses, to dampen the impact of such viruses when they first circulate in naïve human populations.

Using 3D-printed nose models in nasopharyngeal swab training.

BACKGROUND: The COVID-19 pandemic has swept across the globe with massive effects on health care systems as well as global economies. Enhanced testing has been put forward as a means to reduce transmission while awaiting the development of targeted therapy or effective vaccination. However, achieving accurate testing necessitates proper nasopharyngeal swab techniques. METHODS AND RESULTS: We aimed to design and investigate the utility of an anatomically accurate three-dimensional (3D) printed model of the nose in the training for nasopharyngeal swabs. These models were implemented during training sessions for healthcare workers. All participants surveyed felt that the 3D printed models were useful and beneficial in the training of nasopharyngeal swab techniques. CONCLUSIONS: 3D printed nose models are a useful tool in nasopharyngeal swab training. Their usage may help to facilitate the training of potential swabbing manpower in the upscaling of testing capabilities and volumes in this COVID-19 era.

The Emerging Role of Drug-Induced Sleep Endoscopy in the Management of Obstructive Sleep Apnea.

Obstructive sleep apnea is a prevalent sleep disorder characterized by partial or complete obstruction of the upper airway. Continuous positive airway pressure is the first-line therapy for most patients, but adherence is often poor. Alternative treatment options such as mandibular advancement devices, positional therapy, and surgical interventions including upper airway stimulation target different levels and patterns of obstruction with varying degrees of success. Drug-induced sleep endoscopy enables the visualization of upper airway obstruction under conditions mimicking sleep. In the era of precision medicine, this additional information may facilitate better decision-making when prescribing alternative treatment modalities, with the hope of achieving better adherence and/or success rates. This review discusses the current knowledge and evidence on the role of drug-induced sleep endoscopy in the non-positive airway pressure management of obstructive sleep apnea.

Next-Generation Allergic Rhinitis Care in Singapore: 2019 ARIA Care Pathways.

Allergic rhinitis (AR) is prevalent in Singapore, with a significant disease burden. Afflicting up to 13% of the population, AR impairs quality of life, leads to reduced work productivity and is an independent risk factor for asthma. In the last 2 decades, local studies have identified patient and physician behaviours leading to suboptimal control of the disease. Yet, there is an overall lack of attention to address this important health issue. Allergic Rhinitis and its Impact on Asthma (ARIA) is a European organisation aimed at implementing evidence-based management for AR worldwide. Recent focus in Europe has been directed towards empowering patients for self-management, exploring the complementary role of mobile health, and establishing healthcare system-based integrated care pathways. Consolidation of these ongoing efforts has led to the release of the 2019 ARIA care pathways. This review summarises the ARIA update with particular emphasis on the current status of adult AR in Singapore. In addition, we identify unmet needs and future opportunities for research and clinical care of AR in the local context.

Infection of human Nasal Epithelial Cells with SARS-CoV-2 and a 382-nt deletion isolate lacking ORF8 reveals similar viral kinetics and host transcriptional profiles.

The novel coronavirus SARS-CoV-2 is the causative agent of Coronavirus Disease 2019 (COVID-19), a global healthcare and economic catastrophe. Understanding of the host immune response to SARS-CoV-2 is still in its infancy. A 382-nt deletion strain lacking ORF8 (Δ382 herein) was isolated in Singapore in March 2020. Infection with Δ382 was associated with less severe disease in patients, compared to infection with wild-type SARS-CoV-2. Here, we established Nasal Epithelial cells (NECs) differentiated from healthy nasal-tissue derived stem cells as a suitable model for the ex-vivo study of SARS-CoV-2 mediated pathogenesis. Infection of NECs with either SARS-CoV-2 or Δ382 resulted in virus particles released exclusively from the apical side, with similar replication kinetics. Screening of a panel of 49 cytokines for basolateral secretion from infected NECs identified CXCL10 as the only cytokine significantly induced upon infection, at comparable levels in both wild-type and Δ382 infected cells. Transcriptome analysis revealed the temporal up-regulation of distinct gene subsets during infection, with anti-viral signaling pathways only detected at late time-points (72 hours post-infection, hpi). This immune response to SARS-CoV-2 was significantly attenuated when compared to infection with an influenza strain, H3N2, which elicited an inflammatory response within 8 hpi, and a greater magnitude of anti-viral gene up-regulation at late time-points. Remarkably, Δ382 induced a host transcriptional response nearly identical to that of wild-type SARS-CoV-2 at every post-infection time-point examined. In accordance with previous results, Δ382 infected cells showed an absence of transcripts mapping to ORF8, and conserved expression of other SARS-CoV-2 genes. Our findings shed light on the airway epithelial response to SARS-CoV-2 infection, and demonstrate a non-essential role for ORF8 in modulating host gene expression and cytokine production from infected cells.

Host Antiviral Response Suppresses Ciliogenesis and Motile Ciliary Functions in the Nasal Epithelium.

BACKGROUND: Respiratory viral infections are one of the main drivers of development and exacerbation for chronic airway inflammatory diseases. Increased viral susceptibility and impaired mucociliary clearance are often associated with chronic airway inflammatory diseases and served as risk factors of exacerbations. However, the links between viral susceptibility, viral clearance, and impaired mucociliary functions are unclear. Therefore, the objective of this study is to provide the insights into the effects of improper clearance of respiratory viruses from the epithelium following infection, and their resulting persistent activation of antiviral response, on mucociliary functions. METHODS: In order to investigate the effects of persistent antiviral responses triggered by viral components from improper clearance on cilia formation and function, we established an in vitro air-liquid interface (ALI) culture of human nasal epithelial cells (hNECs) and used Poly(I:C) as a surrogate of viral components to simulate their effects toward re-epithelization and mucociliary functions of the nasal epithelium following damages from a viral infection. RESULTS: Through previous and current viral infection expression data, we found that respiratory viral infection of hNECs downregulated motile cilia gene expression. We then further tested the effects of antiviral response activation on the differentiation of hNECs using Poly(I:C) stimulation on differentiating human nasal epithelial stem/progenitor cells (hNESPCs). Using this model, we observed reduced ciliated cell differentiation compared to goblet cells, reduced protein and mRNA in ciliogenesis-associated markers, and increased mis-assembly and mis-localization of ciliary protein DNAH5 following treatment with 25 µg/ml Poly(I:C) in differentiating hNECs. Additionally, the cilia length and ciliary beat frequency (CBF) were also decreased, which suggest impairment of ciliary function as well. CONCLUSION: Our results suggest that the impairments of ciliogenesis and ciliary function in hNECs may be triggered by specific expression of host antiviral response genes during re-epithelization of the nasal epithelium following viral infection. This event may in turn drive the development and exacerbation of chronic airway inflammatory diseases.

An Integrated Analysis of Radial Spoke Head and Outer Dynein Arm Protein Defects and Ciliogenesis Abnormality in Nasal Polyps.

Background: Nasal polyp (NP) is a chronic upper airway inflammatory disease that is frequently triggered by defective host-defense. However, the mechanisms underlying the impaired barrier function such as cilia-mediated mucociliary clearance remain poorly understood. Objective: To assess ciliary ultrastructural and ciliogenesis marker expression and the phenotypes of ciliated cells in NP. Methods: NP biopsy samples were obtained from 97 NP patients and inferior turbinate from 32 healthy controls. Immunofluorescence staining, quantitative polymerase chain reaction, and single-cell cytospin staining were performed. We classified the patterns of radial spoke head protein (RSPH) 1, 4A (RSPH4A), 9 (RSPH9), and dynein axonemal heavy chain 5 (DNAH5) localization. A semi-quantitative scoring system was developed to assess their expression patterns and associations with ciliogenesis markers [centrosomal protein 110 (CP110) and forkhead box j1 (FOXJ1)]. Results: Median scores of RSPH1, RSPH4A, RSPH9, and DNAH5 were significantly higher in NP than in healthy controls, particularly in eosinophilic NPs. Expression pattern scores of RSPH1, RSPH4A, RSPH9, and DNAH5 correlated positively with each other in both groups. In primary-cell specimens, abnormal expression patterns were significantly more common in NP. The total fluorescence intensity of CP110 and FOXJ1 was significantly higher in NPs and correlated positively with expression pattern scores of RSPH1, RSPH4A, RSPH9, and DNAH5. A trend towards lengthened cilia was observed in NP. Conclusion: In the chronic airway inflammatory milieu, the up-regulated ciliogenesis correlates with the abnormal expression of ciliary ultrastructural markers (i.e., DNAH5) in NP (particularly eosinophilic NP).

RNA Sequencing of H3N2 Influenza Virus-Infected Human Nasal Epithelial Cells from Multiple Subjects Reveals Molecular Pathways Associated with Tissue Injury and Complications.

The human nasal epithelium is the primary site of exposure to influenza virus, the initiator of host responses to influenza and the resultant pathologies. Influenza virus may cause serious respiratory infection resulting in major complications, as well as severe impairment of the airways. Here, we elucidated the global transcriptomic changes during H3N2 infection of human nasal epithelial cells from multiple individuals. Using RNA sequencing, we characterized the differentially-expressed genes and pathways associated with changes occurring at the nasal epithelium following infection. We used in vitro differentiated human nasal epithelial cell culture model derived from seven different donors who had no concurrent history of viral infections. Statistical analysis highlighted strong transcriptomic signatures significantly associated with 24 and 48 h after infection, but not at the earlier 8-h time point. In particular, we found that the influenza infection induced in the nasal epithelium early and altered responses in interferon gamma signaling, B-cell signaling, apoptosis, necrosis, smooth muscle proliferation, and metabolic alterations. These molecular events initiated at the infected nasal epithelium may potentially adversely impact the airway, and thus the genes we identified could serve as potential diagnostic biomarkers or therapeutic targets for influenza infection and associated disease management.

Primary Nasopharyngeal Kaposi Sarcoma as Index Diagnosis of AIDS in a Previously Healthy Man.

A 38-year-old, previously healthy man presented with blood-stained saliva and epistaxis. A 3 mm nasopharyngeal lesion was found. A biopsy was performed and microscopic examination revealed a Kaposi sarcoma. The patient was subsequently found to be positive for human immunodeficiency virus (HIV). The diagnosis of Kaposi sarcoma in the presence of HIV infection advanced his disease to Acquired Immunodeficiency Syndrome (AIDS). Primary manifestation of Kaposi sarcoma in the nasopharynx is extremely rare. The histologic differential diagnosis of Kaposi sarcoma in this unusual site, especially without the clinical history of immunosuppression, is broad. Awareness that nasopharynx can be a primary involvement site of Kaposi sarcoma and serves as index diagnosis of AIDS is important given its serious clinical implication.