Anti-inflammatory response to 1,8-Cineol and associated microbial communities in Otitis media patients.

Chronic Otitis Media (COM) is defined as long term inflammation and colonization with pathogenic bacteria due to a defect or retraction of the tympanic membrane. Surgical interventions are often augmented by antibiotic resistance development and therefore, off-label treatment using the natural drug 1,8-Cineol was carried out. All COM patients underwent antibiotic therapy and middle ear surgery and developed antibiotic resistances. Microbiological investigations from the auditory canal and stool samples were performed in correlation with the clinical course. Therapy of COM patients with 1,8-Cineol revealed a clear reduction of inflammatory microbes P. aeruginosa and Proteus mirabilis in ear samples as well as intestinal Prevotella copri, which was associated with an improved clinical outcome in certain individuals. The present off-label study revealed manifold anti-inflammatory effects of the natural monoterpene 1,8-Cineol in Otitis media patients. A better understanding of the underlying mechanisms will improve the current treatment options and possible forms of application of this natural drug.

Immunomodulatory Response of the Middle Ear Epithelial Cells in Otitis Media.

HYPOTHESIS: The middle ear (ME) epithelium transforms because of changed immunomodulation during infection. INTRODUCTION: The epithelial cells of the tympanic cavity represent the first line of defense in the context of otitis media. They can convert from a typical mucosal site into a respiratory epithelium and vice versa. Our goal is to depict the specific immune response of epithelial cells after infection at the molecular level. METHODS: The investigations were carried out on healthy and inflamed ME tissue, removed during surgical interventions in mouse and human models, and in a human in-vitro cell model in human ME epithelial cell line. We determined the epithelial localization of the protein expression of Toll- and NOD-like immune receptors and their associated signaling molecules using immunohistochemistry. In addition, we examined growth behavior and gene expression due to direct stimulation and inhibition. RESULTS: We found clinically and immunobiologically confirmed transformation of the inflamed ME epithelium depending on their origin, as well as differences in the distribution of Toll-like receptors and nucleotide-binding oligomerization domain-like receptors in the epithelial cell lining. Dysregulated gene and protein expression of the inflammatory and apoptotic genes could be modulated by stimulation and inhibition in the epithelial cells. CONCLUSIONS: The local ME mucosal tissue is believed to modulate downstream immune activity after pathogen invasion via intrinsic cellular mechanism. Using translation approaches to target these molecular pathways may offer more reliable clinical resolution of otitis media in the future.

PANEL 3: Otitis media animal models, cell culture, tissue regeneration & pathophysiology.

OBJECTIVE: To review and summarize recently published key articles on the topics of animal models, cell culture studies, tissue biomedical engineering and regeneration, and new models in relation to otitis media (OM). DATA SOURCE: Electronic databases: PubMed, National Library of Medicine, Ovid Medline. REVIEW METHODS: Key topics were assigned to the panel participants for identification and detailed evaluation. The PubMed reviews were focused on the period from June 2019 to June 2023, in any of the objective subject(s) or keywords listed above, noting the relevant references relating to these advances with a global overview and noting areas of recommendation(s). The final manuscript was prepared with input from all panel members. CONCLUSIONS: In conclusion, ex vivo and in vivo OM research models have seen great advancements in the past 4 years. From the usage of novel genetic and molecular tools to the refinement of in vivo inducible and spontaneous mouse models, to the introduction of a wide array of reliable middle ear epithelium (MEE) cell culture systems, the next five years are likely to experience exponential growth in OM pathophysiology discoveries. Moreover, advances in these systems will predictably facilitate rapid means for novel molecular therapeutic studies.

Hearing rehabilitation and microbial shift after middle ear surgery with Vibrant Soundbridge in patients with chronic otitis media.

INTRODUCTION: Patients with otitis media (OM) encounter significant functional hearing impairment with conductive, or a combined hearing loss and long-term sequelae involving impaired speech/language development in children, reduced academic achievement and irreversible disorders of middle and inner ear requiring a long time therapy and/or multiple surgeries. In its persistent chronic form, Otitis media (COM) can often only be treated by undergoing ear surgery for hearing restoration. The persistent inflammatory reaction plays a major role, often caused by multi-resistant pathogens in the ear. Herein, we present outcomes of patients implanted with currently the only FDA approved active Middle Ear Implant Vibrant Soundbridge (VSB), suffering from persistent COM. METHODS: The study enrolled 42 patients, treated by performing middle ear (ME) surgery to different extents and implanted with the VSB to various structures in the ME. Included were 17 children and 25 adults that had recurrent and/or persisting OM and significant hearing loss. Preoperative and postoperative patients' audiometric data were evaluated and the benefit with VSB assessed using the Glasgow Benefit Inventory for adults and pediatric cohorts. The microbial spectrum of pathogens was assessed before and after surgery, exploring the colonization of the otopathogens, as well as the intestinal microbiome from individually burdened patients. RESULTS: The mean functional gain is 29.7 dB HL (range from 10 to 56.2 dB HL) with a significant improvement in speech intelligibility in quiet. Following VSB implantation, no significant differences in coupling were observed at low complication rates. Postoperatively patients showed significantly increased benefit with VSB compared to the untreated situation, including less otorrhea, pain, medical visits, and medication intake, with no recurrent OM and significant bacterial shift in otopathogens. The analysis of the intestinal microbiome displayed a high abundance of bacterial strains that might be linked to chronic and persistent inflammation. CONCLUSIONS: Functional ear surgery including rehabilitation with a VSB in patients suffering from COM present to be safe and effective. The successful acceptance accompanied by the improved audiological performance resulted in significant benefit with VSB, with a shift in the ear pathogens and altered microbiome and thus is a great opportunity to be treated.

Resolution of otitis media in a humanized mouse model.

Otitis media (OM) is one of the largest public health problems of children and has devastating impacts in developing countries. The substantial medical and human costs involved have led to research to understand the disease and improve treatment. Animal models of OM have yielded critical information about the immune, inflammatory and genetic mechanisms of OM. However, it is important to link animal studies to human immune and inflammatory responses. In recent years, "humanized" mice have become a valuable tool to study the human immune system in an animal model. Here we describe the first use of humanized mice to study OM. We demonstrate that humanized mice with a sufficient degree of engraftment recapitulate a normal middle ear (ME) inflammatory response to bacterial infection, including the recruitment of human immune cells, and exhibit normal recovery. Moreover, these animals exhibit regulated expression of human-specific immune and inflammatory genes in the ME. In contrast, mice with insufficient engraftment fail to resolve OM. This model has many potential uses in OM research, including using hematopoietic stem cells from patients with differing degrees of OM susceptibility, to understand the role of human immune responses in proneness to this common childhood disease.

The ECRG4 cleavage product augurin binds the endotoxin receptor and influences the innate immune response during otitis media.

Otitis media (OM), the most common disease of childhood, is typically characterized by bacterial infection of the middle ear (ME). Prominent features of OM include hyperplasia of the ME mucosa, which transforms from a monolayer of simple squamous epithelium with minimal stroma into a full-thickness respiratory epithelium in 2-3 days after infection. Analysis of the murine ME transcriptome during OM showed down-regulation of the tumor suppressor gene Ecrg4 that was temporally related to mucosal hyperplasia and identified stromal cells as the primary ECRG4 source. The reduction in Ecrg4 gene expression coincided with the cleavage of ECRG4 protein to release an extracellular fragment, augurin. The duration of mucosal hyperplasia during OM was greater in Ecrg4 -/- mice, the number of infiltrating macrophages was enhanced, and ME infection cleared more rapidly. ECRG4-null macrophages showed increased bacterial phagocytosis. Co-immunoprecipitation identified an association of augurin with TLR4, CD14 and MD2, the components of the lipopolysaccharide (LPS) receptor. The results suggest that full-length ECRG4 is a sentinel molecule that potentially inhibits growth of the ME stroma. Processing of ECRG4 protein during inflammation, coupled with a decline in Ecrg4 gene expression, also influences the behavior of cells that do not express the gene, limiting the production of growth factors by epithelial and endothelial cells, as well as the activity of macrophages.

Essential Role of the Innate Immune Adaptor RIP2 in the Response to Otitis Media.

Intracellular nucleotide binding and oligomerization domain (NOD) and Toll-like (TLR) receptors have emerged as pivotal sensors of infection. Both Nod1 and Nod2 contain a caspase activation and recruitment domain (CARD) that interacts with the adaptor protein RIP2 (receptor-interaction protein-2). This leads to ubiquitination of RIP2 and in turn to the activation of NFκB and MAPK transcription factors, to command the host defensive response against pathogenic infections. RIP2 is also activated by TLRs 2 and 4, although the mechanism of this activation is less. The role of RIP2 in otitis media (OM) pathogenesis has yet to be examined. Herein, we used in vivo animal models including C57BL/6 wild-type (WT) and RIP2-/- knockout mice inoculated in the middle ear (ME) with non-typeable Haemophilus influenzae (NTHi), a common human OM pathogen, to evaluate the expression of RIP2 and its signaling genes at the cellular level to determine the role of RIP2 in OM pathogenesis and recovery. The Nod1, Nod2, and Ripk2 genes are minimally expressed in the normal ME. However, they are strongly upregulated during acute OM, as are many genes related to RIP2 signaling. However, while signaling genes were expressed by various ME cell types, only mucosal epithelial and stromal cells expressed the NODs, RIP2, and signaling genes required for the activation of the host defensive response. Whereas WT mice clear ME bacteria and recover from OM within 5 days after infection, RIP2-deficient mice show persistent ME bacterial carriage and inflammation to at least 15 days. This includes significantly prolonged mucosal hyperplasia and ME leukocytic infiltration. Recruitment of macrophages is also delayed in comparison to WT mice. Thus, RIP2 is required to elicit a robust innate immune response that promotes bacterial clearance and increases host innate resistance. The results also identify the structural cells of the ME mucosa, as opposed to leukocytes, as the primary sites of NOD/RIP2 activity in the infected ME.

Immunomodulation as a Protective Strategy in Chronic Otitis Media.

Introduction: Major features of the pathogenesis in otitis media, the most common disease in childhood, include hyperplasia of the middle ear mucosa and infiltration by leukocytes, both of which typically resolve upon bacterial clearance via apoptosis. Activation of innate immune receptors during the inflammatory process leads to the activation of intracellular transcription factors (such as NF-κB, AP-1), which regulate both the inflammatory response and tissue growth. We investigated these leading signaling pathways in otitis media using mouse models, human samples, and human middle ear epithelial cell (HMEEC) lines for therapeutic immunomodulation. Methods: A stable otitis media model in wild-type mice and immunodeficient KO-mice, as well as human tissue samples from chronic otitis media, skin from the external auditory canal and middle ear mucosa removed from patients undergoing ear surgery, were studied. Gene and protein expression of innate immune signaling molecules were evaluated using microarray, qPCR and IHC. In situ apoptosis detection determined the apoptotic rate. The influence of bacterial infection on immunomodulating molecules (TNFα, MDP, Tri-DAP, SB203580, Cycloheximide) in HMEEC was evaluated. HMEEC cells were examined after bacterial stimulation/inhibition for gene expression and cellular growth. Results: Persistent mucosal hyperplasia of the middle ear mucosa in chronic otitis media resulted from gene and protein expression of inflammatory and apoptotic genes, including NODs, TNFα, Casp3 and cleaved Casp3. In clinical chronic middle ear samples, these molecules were modulated after a specific stimulation. They also induced a hyposensitive response after bacterial/NOD-/TLR-pathway double stimulation of HMEEC cells in vitro. Hence, they might be suitable targets for immunological therapeutic approaches. Conclusion: Uncontrolled middle ear mucosal hyperplasia is triggered by TLRs/NLRs immunoreceptor activation of downstream inflammatory and apoptotic molecules.

HB-EGF Plays a Pivotal Role in Mucosal Hyperplasia During Otitis Media Induced by a Viral Analog.

Otitis media (OM), the most common childhood illness, can be caused by bacterial and/or viral infection. Hyperplasia of the middle ear (ME) mucosa is an important component of OM that contributes to its deleterious sequelae. Our previous research revealed that ME mucosal hyperplasia in bacterially induced OM was associated with expression of the heparin-binding epidermal growth factor (HB-EGF) gene, and that HB-EGF induced the proliferation of ME mucosal explants in culture. We used single-cell RNA-Seq to identify ME cells that express Hbegf and related genes involved in mediating responses to this factor. To determine the degree to which a viral infection might induce mucosal hyperplasia, and to assess the role of HB-EGF in hyperplasia in vivo, we used, Poly(I:C) to simulate a ME viral infection, Western blotting to confirm ME protein expression, and a specific inhibitor to block the effects of HB-EGF during OM. Genes for HB-EGF and its receptor were expressed in the ME primarily by epithelial, stromal and endothelial cells. Poly(I:C) induced prominent ME mucosal hyperplasia, peaking two days after ME injection. Immunostaining revealed that cleavage of proHB-EGF into its soluble form (sHB-EGF) was strongly induced in response to Poly(I:C). Inhibition of the sHB-EGF receptor dramatically reduced the hyperplastic response of the mucosa. The results demonstrate that a synthetic analog of viral double-stranded RNA interaction can induce OM including a strong proliferative response of the ME mucosa, independent of bacteria. They also indicate that HB-EGF is the dominant growth factor responsible for ME mucosal hyperplasia in vivo.

A transcytotic transport mechanism across the tympanic membrane.

Drug treatments for middle ear diseases are currently delivered systemically, or locally after opening the impermeable tympanic membrane (TM). We previously used bacteriophage display to discover novel peptides that are actively transported across the intact TM, with a variety of transport rates. Peptide structures were analyzed for evidence regarding the mechanism for this unexpected transport, which was then tested by the application of chemical inhibitors. Primary sequences indicated that trans-TM peptides share one of two amino acid motifs. Secondary structures revealed that linear configurations associate with higher transport rates than coiled structures. Tertiary analysis indicated that the shared sequence motifs are prominently displayed at the free ends of rapidly transported peptide phage. The shared motifs were evaluated for similarity to known motifs. The highest probability matches were for protein motifs involved in transmembrane transport and exosomes. Overall, structural findings suggest that the shared motifs represent binding sequences. They also implicate transcytosis, a polarized cell transport mechanism consisting of endocytosis, transcellular transport, and exocytosis. Inhibitor studies indicated that macropinocytosis, retrograde transport through Golgi and exocytosis participate in transport across the TM, consistent with transcytosis. This process can be harnessed to noninvasively deliver therapeutics to the middle ear.

Immunohistochemical and qPCR Detection of SARS-CoV-2 in the Human Middle Ear Versus the Nasal Cavity: Case Series.

Viral infections have already been implicated with otitis media and sudden sensorineural hearing loss. However, the pathophysiology of COVID-19 as it relates to otologic disorders is not well-defined. With the spread of SARS-CoV-2, it is important to evaluate its colonization of middle ear mucosa. Middle ear and nasal tissue samples for quantitative RT-PCR and histologic evaluations were obtained from post-mortem COVID-19 patients and non-diseased control patients. Here we present evidence that SARS-CoV-2 colonizes the middle ear epithelium and co-localizes with the primary viral receptor, angiotensin-converting enzyme 2 (ACE2). Both middle ear and nasal epithelial cells show relatively high expression of ACE2, required for SARS-CoV-2 entry. The epithelial cell adhesion molecule (EpCAM) was use as a biomarker of epithelia. Furthermore, we found that the viral load in the middle ear is lower than that present in the nasal cavity.

[Immunomodulation in Cholesteatoma]. / Immunmodulation im Cholesteatom.

INTRODUCTION: The etiopathogenesis of chronic otitis media epitympanalis/cholesteatoma and its proliferative destructive course with possible complications such as destruction of bony structures with hearing loss, vestibular dysfunction, facial nerve paralysis and intracranial complications are still unexplained. Surgery is still the way to go. New studies are increasingly looking at the innate immune system. METHODS: Our studies were carried out in a mouse model in WT mice and immundeficient KO-mice, as well as in cholesteatoma and healthy ear canal skin and middle ear tissue, which was removed during ear surgery. The expression analyses were carried out at the gene and protein level using TNF as the major target for therapy evaluation. By means of TUNEL staining and immunohistochemistry the level of apoptosis was evaluated. RESULTS: The uncontrolled undirected cholesteatoma growth shows an immunomodulatory profile with up and down-regulation of various gene networks, especially those involved in TNF downstream and upstream signaling pathways. TNF in cholesteatoma is modulated both inflammatorily and apoptotically and therefore is suitable as a possible therapeutic approach in various models. CONCLUSIONS: Cholesteatoma might be immunomodulatory regulated.

A Screen of Autophagy Compounds Implicates the Proteasome in Mammalian Aminoglycoside-Induced Hair Cell Damage.

Introduction: Autophagy is a degradative pathway to safely break down and recycle dysfunctional cellular components. There is prior evidence of autophagy participation during hair cell (HC) damage. Our goal was to screen compounds targeting different aspects of autophagy for their effects on HC loss due to an ototoxic aminoglycoside, gentamicin (GM). Methods: The SELLECKChem autophagy compound library, consisting of 154 compounds with defined autophagy inducing or inhibitory activity, was used for targeted screening in vitro model of ototoxicity. Organ of Corti from postnatal days 3-5 pou4f3/GFP transgenic mice (HCs express green fluorescent protein) were utilized. The organs were micro-dissected, and basal and middle turns divided into micro-explants individually placed into the single wells of a 96-well plate. Samples were treated with 200 µM of GM plus three dosages of tested compound and cultured for 72 h. Negative controls were treated with media only; positive ototoxicity controls were treated with GM only. Results: The majority of the library compounds had no effect on GM-induced HC loss. However, 18 compounds exhibited a significant, protective effect, two compounds were protective at low dosage but showed enhanced GM toxicity at higher doses and one compound was toxic to HCs in the absence of GM. Conclusions: This study evaluated many autophagy compounds that have not been tested previously on HCs. The disparate results obtained underscore the complexity of autophagy events that can influence HC responses to aminoglycosides, but also implicate the proteosome as an important damage mechanism. The screening results can serve as basis for further studies with protective compounds as potential drug targets.

Leukotriene B4 Is a Major Determinant of Leukocyte Recruitment During Otitis Media.

Background: Pathogens of otitis media (OM) induce inflammatory responses in the middle ear (ME), characterized by mucosal hyperplasia, leukocyte infiltration, and inflammatory mediators, including arachidonic acid metabolites. We studied the role of the eicosanoid leukotriene B4 (LTB4) in OM. Methods: Expression of LTB4-related genes was evaluated by gene array and single-cell RNA-Seq in MEs infected with nontypeable Haemophilus influenzae (NTHi). An inhibitor of LTB4 receptor 1 (i.e. U75302) was also used to block LTB4 responses. Results: ME expression of LTB4-related genes was observed by gene arrays and scRNA-Seq. However, not all genes involved in LTB4 generation occurred in any one specific cell type. Moreover, LTB4 receptor inhibition significantly reduced mucosal hyperplasia and virtually eliminated leukocyte infiltration. Conclusions: ME expression of LTB4-related genes suggest a functional role in OM disease. The fact that LTB4-generation is spread across different cell types is consistent with a transcellular pathway of eicosanoid biosynthesis involving cell-to-cell signaling as well as transfer of biosynthetic intermediates between cells. The dramatic reduction in ME leukocyte infiltration caused by U75302 indicates that LTB4 plays a major role in ME inflammatory cell recruitment, acting via the LTB4R1 receptor. Given that there are many other chemotactic factors that occur in the ME during OM, the ability of LTB4 to activate leukocytes and stimulate their extravasation may explain the effects of inhibition. Reduction in mucosal hyperplasia due to U75302 administration may be secondary to the reduction in leukocytes since LTB4R1 is not expressed by mucosal epithelial or stromal cells. The results suggest that LTB4 receptor antagonists could be useful in treating OM.

Macrophage Depletion in CCR2-/- Mice Delays Bacterial Clearance and Enhances Neutrophil Infiltration in an Acute Otitis Media Model.

BACKGROUND: Otitis media (OM) is a common and potentially serious disease of childhood. Although OM is multifactorial on origin, bacterial infection is a unifying component. Many studies have established a critical role for innate immunity in bacterial clearance and OM resolution. A key component of innate immunity is the recruitment of immune and inflammatory cells, including macrophages. METHODS: To explore the role of macrophages in OM, we evaluated the expression of genes related to macrophage function during a complete episode of acute OM in the mouse caused by middle ear (ME) inoculation with Haemophilus influenzae. We also combined CCR2 deficiency with chlodronate liposome toxicity to deplete macrophages during OM. RESULTS: Macrophage genes were robustly regulated during OM. Moreover, macrophage depletion enhanced and prolonged the infiltration of neutrophils into the infected ME and increased the persistence of bacterial infection. CONCLUSIONS: The results illustrate the critical role played by macrophages in OM resolution.

A kinase inhibitor screen identifies signaling pathways regulating mucosal growth during otitis media.

Otitis media, the most common disease of childhood, is characterized by extensive changes in the morphology of the middle ear cavity. This includes hyperplasia of the mucosa that lines the tympanic cavity, from a simple monolayer of squamous epithelium into a greatly thickened, respiratory-type mucosa. The processes that control this response, which is critical to otitis media pathogenesis and recovery, are incompletely understood. Given the central role of protein phosphorylation in most intracellular processes, including cell proliferation and differentiation, we screened a library of kinase inhibitors targeting members of all the major families in the kinome for their ability to influence the growth of middle ear mucosal explants in vitro. Of the 160 inhibitors, 30 were found to inhibit mucosal growth, while two inhibitors enhanced tissue proliferation. The results suggest that the regulation of infection-mediated tissue growth in the ME mucosa involves multiple cellular processes that span the kinome. While some of the pathways and processes identified have been previously implicated in mucosa hyperplasia others are novel. The results were used to generate a global model of growth regulation by kinase pathways. The potential for therapeutic applications of the results are discussed.

Single-Cell Transcriptomes Reveal a Complex Cellular Landscape in the Middle Ear and Differential Capacities for Acute Response to Infection.

Single-cell transcriptomics was used to profile cells of the normal murine middle ear. Clustering analysis of 6770 transcriptomes identified 17 cell clusters corresponding to distinct cell types: five epithelial, three stromal, three lymphocyte, two monocyte, two endothelial, one pericyte and one melanocyte cluster. Within some clusters, cell subtypes were identified. While many corresponded to those cell types known from prior studies, several novel types or subtypes were noted. The results indicate unexpected cellular diversity within the resting middle ear mucosa. The resolution of uncomplicated, acute, otitis media is too rapid for cognate immunity to play a major role. Thus innate immunity is likely responsible for normal recovery from middle ear infection. The need for rapid response to pathogens suggests that innate immune genes may be constitutively expressed by middle ear cells. We therefore assessed expression of innate immune genes across all cell types, to evaluate potential for rapid responses to middle ear infection. Resident monocytes/macrophages expressed the most such genes, including pathogen receptors, cytokines, chemokines and chemokine receptors. Other cell types displayed distinct innate immune gene profiles. Epithelial cells preferentially expressed pathogen receptors, bactericidal peptides and mucins. Stromal and endothelial cells expressed pathogen receptors. Pericytes expressed pro-inflammatory cytokines. Lymphocytes expressed chemokine receptors and antimicrobials. The results suggest that tissue monocytes, including macrophages, are the master regulators of the immediate middle ear response to infection, but that virtually all cell types act in concert to mount a defense against pathogens.

Panel 8: Vaccines and immunology.

OBJECTIVE: To review and highlight significant advances made towards vaccine development and understanding of the immunology of otitis media (OM) since the 19th International Symposium on Recent Advances in Otitis Media (ISOM) in 2015, as well as identify future research directions and knowledge gaps. DATA SOURCES: PubMed database, National Library of Medicine. REVIEW METHODS: Key topics were assigned to each panel member for detailed review. Draft reviews were collated, circulated, and thoroughly discussed when the panel met at the 20th ISOM in June 2019. The final manuscript was prepared with input from all panel members. CONCLUSIONS: Since 2015 there have been a number of studies assessing the impact of licensed pneumococcal vaccines on OM. While these studies have confirmed that these vaccines are effective in preventing carriage and/or disease caused by vaccine serotypes, OM caused by non-vaccine serotype pneumococci and other otopathogens remains a significant health care burden globally. Development of multi-species vaccines is challenging but essential to reducing the global burden of OM. Influenza vaccination has been shown to prevent acute OM, and with novel vaccines against nontypeable Haemophilus influenzae (NTHi), Moraxella catarrhalis and Respiratory Syncytial Virus (RSV) in clinical trials, the potential to significantly prevent OM is within reach. Research into alternative vaccine delivery strategies has demonstrated the power of maternal and mucosal vaccination for OM prevention. Future OM vaccine trials must include molecular diagnostics of middle ear effusion, for detection of viruses and bacteria that are persisting in biofilms and to enable accurate assessment of vaccine impact on OM etiology. Understanding population differences in natural and vaccine-induced immune responses to otopathogens is also important for development of the most effective OM vaccines. Improved understanding of the interaction between otopathogens will also advance development of effective therapies and encourage the assessment of the indirect benefits of vaccination. IMPLICATIONS FOR PRACTICE: While NTHi and M. catarrhalis are the predominant otopathogens, funding opportunities to drive vaccine development for these species are limited due to a focus on prevention of childhood mortality rather than morbidity. Delivery of a comprehensive report on the high financial and social costs of OM, including the potential for OM vaccines to reduce antibiotic use and subsequent development of antimicrobial resistance (AMR), would likely assist in engaging stakeholders to recognize the value of prevention of OM and increase support for efforts on OM vaccine development. Vaccine trials with OM prevention as a clinical end-point are challenging, however a focus on developing assays that measure functional correlates of protection would facilitate OM vaccine development.

Panel 2- recent advance in otitis media bioinformatics.

OBJECTIVES: To update the medical literature on recent large-scale studies employing bioinformatics data analysis tools in otitis media (OM) disease models with a principal focus on developments in the past 5 years. DATA SOURCES: Pubmed indexed peer-reviewed articles. REVIEW METHODS: Comprehensive review of the literature using the following search terms: 'genomics, inflammasome, microRNA, proteomics, transcriptome, bioinformatics' with the term 'otitis media', and 'middle ear'. Included articles published in the English language from January 1, 2015-April 1, 2019. IMPLICATIONS FOR PRACTICE: Large scale bioinformatics tools over the past five years lend credence to the paradigm of innate immune response playing a critical role in host defense against bacteria contributing to Otitis Media (OM) progression from acute to chronic. In total, genomic, miRNAomic, and proteomic analyses all point to the need for a tightly regulated innate immune and inflammatory response in the middle ear. Currently, there is an urgent need for developing novel therapeutic strategies to control immunopathology and tissue damage, improve hearing and enhance host defense for both acute and chronic OM based on full understanding of the basic molecular pathogenesis of OM.

Molecular Screening Strategy to Identify a Non-invasive Delivery Mechanism for the Treatment of Middle Ear Disorders.

Middle ear ailments include a broad range of pathological conditions. Otitis media is the leading middle ear disease of childhood, which incurs significant health care resources in developed countries and, in developing countries, causes significant mortality and morbidity. Recurrent and chronic infections of the middle ear lead to the prolonged presence of inflammatory factors and cellular infiltrates resulting in temporary hearing loss. However, long-term alteration of the middle ear space can pose the risk of permanent damage to the delicate ear structures and cause tissue remodeling. While the etiopathogenesis of middle ear diseases is multifactorial, targeting the biological mechanisms and molecular networks that drive disease development is critical. Yet, a pivotal step in realizing the potential of molecular therapies is the development of methods for local drug delivery, since systemic application risks side effects. Utilizing bacteriophage display in the rat, we discovered rare peptides that are able to transit the intact tympanic membrane from the external canal to the middle ear cavity by an active process. An in vitro assay demonstrated that transport occurs across the tympanic membranes of humans and that the peptides cross the membrane independent of phage. Transport of phage, which is ~900 nm in length, suggests that these peptides could non-invasively deliver drug packages or gene therapy vectors into the middle ear.