Genome-wide association study indicates novel associations of annexin A13 to secretory and GAS2L2 with mucous otitis media.

To evaluate the genetics of chronic nonsuppurative otitis media (OM). We performed a genome-wide association study of 429,599 individuals included in the FinnGen study using three different case definitions: combined chronic nonsuppurative OM (7034 cases) (included serous and mucous chronic OM), mucous chronic OM (5953 cases), and secretory chronic OM (1689 cases). Individuals without otitis media were used as controls (417,745 controls). We used immunohistochemistry (IHC) of the murine middle ear to evaluate the expression of annexin A13. Four loci were significantly associated (p < 1.7 × 10-8) with nonsuppurative OM. Three out of the four association signals included missense variants in genes that may play a role in otitis media pathobiology. According to our subtype-specific analyses, one novel locus, located near ANXA13, was associated with secretory OM. Three loci (near TNFRSF13B, GAS2L2, and TBX1) were associated with mucous OM. Immunohistochemistry of murine middle ear samples revealed annexin A13 expression at the apical pole of the Eustachian tube epithelium as well as variable intensity of the secretory cells of the glandular structure in proximity to the Eustachian tube. We demonstrated that secretory and mucous OM have distinct and shared genetic associations. The association of GAS2L2 with ciliary epithelium function and the pathogenesis of dysfunctional mucosa in mucous OM is suggested. The abundant expression of annexin A13 in the Eustachian tube epithelium, along with its role in apical transport for the binding and transfer of phospholipids, indicates the role of annexin A13 and phospholipids in Eustachian tube dysfunction.

Comparison of Endoplasmic Reticulum Stress Messenger Ribonucleic Acid Expression Between Chronic Otitis Media With and Without Cholesteatoma.

BACKGROUND: We evaluated and compared the role of endoplasmic reticulum stress in chronic otitis media with cholesteatoma and chronic otitis media without cholesteatoma. METHODS: The messenger ribonucleic acid expression of endoplasmic reticulum stress was measured and compared between chronic otitis media with cholesteatoma and chronic otitis media without cholesteatoma according to the presence or absence of bacteria, type of hearing loss, ossicle destruction, and facial canal dehiscence. RESULTS: The expression of immunoglobulin heavy chain-binding protein messenger ribonucleic acid was higher in the chronic otitis media without cholesteatoma group than in the chronic otitis media with cholesteatoma group, and Protein kinase RNA (PKR)-like endoplasmic reticulum kinase and activating transcription factor 6 messenger ribonucleic acid expression were higher in the chronic otitis media with cholesteatoma group than in the chronic otitis media without cholesteatoma group. CONCLUSION: Endoplasmic reticulum stress messenger ribonucleic acids were expressed in both chronic otitis media with cholesteatoma and chronic otitis media without cholesteatoma. The levels of expression of endoplasmic reticulum stress messenger ribonucleic acids differed according to clinical features, suggesting that different endoplasmic reticulum stress pathways are involved in the pathophysiology of different types of chronic otitis media.

Micro RNAs and Circular RNAs in Different Forms of Otitis Media.

The aim of this comprehensive review was to present the current knowledge on the role of microRNAs (miRNAs) in acute, recurrent, and chronic forms of otitis media. Special attention was focused on cholesteatoma of the middle ear. MicroRNAs modulate gene expression, which, in turn, influences the development and likelihood of the recurrence of acute and aggressive chronic middle ear inflammatory processes. Moreover, this study discusses the modulating role of a specific subgroup of noncoding RNA, circular RNA (circRNA). Recognizing the precise potential pathways and the mechanisms of their function may contribute to a better understanding of the molecular bases of middle ear diseases and identifying novel methods for treating this demanding pathology. Articles published between 2009 and 2022 were used in this analysis. In this review, we provide a complete overview of the latest progress in identifying the role and mechanisms of particular miRNAs and circRNAs in acute, recurrent and chronic forms of otitis media.

Investigation of EGF, IL1-α and IL-6 levels and selected hematological parameters (NLR, MPV) in patients with the chronic cholesteatomatous otitis media.

In this study we aimed to investigate epidermal growth factor (EGF), interleukin (IL1)-α and IL-6 levels, and hematological parameters in the serum samples of patients with chronic cholesteatomatous otitis media (CCOM). This prospective included 40 patients who underwent surgery due to CCOM between June 2020 and May 2021. The stage of middle ear cholesteatoma was determined on each chart using the EAONO/JOS system. The control group comprised of 30 adults who were scheduled for septoplasty over the same period in our hospital, had no otological complaints, and had normal otological findings. The demographic, clinical, and laboratory data of the patients were obtained from the electronic medical record system of our hospital. The serum EGF, IL1-α and IL-6 levels, and hematological parameters (neutrophil-lymphocyte ratio (NLR) and mean platelet volume (MPV)) were compared between the CCOM and control groups. Seven patients had Stage 1 and 33 patients had Stage 2 middle ear cholestatoma. There was no statistically significant difference between the CCOM and control groups in terms of age and gender (p=0.092 and p=0.616, respectively). The serum EGF and IL1-α levels of the CCOM group were statistically significantly higher than those of the control group (p=0.047 and p=0.013, respectively). No statistically significant difference was observed in the serum IL-6 levels of the CCOM and control groups (p=0.675). There was also no significant difference between the CCOM and control groups in terms of the mean NLR and MPV values ​​(p=0.887 and p=0.164, respectively). There was no significant difference between the Stage 1 and Stage 2 cholesteatoma subgroups in terms of the mean EGF, IL1-α, IL-6 levels (p=0.204, p=0.557 and p=0.613, respectively), and the mean NLR and MPV values (p=0.487, p=0.439, respectively). Increased serum EGF and IL1-α levels in patients with CCOM suggest that these cytokines may play a role in cholesteatomatous epithelial hyperproliferation.

Effect of Angiogenesis and Lymphangiogenesis in Diesel Exhaust Particles Inhalation in Mouse Model of LPS Induced Acute Otitis Media.

Lymphangiogenesis and angiogenesis might have significant involvement in the pathogenesis of otitis media with effusion. This study investigated the effect of diesel exhaust particles (DEP) on inflammation and lymphangiogenesis in a mouse model of acute otitis media (AOM). BALB/c mice were injected with LPS and exposed to 100 µg/m3 DEP. The mice were divided into four groups: control (no stimulation), AOM, AOM + DEP, and DEP + AOM. The effects of DEP inhalation pre- and post-DEP induction were estimated based on measurements of the auditory brainstem response, mRNA levels of lymphangiogenesis-related genes and cytokines, and histology of the middle ear. Cell viability of human middle ear epithelial cells decreased in a dose-response manner at 24 and 48 hours post-DEP exposure. DEP alone did not induce AOM. AOM-induced mice with pre- or post-DEP exposure showed thickened middle ear mucosa and increased expression of TNF-α and IL1-ß mRNA levels compared to the control group, but increased serum IL-1ß levels were not found in the AOM + Post DEP. The mRNA expression of TLR4, VEGFA, VEGFAC, and VEGFR3 was increased by pre-AOM DEP exposure. The expression of VEFGA protein was stronger in the AOM + Post DEP group than in any other group. The expression of CD31 and CD45 markers in the mouse middle ear tissue was higher in the Pre DEP + AOM group than in the AOM group. This result implies that pre-exposure to DEP more strongly increases inflammation and lymphangiogenesis in a mouse model of acute otitis media.

The Roles of NOD-like Receptors in Innate Immunity in Otitis Media.

Acute otitis media (AOM) can persist or lead to various complications in individuals in which the innate immune system is impaired. In this context, impaired expression of nucleotide-binding oligomerization domain (NOD)-like receptor (NLR), an intracellular pathogen-recognition receptor (PRR), is involved in the etiology of OM in humans and animals, affecting its development, severity, chronicity, recurrence, and associated complications. To assess this relationship, we reviewed literature reports relating NLR expression patterns with the pathophysiology and clinical features of OM in the larger context of impaired innate immunity. We summarized the results of published studies on the expression of NLRs in animals and humans in acute otitis media (AOM), otitis media with effusion (OME), chronic otitis media (COM) with cholesteatoma, and COM without cholesteatoma. NLRs were expressed mainly in association with bacterial infection in AOM, OME, COM with cholesteatoma, and COM without cholesteatoma. In addition, expression of NLRs was affected by the presence or absence of bacteria, fluid characteristics, disease recurrence, tissue type, and repeated surgery. Various factors of the innate immune system are involved in the pathogenesis of OM in the middle ear. NLRs are expressed in AOM, OME, COM with cholesteatoma, and COM without cholesteatoma. Impaired NLR expression induced the development, chronicity and recurrence of OM and exacerbated associated complications, indicating that NLRs have important roles in the pathogenesis of OM.

Altered secretome by diesel exhaust particles and lipopolysaccharide in primary human nasal epithelium.

BACKGROUND: Airway epithelial cells can actively participate in the defense against environmental pathogens to elicit local or systemic inflammation. Diesel exhaust particles (DEP), a main component of urban air pollution with particulate matter, are associated with the occurrence of acute and chronic upper airway inflammatory diseases. OBJECTIVES: We sought to investigate the effect of DEP alone or in combination with lipopolysaccharide on the secretome in the primary human nasal epithelium (PHNE) and to find potential biomarkers to relate DEP exposure to upper airway inflammatory diseases. METHODS: PHNE was cultured at an air-liquid interface to create a differentiated in vivo-like model. Secreted proteins (secretome) on the bottom media of the PHNE were analyzed by mass spectrometry-based label-free quantitative proteomics and ELISA. RESULTS: Considerably more differentially expressed secreted proteins were identified in response to DEP plus lipopolysaccharide than to DEP alone. Some canonical pathways related to inflammation and cancer such as the p53, ß-catenin, and extracellular signal-regulated kinase 1/2 pathways were involved. Among differentially expressed secreted proteins, leukemia inhibitory factor was also detected at a high level in the middle ear effusions of otitis media patients, and the leukemia inhibitory factor level was significantly correlated with daily mean mass concentrations of atmospheric particulate matter averaged over 8 days before sample collection. CONCLUSIONS: Apical stimulation with DEP and lipopolysaccharide can significantly alter the basal secretome in PHNE, and this alteration can be reflected by surrounding inflammation with effusion of fluids in vivo such as middle ear effusions in otitis media patients.

Toll-Like Receptors: Expression and Roles in Otitis Media.

Otitis media is mainly caused by upper respiratory tract infection and eustachian tube dysfunction. If external upper respiratory tract infection is not detected early in the middle ear, or an appropriate immune response does not occur, otitis media can become a chronic state or complications may occur. Therefore, given the important role of Toll-like receptors (TLRs) in the early response to external antigens, we surveyed the role of TLRs in otitis media. To summarize the role of TLR in otitis media, we reviewed articles on the expression of TLRs in acute otitis media (AOM), otitis media with effusion (OME), chronic otitis media (COM) with cholesteatoma, and COM without cholesteatoma. Many studies showed that TLRs 1-10 are expressed in AOM, OME, COM with cholesteatoma, and COM without cholesteatoma. TLR expression in the normal middle ear mucosa is absent or weak, but is increased in inflammatory fluid of AOM, effusion of OME, and granulation tissue and cholesteatoma of COM. In addition, TLRs show increased or decreased expression depending on the presence or absence of bacteria, recurrence of disease, tissue type, and repeated surgery. In conclusion, expression of TLRs is associated with otitis media. Inappropriate TLR expression, or delayed or absent induction, are associated with the occurrence, recurrence, chronicization, and complications of otitis media. Therefore, TLRs are very important in otitis media and closely related to its etiology.

Osteoclastic activity in chronic otitis media with cholesteatoma-related bone destruction.

BACKGROUND: Cholesteatoma-related bone destruction is the cause of many complications due to chronic otitis media. This study aimed to evaluate osteoclastic activity in cholesteatoma-related bone destruction using tartrate-resistant acid phosphatase 5b, an enzyme specific to osteoclastic activity. METHOD: Seventy-two patients diagnosed with chronic otitis media were included in this study and were divided into two groups: with and without bone destruction. The blood serum and tissue tartrate-resistant acid phosphatase 5b levels from both groups were compared. RESULTS: There were no significant differences in the level of serum enzymes between both groups. However, in tissue samples, tartrate-resistant acid phosphatase 5b levels were significantly lower in the bone destruction group than the group without bone destruction. CONCLUSION: This study determined that the level of tartrate-resistant acid phosphatase 5b, a specific enzyme for osteoclastic activity in cholesteatoma-related bone destruction, is locally decreased. This data suggests that osteoclastic activity may decrease in cholesteatoma-related bone destruction. However, further experimental and clinical studies are required to clarify this highly complex mechanism.

Glycolytic Metabolism Is Critical for the Innate Antibacterial Defense in Acute Streptococcus pneumoniae Otitis Media.

Objective: Streptococcus pneumoniae (S.pn) is a common respiratory pathogen and a frequent cause of acute otitis media (AOM) in children. However, little is known about the immunometabolism during AOM. This study was to assess the presence of glucose metabolic reprogramming during AOM and its underlying mechanism affecting inflammatory response and middle ear injury. Methods: The levels of glycolytic metabolism were evaluated by measuring the expression of glycolysis-related genes and the production of metabolites. HE stain, immunofluorescence, immunohistochemistry, enzyme-linked immunosorbent assay (ELISA) and Western blot were performed to measure the effect of glucose metabolic reprogramming on inflammatory response, pneumococcal clearance, hypoxia-inducible factor 1 alpha (HIF-1α) expression and cytokine secretion during AOM, respectively. Results: The analysis of microarray revealed an increase of the expression of glycolysis-related genes during S.pn-induced AOM, which was verified by real-time PCR. Increased glycolysis promoted the production of IL-1ß and TNF-α and facilitated the clearance of S.pn by enhancing phagocytosis and killing capability of neutrophils, but also aggravated the middle ear injury. Furthermore, these pathogenic effects could be reversed after glycolytic inhibitor 2DG treatment. Additionally, HIF-1α was observed to involve in glycolytic metabolism during AOM. Conclusion: S.pn infection induced increased glycolysis conversion during AOM, which promoted inflammatory responses and bacterial clearance, but also aggravated tissue damage.

The transcriptional landscape of the cultured murine middle ear epithelium in vitro.

Otitis media (OM) is the most common paediatric disease and leads to significant morbidity. Although understanding of underlying disease mechanisms is hampered by complex pathophysiology, it is clear that epithelial abnormalities underpin the disease. The mechanisms underpinning epithelial remodelling in OM remain unclear. We recently described a novel in vitro model of mouse middle ear epithelial cells (mMEECs) that undergoes mucociliary differentiation into the varied epithelial cell populations seen in the middle ear cavity. We now describe genome wide gene expression profiles of mMEECs as they undergo differentiation. We compared the gene expression profiles of original (uncultured) middle ear cells, confluent cultures of undifferentiated cells and cells that had been differentiated for 7 days at an air liquid interface (ALI). >5000 genes were differentially expressed among the three groups of cells. Approximately 4000 genes were differentially expressed between the original cells and day 0 of ALI culture. The original cell population was shown to contain a mix of cell types, including contaminating inflammatory cells that were lost on culture. Approximately 500 genes were upregulated during ALI induced differentiation. These included some secretory genes and some enzymes but most were associated with the process of ciliogenesis. The data suggest that the in vitro model of differentiated murine middle ear epithelium exhibits a transcriptional profile consistent with the mucociliary epithelium seen within the middle ear. Knowledge of the transcriptional landscape of this epithelium will provide a basis for understanding the phenotypic changes seen in murine models of OM.

Immunoglobulins and Transcription Factors in Otitis Media.

The causes of otitis media (OM) involve bacterial and viral infection, anatomo-physiological abnormalities of the Eustachian canal and nasopharynx, allergic rhinitis, group childcare centers, second-hand smoking, obesity, immaturity and defects of the immune system, formula feeding, sex, race, and age. OM is accompanied by complex and diverse interactions among bacteria, viruses, inflammatory cells, immune cells, and epithelial cells. The present study summarizes the antibodies that contribute to immune reactions in all types of otitis media, including acute otitis media, otitis media with effusion, and chronic otitis media with or without cholesteatoma, as well as the transcription factors that induce the production of these antibodies. The types and distribution of B cells; the functions of B cells, especially in otorhinolaryngology; antibody formation in patients with otitis media; and antibodies and related transcription factors are described. B cells have important functions in host defenses, including antigen recognition, antigen presentation, antibody production, and immunomodulation. The phenotypes of B cells in the ear, nose, and throat, especially in patients with otitis media, were shown to be CD5low, CD23high, CD43low, B220high, sIgMlow, sIgDhigh, Mac-1low, CD80(B7.1)low, CD86(B7.2)low, and Syndecam-1low. Of the five major classes of immunoglobulins produced by B cells, three (IgG, IgA, and IgM) are mainly involved in otitis media. Serum concentrations of IgG, IgA, and IgM are lower in patients with OM with effusion (OME) than in subjects without otitis media. Moreover, IgG, IgA, and IgM concentrations in the middle ear cavity are increased during immune responses in patients with otitis media. B cell leukemia/lymphoma-6 (Bcl-6) and paired box gene 5 (Pax-5) suppress antibody production, whereas B lymphocyte inducer of maturation program 1 (Blimp-1) and X-box binding protein 1 (XBP-1) promote antibody production during immune responses in patients with otitis media.

Expression of endoplasmic reticulum stress mRNAs in otitis media.

BACKGROUND: Endoplasmic reticulum (ER) stress is a cellular defense mechanism that occurs when ER function is impaired. OBJECTIVE: This study was designed to evaluate the expression of major mRNAs of ER stress in patients with otitis media with effusion (OME), chronic otitis media (COM), and COM with cholesteatoma (CholeOM). MATERIAL AND METHODS: Specimens were collected during surgery from patients with OME, COM, and CholeOM, and the levels of ER stress mRNAs measured by real-time polymerase chain reaction. Levels of ER stress mRNAs were compared in the three groups and correlated with clinical findings and pus culture results. RESULTS: The level of CHOP mRNA was higher, and the levels of sXBP1 and ATF6 mRNAs lower, in the OME than in the other two groups (p < .05 each). Evaluation of bacterial pus culture negative patients showed that the level of ATF6 mRNA was higher in the CholeOM than in the other two groups (p < .05), whereas evaluation of bacterial pus culture positive patients showed that the level of CHOP mRNA was higher in the OME than in the other groups (p < .05). CONCLUSIONS AND SIGNIFICANCE: ER stress may be involved in the pathophysiology of OM and the levels of ER stress mRNAs were expressed differently in each type of otitis media according to bacterial culture test results.

The association between ear involvement and clinical features and prognosis in ANCA-associated vasculitis.

OBJECTIVE: The concept of otitis media with ANCA-associated vasculitis (OMAAV) was recently proposed by the study group of the Japan Otological Society. However, little is known about the effect of ear involvement on the clinical features and prognosis of AAV. We investigate this issue in this study. METHODS: We retrospectively examined 36 patients diagnosed with OMAAV and 44 patients diagnosed with AAV without ear involvement (non-OMAAV) at Ehime University Hospital from 2013 to 2018. We collected serological findings including ANCA type and titer, C-reactive protein (CRP), serum creatinine level, organ involved at initial diagnosis, treatment, remission, disease relapse, and mortality from medical records. We investigated whether clinical features and outcomes differed between the OMAAV and non-OMAAV groups. RESULTS: Age, ANCA titer, and CRP at initial diagnosis were not significantly different between the two groups, and the rate of intravenous cyclophosphamide (IVCY) use also did not differ. The proportions of patients with concurrent eye involvement, facial palsy (FP), and hypertrophic pachymeningitis (HCP) were significantly higher in the OMAAV than in the non-OMAAV group (p = 0.005, 0.005 and 0.049, respectively), while both renal and peripheral nerve involvement were significantly less common in OMAAV patients (p = 0.04). Among the 30 patients with renal involvement, serum creatinine level at diagnosis was significantly lower in the OMAAV group (p = 0.04). The mortality rate was 8.3% in OMAAV and 6.8% in non-OMAAV cases, but this difference was not significant. The rate of relapse was 33.3% in OMAAV and 13.6% in non-OMAAV cases; this difference was significant (p = 0.04). CONCLUSIONS: Serological measurements of disease activity did not differ between the groups. Eye involvement, FP, and HCP, however, were significantly more common in AAV with ear involvement. In addition, renal involvement was less common and renal impairment was milder in AAV with ear involvement. These findings can be considered clinical features. The relapse rate was significantly higher in AAV with ear involvement.

Bitter Taste Receptors and Chronic Otitis Media.

OBJECTIVE: To evaluate the presence of bitter taste receptors (T2Rs) in the middle ear and to examine their relationship with chronic ear infections. STUDY DESIGN: Cross-sectional study. SETTING: Tertiary care hospital. METHODS: This study enrolled 84 patients being evaluated for otologic surgery: 40 for chronic otitis media (COM) and 44 for other surgical procedures (controls). We collected a small piece of mucosa from 14 patients for mRNA analysis and from 23 patients for immunohistochemistry. A total of 55 patients underwent a double-blind taste test to gauge sensitivity to phenylthiocarbamide, denatonium, quinine, sucrose, and sodium chloride; 47 patients gave a salivary sample for single-nucleotide polymorphism analysis of rs1376251 (TAS2R50) and rs1726866 (TAS2R38). RESULTS: Bitter taste receptors were found in all samples, but the repertoire varied among patients. T2R50 was the most consistently identified receptor by mRNA analysis. Its rs1376251 allele was related to susceptibility to COM but not the expression pattern of T2R50. Ratings of bitterness intensity of phenylthiocarbamide, a ligand for T2R38, differed significantly between the COM and control groups. CONCLUSION: T2Rs were found within the middle ear of every patient sampled; the rs1376251 allele of TAS2R50 appears to be related to chronic ear infections. These receptors are an intriguing target for future research and possible drug targeting.

Analysis of Inflammatory Signaling in Human Middle Ear Cell Culture Models of Pediatric Otitis Media.

OBJECTIVES/HYPOTHESIS: Cell culture models are valuable tools for investigation of the molecular pathogenesis of diseases including otitis media (OM). Previous study indicates that age-, sex-, and race-associated differences in molecular signaling may impact disease pathophysiology. Currently, a singular immortalized middle ear epithelial (MEE) cell line exists, HMEEC-1, derived from an adult without known middle ear disease. In this study, HMEEC-1 and primary MEE cultures from pediatric patients with and without OM were stimulated with inflammatory cytokines or OM-pathogenic bacterial lysates to examine differences in the response of molecules associated with OM pathogenesis. STUDY DESIGN: Case-control series. METHODS: MEE cultures were established from patients aged <6 years: two with recurrent OM (ROM), two with OM with effusion (OME), and one patient without OM who was undergoing cochlear implant surgery control undergoing cochlear implantation (Peds CI). Primary MEE cultures and HMEEC-1 cells were stimulated with tumor necrosis factor-α, interleukin (IL)-1ß, or nontypeable Haemophilus influenzae lysate. TNFA, IL1B, IL6, IL8, IL10, and MUC5B were assayed via quantitative polymerase chain reaction. IL-8 was assayed by enzyme-linked immunosorbent assay. RESULTS: Gene/protein target expressions were frequently higher in pediatric OM lines than in HMEEC-1 and Peds CI. HMEEC-1 cells were frequently less responsive to stimuli than all pediatric lines. OME lines were often more responsive than ROM lines. CONCLUSIONS: OM may be associated with specific molecular phenotypes that are retained in primary cell culture. Adult-derived HMEEC-1 cells differ significantly in baseline expression and response of OM-associated molecules relative to pediatric MEE cells. Work is underway to immortalize pediatric OM MEE cultures as improved tools for the OM research community. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:410-416, 2021.

The FUT2 Variant c.461G>A (p.Trp154*) Is Associated With Differentially Expressed Genes and Nasopharyngeal Microbiota Shifts in Patients With Otitis Media.

Otitis media (OM) is a leading cause of childhood hearing loss. Variants in FUT2, which encodes alpha-(1,2)-fucosyltransferase, were identified to increase susceptibility to OM, potentially through shifts in the middle ear (ME) or nasopharyngeal (NP) microbiotas as mediated by transcriptional changes. Greater knowledge of differences in relative abundance of otopathogens in carriers of pathogenic variants can help determine risk for OM in patients. In order to determine the downstream effects of FUT2 variation, we examined gene expression in relation to carriage of a common pathogenic FUT2 c.461G>A (p.Trp154*) variant using RNA-sequence data from saliva samples from 28 patients with OM. Differential gene expression was also examined in bulk mRNA and single-cell RNA-sequence data from wildtype mouse ME mucosa after inoculation with non-typeable Haemophilus influenzae (NTHi). In addition, microbiotas were profiled from ME and NP samples of 65 OM patients using 16S rRNA gene sequencing. In human carriers of the FUT2 variant, FN1, KMT2D, MUC16 and NBPF20 were downregulated while MTAP was upregulated. Post-infectious expression in the mouse ME recapitulated these transcriptional differences, with the exception of Fn1 upregulation after NTHi-inoculation. In the NP, Candidate Division TM7 was associated with wildtype genotype (FDR-adj-p=0.009). Overall, the FUT2 c.461G>A variant was associated with transcriptional changes in processes related to response to infection and with increased load of potential otopathogens in the ME and decreased commensals in the NP. These findings provide increased understanding of how FUT2 variants influence gene transcription and the mucosal microbiota, and thus contribute to the pathology of OM.

TLR4 regulates ROS and autophagy to control neutrophil extracellular traps formation against Streptococcus pneumoniae in acute otitis media.

BACKGROUND: Otitis media (OM), a prevalent pediatric infectious disease, is mainly caused by Streptococcus pneumoniae (S.pn). Neutrophil extracellular traps (NETs), a novel antimicrobial strategy, were reported in 2004. We found that NETs formed in the middle ear with acute otitis media (AOM) induced by S.pn. However, the mechanisms of NETs formation are not entirely clear. METHODS: We stimulated neutrophils isolated from mouse bone marrow with S.pn clinical stain 19F in vitro, and established mouse model of AOM via transbullar injection with S.pn. NETs formation, reactive oxygen species (ROS) production, autophagy activation and bacterial load were analyzed in TLR4-/- and wild-type neutrophils stimulated in vitro with S.pn and in vivo during AOM. RESULTS: We found that autophagy and ROS were required for S.pn-induced NETs formation. Moreover, TLR4 partly mediated NETs formation in response to S.pn in vitro and in vivo during AOM. We also showed that attenuated NETs formation in TLR4-/- neutrophils correlated with an impaired ROS production and autophagy activation in vitro and in vivo. In addition, both the in vivo and in vitro-produced NETs were able to engulf and kill S.pn. CONCLUSIONS: TLR4 regulates ROS and autophagy to control NETs formation against S.pn in the course of AOM. IMPACT: S.pn can induce NETs formation in vitro and in vivo; TLR4 regulates NETs formation by ROS and autophagy; NETs contribute to the clearance of bacteria in acute otitis media. In this study, we firstly found that autophagy and ROS were required for S.pn-induced NETs formation in the model of acute otitis media (AOM). And to some extent, TLR4 mediated NETs formation during AOM. Our research might provide a potential strategy for the treatment of otitis media.

Proteomic Study Identifies Glycolytic and Inflammation Pathways Involved in Recurrent Otitis Media.

Recurrent acute otitis media (RAOM) in children is clinically defined as the occurrence of at least three episodes of acute otitis media over a course of 6 months. A further common pathological condition of interest in the context of pediatric otolaryngology is adenotonsillar hypertrophy (ATH), a common cause of obstructive sleep apnea syndrome. Aimed at unraveling the differential modulation of proteins in the two pathologies and at understanding the possible pathways involved in their onset, we analyzed the proteomic profile of the adenoids from 14 RAOM and ATH patients by using two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS). The 2-DE coupled with MS allowed us to identify 23 spots with significant (p-value < 0.05) changes in protein amount, recognizing proteins involved in neutrophil degranulation and glycolysis pathways.

Periostin in middle ear mucosa according to eosinophilic otitis media severity: Middle ear pathology-based treatment.

OBJECTIVE: Eosinophilic otitis media (EOM) is an intractable disorder associated with bronchial asthma and chronic rhinosinusitis with nasal polyposis. Periostin is an extracellular matrix protein secreted by fibroblasts in response to interleukin (IL)-4 and/or IL-13 and is a known marker for eosinophilic disorders. We assessed serum periostin levels and expression of periostin in the middle ear mucosa according to three grade of EOM severity (grade1 to 3). METHODS: 68 patients of blood and serum samples were corrected by whom diagnose bilateral EOM in Jichi Medical University Saitama Medical Center from January 2015 to June 2017.Immunohistochemical evaluation was performed to 18 EOM middle ears mucosa samples, which cauterized in tree groups and compared to that of chronic otitis media (COM). RESULTS: Serum periostin levels was significantly higher in EOM patients than in COM patients (EOM, 125.0 ± 45.5 ng/mL; COM, 79.4 ± 38.3 ng/mL; P<0.0001). The expression of periostin immunopositivity in the EOM middle ear mucosa was significantly greater in severe cases (grade3 samples) than others (grade1 and grade2 samples) (P <0.001 and P = 0.011, respectively). Periostin was expressed at the lamina propria especially in severe EOM cases and the cases had little response to glucocorticoids treatment. CONCLUSION: This study showed that periostin in the middle ear mucosa was correlated with EOM severity, and EOM with highly expressed periostin had difficulty in glucocorticoids treatment.