Organoids-the key to novel therapies for the inner ear?

The sensitivity and the complexity of the human inner ear in conjunction with the lack of regenerative capacity are the main reasons for hearing loss and tinnitus. Progress in the development of protective and regenerative therapies for the inner ear often failed in the past not least due to the fact that no suitable model systems for cell biological and pharmacological in vitro studies were available. A novel technology for creating "mini-organs", so-called organoids, could solve this problem and has now also reached inner ear research. It makes it possible to produce inner ear organoids from cochlear stem/progenitor cells, embryonic and induced pluripotent stem cells that mimic the structural characteristics and functional properties of the natural inner ear. This review focuses on the biological basis of these inner ear organoids, the current state of research and the promising prospects that are now opening up for basic and translational inner ear research.

High-Dose Glucocorticoids for the Treatment of Sudden Hearing Loss.

BACKGROUND: Systemic glucocorticoids are commonly used for primary therapy of idiopathic sudden sensorineural hearing loss (ISSNHL). However, the comparative effectiveness and risk profiles of high-dose over lower-dose regimens remain unknown. METHODS: We randomly assigned patients with sudden hearing loss of greater than or equal to 50 dB within 7 days from onset to receive either 5 days of high-dose intravenous prednisolone at 250 mg/d (HD-Pred), 5 days of high-dose oral dexamethasone at 40 mg/d (HD-Dex), or, as a control, 5 days of oral prednisolone (Pred-Control) at 60 mg/d followed by 5 days of tapering doses. The primary outcome was the change in hearing threshold (pure tone average) in the three most affected contiguous frequencies from baseline to day 30. Secondary outcomes included speech understanding, tinnitus, communication competence, quality of life, hypertension, and insulin resistance. RESULTS: A total of 325 patients were randomly assigned. Mean change in 3PTAmost affected hearing threshold from baseline to 30 days was 34.2 dB (95% CI, 28.4 to 40.0) in the HD-Pred group, 41.4 dB (95% CI, 35.6 to 47.2) in the HD-Dex group, and 41.0 dB (95% CI, 35.2 to 46.8) in the Pred-Control group (P=0.09 for analysis of variance). There were more adverse events related to trial medication in the HD-Pred (n=73) and HD-Dex (n=76) groups than in the Pred-Control group (n=46). CONCLUSIONS: Systemic high-dose glucocorticoid therapy was not superior to a lower-dose regimen in patients with ISSNHL, and it was associated with a higher risk of side effects. (Funded by the Federal Ministry of Education and Research [BMBF]; EudraCT number, 2015­002602­36.)

[Organoids-the key to novel therapies for the inner ear? German version]. / Organoide ­ der Schlüssel zu neuen Therapien für das Innenohr?

The sensitivity and the complexity of the human inner ear in conjunction with the lack of regenerative capacity are the main reasons for hearing loss and tinnitus. Progress in the development of protective and regenerative therapies for the inner ear often failed in the past not least due to the fact that no suitable model systems for cell biological and pharmacological in vitro studies were available. A novel technology for creating "mini-organs", so-called organoids, could solve this problem and has now also reached inner ear research. It makes it possible to produce inner ear organoids from cochlear stem/progenitor cells, embryonic and induced pluripotent stem cells that mimic the structural characteristics and functional properties of the natural inner ear. This review focuses on the biological basis of these inner ear organoids, the current state of research and the promising prospects that are now opening up for basic and translational inner ear research.

[Has the SARS-CoV-2 pandemic improved teaching?-Virtual education in otorhinolaryngology from the students' perspective]. / Hat die SARS-CoV-2-Pandemie die Lehre verbessert? ­ Virtueller Unterricht im Fach HNO-Heilkunde aus Sicht der Studierenden.

BACKGROUND: The Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV­2) pandemic has significantly changed the education of medical students. Due to the contact restrictions and the associated requirement for distance learning, digital teaching formats had to be implemented within a short period of time. The aim of our work was to analyze student evaluation data for virtual teaching in otorhinolaryngology (ORL) during the SARS-CoV­2 pandemic and to compare the data with previously obtained evaluation data under face-to-face conditions. MATERIALS AND METHODS: Evaluation data for the block practical courses in winter semester 2020/21 and summer semester 2021, which were carried out in a virtual format with a short face-to-face phase as well as those for the block practical courses from summer semester 2018 to winter semester 2019/20, which had been performed completely in a conventional face-to-face format, were analyzed. The anonymous survey of the students focused on various aspects of the courses such as organization, didactics and learning atmosphere. RESULTS: Of 16 surveyed categories, 14 (87.5%) showed significantly better evaluation results for the virtual courses compared to the courses carried out previously under face-to-face conditions. This very positive assessment of the digital teaching offer showed no significant change during the course of the pandemic over the period of two semesters. CONCLUSIONS: Our data show a high acceptance of digital teaching in ORL for students. Even though essential components of the medical education such as teaching on the patient and clinical-practical skills can still only be realized in a face-to-face format, our data suggest that digital elements could also play a role in medical education after the SARS-CoV­2 pandemic.

Powered air-purifying respirators used during the SARS-CoV-2 pandemic significantly reduce speech perception.

BACKGROUND: Due to the coronavirus disease 2019 (COVID-19) pandemic, interventions in the upper airways are considered high-risk procedures for otolaryngologists and their colleagues. The purpose of this study was to evaluate limitations in hearing and communication when using a powered air-purifying respirator (PAPR) system to protect against severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) transmission and to assess the benefit of a headset. METHODS: Acoustic properties of the PAPR system were measured using a head and torso simulator. Audiological tests (tone audiometry, Freiburg speech test, Oldenburg sentence test (OLSA)) were performed in normal-hearing subjects (n = 10) to assess hearing with PAPR. The audiological test setup also included simulation of conditions in which the target speaker used either a PAPR, a filtering face piece (FFP) 3 respirator, or a surgical face mask. RESULTS: Audiological measurements revealed that sound insulation by the PAPR headtop and noise, generated by the blower-assisted respiratory protection system, resulted in significantly deteriorated hearing thresholds (4.0 ± 7.2 dB hearing level (HL) vs. 49.2 ± 11.0 dB HL, p < 0.001) and speech recognition scores in quiet (100.0 ± 0.0% vs. 2.5 ± 4.2%, p < 0.001; OLSA: 20.8 ± 1.8 dB vs. 61.0 ± 3.3 dB SPL, p < 0.001) when compared to hearing without PAPR. Hearing with PAPR was significantly improved when the subjects were equipped with an in-ear headset (p < 0.001). Sound attenuation by FFP3 respirators and surgical face masks had no clinically relevant impact on speech perception. CONCLUSIONS: The PAPR system evaluated here can be considered for high-risk procedures in SARS-CoV-2-positive patients, provided that hearing and communication of the surgical team are optimized by the additional use of a headset.

Simultaneous treatment with pentoxifylline does not adversely affect the neurotrophic effects of brain-derived neurotrophic factor on spiral ganglion neurons.

The hemorheologic drug pentoxifylline is applied for the treatment of sudden sensorineural hearing loss and tinnitus to improve cochlear microcirculation. Recent studies also suggest protective and trophic effects on neuronal cells. Because the preservation of sensorineural structures of the inner ear is fundamental for normal hearing and hearing restoration with auditory prostheses, pentoxifylline and neurotrophic factors such as brain-derived neurotrophic factor (BDNF) are promising candidates to treat degenerative disorders of the inner ear. We used an in-vitro model to determine the neurotrophic effects of these factors on spiral ganglion cells from postnatal rats. Pentoxifylline, alone and in combination with BDNF, was added at various concentrations to the cultured cells. Cells were immunolabeled and analyzed to determine neuronal survival, neurite length, neuronal branching and morphology. Pentoxifylline did not significantly increase or decrease neuronal survival, neurite length and neuronal branching compared to control cultures. Analysis of cellular morphology showed that diverse neuronal subtypes developed in the presence of pentoxifylline. Our data revealed that pentoxifylline did not interfere with the robust neurotrophic effects of BDNF on spiral ganglion neurons when cultured cells were treated with pentoxifylline and BDNF simultaneously. The results of our study do not suggest major neurotrophic effects of pentoxifylline on cultured spiral ganglion neurons. Because pentoxifylline has no detrimental effects on spiral ganglion neurons and does not reduce the effects of BDNF, both agents could be combined to treat diseases of the inner ear provided that future in vivo experiments and clinical studies support these findings.

Typical symptoms of common otorhinolaryngological diseases may mask a SARS-CoV-2 infection.

PURPOSE: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) replicates predominantly in the upper respiratory tract and is primarily transmitted by droplets and aerosols. Taking the medical history for typical COVID-19 symptoms and PCR-based SARS-CoV-2 testing have become established as screening procedures. The aim of this work was to describe the clinical appearance of SARS-CoV-2-PCR positive patients and to determine the SARS-CoV-2 contact risk for health care workers (HCW). METHODS: The retrospective study included n = 2283 SARS-CoV-2 PCR tests from n = 1725 patients with otorhinolaryngological (ORL) diseases performed from March to November 2020 prior to inpatient treatment. In addition, demographic data and medical history were assessed. RESULTS: n = 13 PCR tests (0.6%) were positive for SARS-CoV-2 RNA. The positive rate showed a significant increase during the observation period (p < 0.01). None of the patients had clinical symptoms that led to a suspected diagnosis of COVID-19 before PCR testing. The patients were either asymptomatic (n = 4) or had symptoms that were interpreted as symptoms typical of the ORL disease or secondary diagnoses (n = 9). CONCLUSION: The identification of SARS-CoV-2-positive patients is a considerable challenge in clinical practice. Our findings illustrate that taking a medical history alone is of limited value and cannot replace molecular SARS-CoV-2 testing, especially for patients with ORL diseases. Our data also demonstrate that there is a high probability of contact with SARS-CoV-2-positive patients in everyday clinical practice, so that the use of personal protective equipment, even in apparently "routine cases", is highly recommended.

In vitro comparison of surgical techniques in times of the SARS-CoV-2 pandemic: electrocautery generates more droplets and aerosol than laser surgery or drilling.

INTRODUCTION: Based on current knowledge, the SARS-CoV-2 is transmitted via droplet, aerosols and smear infection. Due to a confirmed high virus load in the upper respiratory tract of COVID-19 patients, there is a potential risk of infection for health care professionals when performing surgical procedures in this area. The aim of this study was the semi-quantitative comparison of ENT-typical interventions in the head and neck area with regard to particle and aerosol generation. These data can potentially contribute to a better risk assessment of aerogenic SARS-CoV-2-transmission caused by medical procedures. MATERIALS AND METHODS: As a model, a test chamber was created to examine various typical surgical interventions on porcine soft and hard tissues. Simultaneously, particle and aerosol release were recorded and semi-quantitatively evaluated time-dependently. Five typical surgical intervention techniques (mechanical stress with a passive instrument with and without suction, CO2 laser treatment, drilling and bipolar electrocoagulation) were examined and compared regarding resulting particle release. RESULTS: Neither aerosols nor particles could be detected during mechanical manipulation with and without suction. The use of laser technique showed considerable formation of aerosol. During drilling, mainly solid tissue particles were scattered into the environment (18.2 ± 15.7 particles/cm2/min). The strongest particle release was determined during electrocoagulation (77.2 ± 30.4 particles/cm2/min). The difference in particle release between electrocoagulation and drilling was significant (p < 0.05), while particle diameter was comparable. In addition, relevant amounts of aerosol were released during electrocoagulation (79.6% of the maximum flue gas emission during laser treatment). DISCUSSION: Our results demonstrated clear differences comparing surgical model interventions. In contrast to sole mechanical stress with passive instruments, all active instruments (laser, drilling and electrocoagulation) released particles and aerosols. Assuming that particle and aerosol exposure is clinically correlated to the risk of SARS-CoV-2-transmission from the patient to the physician, a potential risk for health care professionals for infection cannot be excluded. Especially electrocautery is frequently used for emergency treatment, e.g., nose bleeding. The use of this technique may, therefore, be considered particularly critical in potentially infectious patients. Alternative methods may be given preference and personal protective equipment should be used consequently.

[Implementation of an operational concept in an ENT clinic in the context of the SARS-CoV-2 pandemic]. / Implementierung eines Betriebskonzeptes in einer HNO-Klinik im Rahmen der SARS-CoV-2-Pandemie.

The SARS-CoV-2 pandemic poses major challenges for the entire medical care system. Especially in university institutions as maximum care providers, a higher exposure to potentially infectious patients or actual COVID-19 patients is to be expected. In a short period of time, an operational concept had to be developed regarding the current hygiene recommendations of the Robert Koch Institute (RKI), the leading medical societies and the internal hospital hygiene plan. Here, patient safety and employee protection are equally important.In cooperation with the Institute for Medical Microbiology and Hospital Hygiene and the occupational medical service, interventions were defined to develop solutions to minimize the COVID-19 transmission risk for examiners and patients despite limited diagnostic and equipment resources. For this purpose, an operational concept was developed, consisting of various individual actions, e. g. the reduction of outpatient treatment to emergencies, life-threatening diseases and urgent aftercare, a double triage of patients and the introduction of treatment teams.The newly developed operational concept was successfully implemented within a few days. After the initial rollout and several "hygiene inspections" only minor improvements to the concept were necessary. All measures were documented in the internal quality handbook and are accessible to all employees. Since the SARS-CoV-2 pandemic is a dynamic process with regular changes in the development and information status, the operational concept is regularly reviewed for validity and adjusted as necessary.

Relevance of Neurotrophin Receptors CD271 and TrkC for Prognosis, Migration, and Proliferation in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide and often has a poor prognosis. The present study investigated the role of the low affinity nerve growth factor receptor CD271 as a putative therapy target in HNSCC. Neurotrophins that bind to CD271 also have a high affinity for the tropomyosin receptor kinase family (Trk), consisting of TrkA, TrkB, and TrkC, which must also be considered in addition to CD271. A retrospective study and functional in vitro cell line tests (migration assay and cell sorting) were conducted in order to evaluate the relevance of CD271 expression alone and with regard to Trk expression. CD271 and Trks were heterogeneously expressed in human HNSCC. The vast majority of tumors exhibited CD271 and TrkA, whereas only half of the tumors expressed TrkB and TrkC. High expression of CD271-positive cells predicted a bad clinical outcome of patients with HNSCC and was associated with distant metastases. However, the human carcinomas that also expressed TrkC had a reduced correlation with distant metastases and better survival rates. In vitro, CD271 expression marked a subpopulation with higher proliferation rates, but proliferation was lower in tumor cells that co-expressed CD271 and TrkC. The CD271 inhibitor LM11A 31 suppressed cell motility in vitro. However, neither TrkA nor TrkB expression were linked to prognosis or cell proliferation. We conclude that CD271 is a promising candidate that provides prognostic information for HNSCC and could be a putative target for HNSCC treatment.

Growth hormone promotes neurite growth of spiral ganglion neurons.

Intact spiral ganglion neurons are a specific requirement for hearing rehabilitation in deaf patients by cochlear implantation. Neurotrophic growth factors have been proposed as effective tools to protect and regenerate spiral ganglion neurons that are degenerated in the majority of patients suffering from hearing loss. Here, we show that growth hormone (GH), a pleiotropic growth factor whose neurotrophic role in the inner ear is still unclear, significantly increases neurite extension, as well as neuronal branching, in spiral ganglion cell cultures derived from early postnatal rats. Our data suggest that GH can act as a potent neurotrophic factor for inner ear neurons, which specifically promotes neurite growth. These effects might be elicited in a direct way or, alternatively, by induction of other growth factors that account for the observed neurotrophic effects. Thus, we conlude that GH might represent a novel candidate for the treatment of neurodegeneration in the hearing-impaired inner ear that has the potential to ultimately improve the performance and outcome of modern auditory implants.

Expression of ALDH1A1 and CD44 in primary head and neck squamous cell carcinoma and their value for carcinogenesis, tumor progression and cancer stem cell identification.

In head and neck squamous cell carcinoma (HNSCC), aldehyde dehydrogenase 1 family, member A1 (ALDH1A1) and hyaluronan receptor cluster of differentiation 44 (CD44) are often used as cancer stem cell (CSC) markers. The aim of the present study was to examine the relevance of these proteins for HNSCC in general and for the identification of CSCs. Tumors from 48 patients with primary HNSCC were analyzed for the expression of ALDH1A1 and CD44. Additionally, the association of the proteins with the proliferation rate and epidermal growth factor receptor (EGFR) expression was analyzed. ALDH1A1 was expressed in 54.2% of the carcinoma samples while CD44 was expressed in 89.6% of the carcinoma samples. Most notably, these proteins were often not expressed exclusively in a subpopulation, but also in the majority of tumor cells (ALDH1A1: 30.8% of ALDH1A1+ tumors; CD44: 65.1% of CD44+ tumors). Furthermore, patients with ALDH1A1+ tumors exhibited worse survival rates. CD44 and EGFR expression patterns were overlapping within the tumors and the expression rates were significantly connected. Ki-67+ tumor cells often expressed CD44. ALDH1A1 and CD44 expression patterns only partly overlapped. Consequently, ALDH1A1 and CD44 play significant roles in carcinogenesis and tumor progression. Within the present study, CD44 appeared to interact with EGFR and was more often expressed in primary HNSCC than the marker ALDH1A1. However, ALDH1A1 was a better marker to define a subpopulation of tumor cells. Finally, neither ALDH1A1 nor CD44, alone or combined, were sufficient to determine the CSC population in HNSCC.

Clonal colony formation from spiral ganglion stem cells.

Neural stem cells from the central nervous system have the distinct capacity to give rise to clonal neurospheres. These clonal spheres are derived from a single clone-forming cell and represent homogenous, pure cell colonies. Recently, stem/progenitor cells have been isolated from the spiral ganglion of the inner ear using sphere-forming assays. However, the clonality of spiral ganglion-derived spheres has not yet been addressed in detail. Here, we report the isolation of clonal colonies from the spiral ganglion of early postnatal mice. We analyze sphere clonality using coculture experiments with transgenic cells, a semisolid assay, and culture of single cells in isolation. Our data show that sphere clonality differs in primary and secondary cultures and indicate that clonal sphere formation is dependent on specific culture parameters. We also show that the initiation of clonal colony formation does not require cell-to-cell interactions or paracrine signaling from surrounding cells. Generation of clonal colonies from spiral ganglion stem/progenitor cells might be crucial for future clinical applications because pure cell populations are considered to be more efficient and safe for therapeutic use than chimeric, heterogeneous spheres.

Spiral ganglion stem cells can be propagated and differentiated into neurons and glia.

The spiral ganglion is an essential functional component of the peripheral auditory system. Most types of hearing loss are associated with spiral ganglion cell degeneration which is irreversible due to the inner ear's lack of regenerative capacity. Recent studies revealed the existence of stem cells in the postnatal spiral ganglion, which gives rise to the hope that these cells might be useful for regenerative inner ear therapies. Here, we provide an in-depth analysis of sphere-forming stem cells isolated from the spiral ganglion of postnatal mice. We show that spiral ganglion spheres have characteristics similar to neurospheres isolated from the brain. Importantly, spiral ganglion sphere cells maintain their major stem cell characteristics after repeated propagation, which enables the culture of spheres for an extended period of time. In this work, we also demonstrate that differentiated sphere-derived cell populations not only adopt the immunophenotype of mature spiral ganglion cells but also develop distinct ultrastructural features of neurons and glial cells. Thus, our work provides further evidence that self-renewing spiral ganglion stem cells might serve as a promising source for the regeneration of lost auditory neurons.

HPV status and overall survival of patients with oropharyngeal squamous cell carcinoma--a retrospective study of a German head and neck cancer center.

BACKGROUND/AIM: A change in epidemiology of head and neck squamous cell carcinoma (HNSCC) has been noticed: while overall incidence has decreased, the incidence of oropharyngeal SCC (OSCC) has been increasing over the past decades. A growing body of evidence suggests a causative role of the human papillomavirus (HPV) as an independent risk factor in development of OSCC. The aim of this study was to determine the HPV status in all OSCC specimens collected in our biological database since 1988, correlating the results with overall survival, and to compare them with the current literature data. PATIENTS AND METHODS: A total of 104 tumor samples were obtained and included in this study. Patient records were reviewed. HPV status was determined by a two-step polymerase chain reaction (PCR) combined with p16 immunohistochemistry. Statistical analysis was performed with BiAS™. RESULTS: Overall 12 (12%) of the 104 tumor samples were HPV-positive. Most of the patients had advanced disease [(UICC) stage III or IV)]: 91.7 % in the HPV-positive group versus 78.2% in the HPV-negative group. Multivariate analysis showed that HPV status (p=0.04), UICC stage (p=0.01) and age at initial diagnosis (p=0.0006) were all independent determinants of overall survival. A positive HPV status (hazard ratio=0.52; 95%) was associated with a 48% increase of overall survival compared to patients with HPV-negative tumors. CONCLUSION: Our findings confirm a prevalence of HPV-positive tumors within OSCC. Due to its epidemiologic and prognostic relevance, HPV status should be considered an important part of tumor staging. For this purpose, HPV detection via two-step PCR combined with p16 immunohistochemistry seems reliable.

The role of p16 expression as a predictive marker in HPV-positive oral SCCHN--a retrospective single-center study.

UNLABELLED: Squamous cell carcinoma of the head and neck (SCCHN) is the sixth most common type of cancer worldwide; 600,000 new cases are diagnosed every year. Infected with high-risk human papilloma virus (HPV) types are particularly linked to oropharyngeal cancer. Among over 100 different HPV types, HPV-16 and HPV-18 are detected in the majority of HPV-positive SCCHNs. The p16 gene is often mutated in SCCHN, its overexpression is caused by the viral E7 protein. Consequently, p16 is assumed to be an indirect marker of HPV-induced SCCHN. The aim of the present study was to determine the role of p16 expression as a predictive marker of HPV infection in SCCHN tumors in a retrospective single-center study. MATERIALS AND METHODS: Oropharyngeal tumor samples from 45 patients (34 males, 11 females) were analyzed. Tumor samples were examined for HPV infection using a two-step PCR. p16 staining by immunohistochemistry was then performed. RESULTS: Samples with strong p16 signal were typed HPV-16-positive. Out of 14 tumor samples with HPV-positive PCR results, 13 samples contained the high risk variant HPV-16. In one sample, HPV-6 DNA was detected. All HPV-16-positive tumors overexpressed p16 (p16(+++)), whereas the HPV-6 sample was p16-negative. CONCLUSION: p16 is not a surrogate marker for replacing PCR testing, but both methods in combination, PCR and immunohistochemistry, could lead to a higher diagnostic validation.