Microbiome and mycobiome analyses in continuous positive airway pressure devices.

Objectives: Microorganisms are likely present in continuous positive airway pressure (CPAP) devices used daily. Considering the potential risk of infections among CPAP device users, here we aimed to compare the microbiomes in CPAP devices with those in nasal mucosal samples obtained from corresponding individuals using these devices. Methods: We conducted a prospective cohort study at tertiary medical institutes. Samples were collected from the tubes and filters of CPAP devices and the nasal mucosa of corresponding individuals using these devices. Microbiomes and mycobiomes were analyzed using 16S ribosomal RNA and internal transcribed spacer region sequencing. Results were compared according to the sampling site and usage duration for each patient. Results: Overall, 27 paired human nasal mucosa and CPAP samples were analyzed. Bacteria were present in 7 of 27 tubes (29.6%) and 22 of 27 filters (81.5%). Fungi were present in 2 of the 27 tubes (7.4%) and 16 of the 27 filters (59.3%). Actinobacteria and Firmicutes were the predominant phyla among all samples. Fungi were not detected in any of the nasal mucosal samples. However, Basidiomycota and Ascomycota were predominant in the CPAP filters and tube samples. No significant associations were identified among the results according to sampling site and usage duration. Conclusion: Bacteria or fungi can be detected to some extent in CPAP samples even if the CPAP usage period is short. The association between respiratory infections and these microbiomes or mycobiomes was not investigated. Further research might be required to determine the risk posed by CPAP devices as a microbial contamination source.

Non-invasive strategy: Developing a topical IL-4Rα-specific nanobody for the treatment of allergic airway diseases.

Inhibiting IL-4 and IL-13 are critical cytokines that induce the pathogenic responses of allergic airway diseases. Currently, monoclonal antibodies targeting IL-4Rα are administered subcutaneously to treat eosinophilic rhinosinusitis and allergic asthma. However, these treatments have several drawbacks. To address these issues, we have developed a novel IL-4Rα-targeting nanobody designed for non-invasive delivery to local inflammatory sites in allergic airway diseases. H5, selected via the ribosomal display applied screening from synthetic nanobody library, underwent dimerization and in-silico affinity maturation using AlphaFold2 and GROMACS resulting in a substantial/dramatic enhancement of its binding affinity. H5 effectively controlled inflammatory markers such as MUC5AC, CCL26, and FOXJ1 in human nasal epithelial cells (HNECs) by inhibiting IL-4 and IL-13 signaling. The bivalent form of H5 showed efficacy in easily accessible cells, such as multi-ciliated cells, while the monovalent variant targeted hard-to-reach cells, such as basal cells of HNECs. In summary, we developed a nanobody that could effectively inhibit inflammatory signaling in HNECs via intranasal administration, showing promise as a non-invasive rhinitis treatment.

Outcomes and Predictors of Success Following Multilevel Surgery in Positional and Nonpositional Obstructive Sleep Apnea.

OBJECTIVE: To assess the surgical outcomes and identify predictors of surgical success in patients with positional and non-positional obstructive sleep apnea following multilevel airway surgery. STUDY DESIGN: Retrospective cohort study. SETTING: Singe-tertiary medical center. METHODS: This study included 158 patients with obstructive sleep apnea who underwent multilevel airway surgery. Patients were divided into 2 groups according to position dependency: "positional patients" group (n = 100), and "nonpositional patients" group (n = 58). The characteristics and surgical outcomes of the 2 groups were compared. RESULTS: The nonpositional group included younger and more obese patients in comparison to the positional group. Moreover, the nonpositional group had more severe disease than the positional group. Both groups showed overall improvement after surgery, and the surgical success rate did not differ significantly between the 2 groups (nonpositional, 41.4% vs positional, 48.0%; P = .424). Notably, 69.0% of patients belonging to the non-positional group converted to positional group postoperatively. Logistic regression analysis revealed that larger tonsil size, female sex, and higher mean O2 saturation were associated with higher success rate in the positional group, whereas larger tonsil size was associated with surgical success in the nonpositional group. CONCLUSION: Both nonpositional and positional groups showed improvements following multilevel airway surgery, and surgery induced a transition from nonpositional to positional group. Given that the factors related to surgical success differed between the two groups, surgeons should consider position dependency and these distinct factors during decision-making.

Intermittent hypoxia induces Th17/Treg imbalance in a murine model of obstructive sleep apnea.

Obstructive sleep apnea (OSA) is characterized by cyclic normoxic and hypoxic conditions (intermittent hypoxia, IH) induced by the repeated closure of the upper-airway respiratory tract. As a pathomechanism of OSA, IH results in various comorbidities via chronic inflammation and related pathways. However, the role of other inflammatory cells, such as lymphocytes, has not been well-explored. This study aimed to examine the effects of IH on the distribution and balance of T cell subsets and other related cytokines, and mechanisms in the immune system. We modified OSA mouse model (male C57BL/6N male) using our customized chamber that controls specific sleep and oxygenic cycles. To induce hypoxia, the IH group was repeatedly exposed to 5% O2 and 21% O2 lasting for 120 s each for 7 h daily for 4 weeks. Mice were then subjected to a recovery period of 4 weeks, in which IH stimulation was ceased. T cells and related cytokines were analyzed using flow cytometry and immunohistochemistry. Compared with the control group, the IH group had significantly lower levels of CD4+CD25+Foxp3+ regulatory T cells but higher levels of Th 17, IL-4, HIF-1, and inflammatory cytokines. After the recovery period, these altered changes in the immune cells were recovered, and we found no significant difference in their levels between the control and recovery groups. This study revealed that the Th17/Treg ratio is increased by intermittent hypoxia, and this imbalance can explain immune-related diseases, including recently reported allergies, autoimmune, and even cancer diseases, arising from OSA.

Nicotiana benthamiana-derived dupilumab-scFv reaches deep into the cultured human nasal epithelial cells and inhibits CCL26 expression.

Plants offer a cost-effective and scalable pharmaceutical platform devoid of host-derived contamination risks. However, their medical application is complicated by the potential for acute allergic reactions to external proteins. Developing plant-based protein therapeutics for localized diseases with non-invasive treatment modalities may capitalize on the benefits of plant proteins while avoiding their inherent risks. Dupilumab, which is effective against a variety of allergic and autoimmune diseases but has systemic responses and injection-related side effects, may be more beneficial if delivered locally using a small biological form. In this study, we engineered a single-chain variable fragment (scFv) of dupilumab, termed Dup-scFv produced by Nicotiana benthamiana, and evaluated its tissue permeability and anti-inflammatory efficacy in air-liquid interface cultured human nasal epithelial cells (HNECs). Despite showing 3.67- and 17-fold lower binding affinity for IL-4Ra in surface plasmon resonance assays and cell binding assays, respectively, Dup-scFv retained most of the affinity of dupilumab, which was originally high, with a dissociation constant (KD) of 4.76 pM. In HNECs cultured at the air-liquid interface, Dup-scFv administered on the air side inhibited the inflammatory marker CCL26 in hard-to-reach basal cells more effectively than dupilumab. In addition, Dup-scFv had an overall permeability of 0.8% across cell layers compared to undetectable levels of dupilumab. These findings suggest that plant-produced Dup-scFv can be delivered non-invasively to cultured HNESc to alleviate inflammatory signaling, providing a practical approach to utilize plant-based proteins for topical therapeutic applications.

Airway epithelial CD47 plays a critical role in inducing influenza virus-mediated bacterial super-infection.

Respiratory viral infection increases host susceptibility to secondary bacterial infections, yet the precise dynamics within airway epithelia remain elusive. Here, we elucidate the pivotal role of CD47 in the airway epithelium during bacterial super-infection. We demonstrated that upon influenza virus infection, CD47 expression was upregulated and localized on the apical surface of ciliated cells within primary human nasal or bronchial epithelial cells. This induced CD47 exposure provided attachment sites for Staphylococcus aureus, thereby compromising the epithelial barrier integrity. Through bacterial adhesion assays and in vitro pull-down assays, we identified fibronectin-binding proteins (FnBP) of S. aureus as a key component that binds to CD47. Furthermore, we found that ciliated cell-specific CD47 deficiency or neutralizing antibody-mediated CD47 inactivation enhanced in vivo survival rates. These findings suggest that interfering with the interaction between airway epithelial CD47 and pathogenic bacterial FnBP holds promise for alleviating the adverse effects of super-infection.

Sinonasal seromucinous hamartoma: a single institution case series combined with a narrative review of the literature.

PURPOSE: This study aimed to investigate the clinical and histopathological characteristics of sinonasal seromucinous hamartomas (SHs). METHODS: Eight patients with sinonasal SH and treated at a tertiary hospital between November 2005 and September 2023 were included. Additionally, a systematic review of published articles was conducted, analyzing 48 cases of SH described in the literature. RESULTS: Among the eight patients treated at our institution, tumors originated from the posterior nasal cavity in four patients and middle turbinate and middle meatus were the primary origin in two patients each. Coexistence of inflammatory nasal polyps (NPs) was observed in four cases. Histopathologically, four patients exhibited focal respiratory epithelial adenomatoid hamartoma (REAH) features, and low-grade dysplasia was found in one patient. A combined analysis with previous literature revealed that 46.3% of all cases originated in the anterior nasal cavity. The proportions of cases accompanied by NPs and those with focal REAH features were 20.5% and 39.1%, respectively. Additionally, the frequencies of cases exhibiting dysplastic features (5.4%) and recurrence (2.1%) were low. Remarkably, tumors originating from the anterior region tended to have a higher frequency of dysplasia than those originating from the posterior region, although this difference was not statistically significant (p = 0.0996). CONCLUSION: Patients with sinonasal SH showed favorable treatment outcomes following surgical resection. Focal REAH features and accompanying NPs were frequently observed. A substantial proportion of cases originate in the anterior nasal cavity, and these tumors may exhibit a high tendency for dysplasia.

Diagnosis of Primary Ciliary Dyskinesia via Whole Exome Sequencing and Histologic Findings.

PURPOSE: To assess the diagnostic potential of whole-exome sequencing (WES) and elucidate the clinical and genetic characteristics of primary ciliary dyskinesia (PCD) in the Korean population. MATERIALS AND METHODS: Forty-seven patients clinically suspected of having PCD were enrolled at a tertiary medical center. WES was performed in all patients, and seven patients received biopsy of cilia and transmission electron microscopy (TEM). RESULTS: Overall, PCD was diagnosed in 10 (21.3%) patients: eight by WES (8/47, 17%), four by TEM. Among patients diagnosed as PCD based on TEM results, two patients showed consistent results with WES and TEM of PCD (2/4, 50%). In addition, five patients, who were not included in the final PCD diagnosis group, had variants of unknown significance in PCD-related genes (5/47, 10.6%). The most frequent pathogenic (P)/likely pathogenic (LP) variants were detected in DNAH11 (n=4, 21.1%), DRC1 (n=4, 21.1%), and DNAH5 (n=4, 21.1%). Among the detected 17 P/LP variants in PCD-related genes in this study, 8 (47.1%) were identified as novel variants. Regarding the genotype-phenotype correlation in this study, the authors experienced severe PCD cases caused by the LP/P variants in MCIDAS, DRC1, and CCDC39. CONCLUSION: Through this study, we were able to confirm the value of WES as one of the diagnostic tools for PCD, which increases with TEM, rather than single gene tests. These results will prove useful to hospitals with limited access to PCD diagnostic testing but with relatively efficient in-house or outsourced access to genetic testing at a pre-symptomatic or early disease stage.

Serum high-mobility group box 1 protein level correlates with the lowest SaO2 in patients with sleep apnea: a preliminary study / Nível sérico da proteína de alta mobilidade do grupo Box-1 está associado à SaO2 mais baixa em pacientes com apneia do sono: um estudo preliminar

Abstract Introduction: Serum level of high-mobility group box 1 protein is reportedly correlated with the severity of obstructive sleep apnea. Objective: We tried to evaluate the possibility of using the serum high-mobility group box 1 protein level as a biologic marker in obstructive sleep apnea patients. Methods: We generated a chronic intermittent hypoxia murine model that reflected human obstructive sleep apnea. Obstructive sleep apnea patients who underwent polysomnography were prospectively enrolled. Serum samples were obtained from mice and obstructive sleep apnea patients, and the serum high-mobility group box1 protein level was measured by enzyme-linked immunosorbent assay. Results: Serum high-mobility group box 1 protein level was 56.16 ± 30.33 ng/mL in chronic intermittent hypoxia and 18.63 ± 6.20 ng/mL in control mice (p<0.05). The mean apnea-hypopnea index and respiratory disturbance index values of enrolled obstructive sleep apnea patients were 50.35 ± 27.96 and 51.56 ± 28.53, respectively, and the mean serum high-mobility group box 1 protein level was 30.13 ± 19.97 ng/mL. The apnea-hypopnea index and respiratory disturbance index were not significantly correlated with the serum high-mobility group box 1 protein level (p>0.05). Instead, this protein level was significantly correlated with lowest arterial oxygen concentration (SaO2) (p<0.05). Conclusion: High-mobility group box 1 protein may be involved in the pathogenesis of obstructive sleep apnea, and the possibility of this protein being a useful biologic marker in obstructive sleep apnea should be further evaluated.

Resumo Introdução: O nível sérico da proteína de alta mobilidade do grupo Box-1 está relacionado com a gravidade da apneia obstrutiva do sono. Objetivo: Avaliar o uso do nível sérico da proteína de alta mobilidade do grupo Box-1 como um marcador biológico em pacientes com apneia obstrutiva do sono. Método: Geramos um modelo murino de hipóxia intermitente crônica que imita a apneia obstrutiva do sono em humanos. Pacientes com apneia obstrutiva do sono que fizeram polissonografia foram incluídos prospectivamente. Amostras de soro foram obtidas de camundongos e pacientes com apneia obstrutiva do sono e o nível sérico da proteína de alta mobilidade do grupo Box-1 foi medido por enzyme-linked immunosorbent assay. Resultados: O nível sérico da proteína de alta mobilidade do grupo Box-1 foi 56,16 ± 30,33 ng/mL em hipóxia intermitente crônica e 18,63 ± 6,20 ng/mL em camundongos controle (p < 0,05). Os valores médios do índice de apneia-hipopneia e do índice de distúrbio respiratório nos pacientes com apneia obstrutiva do sono foram 50,35 ± 27,96 e 51,56 ± 28,53, respectivamente, e o nível médio da proteína de alta mobilidade do grupo Box-1 foi 30,13 ± 19,97 ng/mL. O índice de apneia-hipopneia e o índice de distúrbio respiratório não foram significantemente associados com o nível da proteína de alta mobilidade do grupo Box-1 p> 0,05). Em vez disso, esse nível de proteína foi significantemente associado com o valor mais baixo da concentração arterial de oxigênio (SaO2) (p <0,05). Conclusão: A proteína de alta mobilidade do grupo Box-1 pode estar envolvida na patogênese da apneia obstrutiva do sono e a possibilidade de que essa proteína possa ser um marcador biológico útil na apneia obstrutiva do sono deve ser avaliada mais detalhadamente.