Temporal Bone Fractures and Related Complications in Pediatric and Adult Cranio-Facial Trauma: A Comparison of MDCT Findings in the Acute Emergency Setting.

PURPOSE: The purpose of this study was to analyze the prevalence of and complications resulting from temporal bone fractures in adult and pediatric patients evaluated for cranio-facial trauma in an emergency setting. METHODS: A retrospective blinded analysis of CT scans of a series of 294 consecutive adult and pediatric patients with cranio-facial trauma investigated in the emergency setting was conducted. Findings were compared between the two populations. Preliminary reports made by on-call residents were compared with the retrospective analysis, which was performed in consensus by two experienced readers and served as reference standard. RESULTS: CT revealed 126 fractures in 116/294 (39.5%) patients, although fractures were clinically suspected only in 70/294 (23.8%); p < 0.05. Fractures were longitudinal, transverse and mixed in 69.5%, 10.3% and 19.8% of cases, respectively. Most fractures were otic-sparing fractures (95.2%). Involvement of the external auditory canal, ossicular chain and the osseous structures surrounding the facial nerve was present in 72.2%, 8.7% and 6.3% of cases, respectively. Temporal bone fractures extended into the venous sinuses/jugular foramen and carotid canal in 18.3% and 17.5% of cases, respectively. Vascular injuries (carotid dissection and venous thrombosis) were more common in children than in adults (13.6% versus 5.3%); however, the observed difference did not reach statistical significance. 79.5% of patients with temporal bone fractures had both brain injuries and fractures of the facial bones and cranial vault. Brain injuries were more common in adults (90.4%) than in children (63.6%), p = 0.001. Although on-call residents reliably detected temporal bone fractures (sensitivity = 92.8%), they often missed trauma-associated ossicular dislocation (sensitivity = 27.3%). CONCLUSIONS: Temporal bone fractures and related complications are common in patients with cranio-facial trauma and need to be thoroughly looked for; the pattern of associated injuries is slightly different in children and in adults.

Functional MRI Neurofeedback Outperforms Cognitive Behavioral Therapy for Reducing Tinnitus Distress: A Prospective Randomized Clinical Trial.

Background Cognitive behavioral therapy (CBT) is the current standard treatment for chronic severe tinnitus; however, preliminary evidence suggests that real-time functional MRI (fMRI) neurofeedback therapy may be more effective. Purpose To compare the efficacy of real-time fMRI neurofeedback against CBT for reducing chronic tinnitus distress. Materials and Methods In this prospective controlled trial, participants with chronic severe tinnitus were randomized from December 2017 to December 2021 to receive either CBT (CBT group) for 10 weekly group sessions or real-time fMRI neurofeedback (fMRI group) individually during 15 weekly sessions. Change in the Tinnitus Handicap Inventory (THI) score (range, 0-100) from baseline to 6 or 12 months was assessed. Secondary outcomes included four quality-of-life questionnaires (Beck Depression Inventory, Pittsburgh Sleep Quality Index, State-Trait Anxiety Inventory, and World Health Organization Disability Assessment Schedule). Questionnaire scores between treatment groups and between time points were assessed using repeated measures analysis of variance and the nonparametric Wilcoxon signed rank test. Results The fMRI group included 21 participants (mean age, 49 years ± 11.4 [SD]; 16 male participants) and the CBT group included 22 participants (mean age, 53.6 years ± 8.8; 16 male participants). The fMRI group showed a greater reduction in THI scores compared with the CBT group at both 6 months (mean score change, -28.21 points ± 18.66 vs -12.09 points ± 18.86; P = .005) and 12 months (mean score change, -30 points ± 25.44 vs -4 points ± 17.2; P = .01). Compared with baseline, the fMRI group showed improved sleep (mean score, 8.62 points ± 4.59 vs 7.25 points ± 3.61; P = .006) and trait anxiety (mean score, 44 points ± 11.5 vs 39.84 points ± 10.5; P = .02) at 1 month and improved depression (mean score, 13.71 points ± 9.27 vs 6.53 points ± 5.17; P = .01) and general functioning (mean score, 24.91 points ± 17.05 vs 13.06 points ± 10.1; P = .01) at 6 months. No difference in these metrics over time was observed for the CBT group (P value range, .14 to >.99). Conclusion Real-time fMRI neurofeedback therapy led to a greater reduction in tinnitus distress than the current standard treatment of CBT. ClinicalTrials.gov registration no.: NCT05737888; Swiss Ethics registration no.: BASEC2017-00813 © RSNA, 2024 Supplemental material is available for this article.

Axonal Regrowth of Olfactory Sensory Neurons In Vitro.

One of the most prevalent causes of olfactory loss includes traumatic brain injury with subsequent shearing of olfactory axons at the level of the cribriform plate (anterior skull base). Scar tissue at this level may prevent axonal regrowth toward the olfactory bulb. Currently, there is no cure for this debilitating and often permanent condition. One promising therapeutic concept is to implant a synthetic scaffold with growth factors through the cribriform plate/scar tissue to induce neuroregeneration. The first step toward this goal is to investigate the optimum conditions (growth factors, extracellular matrix proteins) to boost this regeneration. However, the lack of a specifically tailored in vitro model and an automated procedure for quantifying axonal length limits our ability to address this issue. The aim of this study is to create an automated quantification tool to measure axonal length and to determine the ideal growth factors and extracellular proteins to enhance axonal regrowth of olfactory sensory neurons in a mouse organotypic 2D model. We harvested olfactory epithelium (OE) of C57BL/6 mice and cultured them during 15 days on coverslips coated with various extracellular matrix proteins (Fibronectin, Collagen IV, Laminin, none) and different growth factors: fibroblast growth factor 2 (FGF2), brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), nerve growth factor (NGF), retinoic acid (RA), transforming growth factor ß (TGFß), and none. We measured the attachment rate on coverslips, the presence of cellular and axonal outgrowth, and finally, the total axonal length with a newly developed automated high-throughput quantification tool. Whereas the coatings did not influence attachment and neuronal outgrowth rates, the total axonal length was enhanced on fibronectin and collagen IV (p = 0.001). The optimum growth factor supplementation media to culture OE compared to the control condition were as follows: FGF2 alone and FGF2 from day 0 to 7 followed by FGF2 in combination with NGF from day 7 to 15 (p < 0.0001). The automated quantification tool to measure axonal length outperformed the standard Neuron J application by reducing the average analysis time from 22 to 3 min per specimen. In conclusion, robust regeneration of murine olfactory neurons in vitro can be induced, controlled, and efficiently measured using an automated quantification tool. These results will help advance the therapeutic concept closer toward preclinical studies.

3D-printing a cost-effective model for mastoidectomy training.

BACKGROUND: 3D-printed temporal bone models can potentially provide a cost-effective alternative to cadaver surgery that can be manufactured locally at the training department. The objective of this study was to create a cost-effective 3D-printed model suitable for mastoidectomy training using entry level and commercially available print technologies, enabling individuals, without prior experience on 3D-printing, to manufacture their own models for basic temporal bone training. METHODS: Expert technical professionals and an experienced otosurgeon identified the best material for replicating the temporal bone and created a cost-effective printing routine for the model using entry-level print technologies. Eleven participants at a temporal bone dissection course evaluated the model using a questionnaire. RESULTS: The 3D-printed temporal bone model was printed using a material extrusion 3D-printer with a heat resistant filament, reducing melting during drilling. After printing, a few simple post-processing steps were designed to replicate the dura, sigmoid sinus and facial nerve. Modifying the 3D-printer by installing a direct-drive and ruby nozzle resulted in more successful prints and less need for maintenance. Upon evaluation by otorhinolaryngology trainees, unanimous feedback was that the model provided a good introduction to the mastoidectomy procedure, and supplementing practice to cadaveric temporal bones. CONCLUSION: In-house production of a cost-effective 3D-printed model for temporal bone training is feasible and enables training institutions to manufacture their own models. Further, this work demonstrates the feasibility of creating new temporal bone models with anatomical variation to provide ample training opportunity.

Closing the Gap between the Auditory Nerve and Cochlear Implant Electrodes: Which Neurotrophin Cocktail Performs Best for Axonal Outgrowth and Is Electrical Stimulation Beneficial?

Neurotrophins promote neurite outgrowth of auditory neurons and may help closing the gap to cochlear implant (CI) electrodes to enhance electrical hearing. The best concentrations and mix of neurotrophins for this nerve regrowth are unknown. Whether electrical stimulation (ES) during outgrowth is beneficial or may direct axons is another open question. Auditory neuron explant cultures of distinct cochlear turns of 6-7 days old mice were cultured for four days. We tested different concentrations and combinations of BDNF and NT-3 and quantified the numbers and lengths of neurites with an advanced automated analysis. A custom-made 24-well electrical stimulator based on two bulk CIs served to test different ES strategies. Quantification of receptors trkB, trkC, p75NTR, and histological analysis helped to analyze effects. We found 25 ng/mL BDNF to perform best, especially in basal neurons, a negative influence of NT-3 in combined BDNF/NT-3 scenarios, and tonotopic changes in trk and p75NTR receptor stainings. ES largely impeded neurite outgrowth and glia ensheathment in an amplitude-dependent way. Apical neurons showed slight benefits in neurite numbers and length with ES at 10 and 500 µA. We recommend BDNF as a potent drug to enhance the man-machine interface, but CIs should be better activated after nerve regrowth.

[Molecular genetic diagnosis in children with cochlear implants in the Western french speaking part of Switzerland]. / Diagnostic génétique moléculaire des enfants implantés cochléaires en Suisse romande.

Hearing loss is the most frequent sensory deficit at birth. Newborn hearing screening helps with early identification and clinical management of hearing deficits. A cochlear implantation is advised for profound hearing loss. Previously, an etiologic diagnosis was difficult to obtain, and many laboratory tests were required. Today, genetics has up to 60% success rate in etiologic diagnosis and is now part of the international pediatric ENT recommendations. The Centre Universitaire Romand des Implants Cochléaires (CURIC) follows children with cochlear implants. From 2015 to 2021, 26 implanted children received testing, with a 73% success rate. The genetic diagnosis helped guide their clinical management and helped to avoid unnecessary and costly clinical testing.

Le déficit auditif (DA) est le déficit neurosensoriel le plus fréquent à la naissance. Le dépistage auditif permet l'identification et la prise en charge précoces des problèmes d'audition. Dans le cas de surdités profondes, une implantation cochléaire est conseillée. Auparavant, le diagnostic étiologique était difficile à poser malgré de nombreux examens complémentaires. Depuis 10 ans, la médecine génétique aboutit à un diagnostic étiologique dans 60% des cas et fait partie des recommandations internationales d'ORL pédiatrique. Le Centre universitaire romand des implants cochléaires prend en charge les enfants implantés. Entre 2015 et 2021, 26 enfants implantés ont eu une analyse génétique, dont 73% avec succès. Ceci permet d'orienter la prise en charge spécifiquement au profil génétique et diminue les examens complémentaires.

Diagnostic value and prognostic significance of MRI findings in sudden sensorineural hearing loss.

Purpose: We evaluated the clinical significance of magnetic resonance imaging (MRI) findings and their prognostic value for initial hearing loss and recovery in patients with sudden sensorineural hearing loss (SSNHL). Materials and Methods: This retrospective study included consecutive adult patients with unilateral SSNHL, contrast-enhanced MRI and audiometric testing evaluated in our institution between 2005 and 2017. MRI reports, patient data, treatment, and audiometric tests were reviewed, with the relationship between MRI findings and hearing loss/recovery analyzed. Results: Overall, 266 patients were included. Additional symptoms comprised tinnitus (114/266; 43%), vertigo (45/266; 17%), ear pain (26/266; 10%), and ear pressure (6/266; 2%). At least one cardiovascular risk factor (hypertension, diabetes, hypercholesterolemia, cardiopathy, and active smoking) existed in 167/266 (63%) patients. Corticosteroid treatment was followed by 198/266 (74%) patients while contraindications/refusal/compliance precluded treatment in 68/266(26%). Complete, partial or slight hearing recovery occurred in 167/266 (63%) patients. Three MRI patient groups were identified: a group with normal MRI examinations or incidentalomas (128/266; 48%), a group with peripheral auditory system (PAS) lesions (95/266; 36%), and a group with central nervous system (CNS) lesions (43/266; 16%). PAS lesions included lesions from the cochlea to the brain stem (e.g., schwannoma, meningioma, labyrinthitis, intracochlear hemorrhage, vestibulocochlear neuritis), whereas CNS lesions corresponded in 42/43(98%) of cases to leukoaraiosis and other vascular lesions (e.g., stroke, hemorrhage, aneurysm, venous sinus thrombosis, and cavernoma). Belonging to one of the three MRI groups did not influence the degree of initial hearing loss, affected frequencies or treatment, p > .05. Gender and cardiovascular risk factors did neither affect initial hearing loss nor recovery. However, age > 70 years negatively affected initial hearing loss in all frequencies, as well as recovery in all frequencies except 1000 Hz. Also, poor recovery of initial high-frequency hearing loss (>1000 Hz) was significantly associated with CNS lesions. Conclusion: Age > 70 years and CNS lesions depicted by MRI independently predicted poor auditory recovery, albeit in different frequencies. Lay Summary: In patients with sudden hearing loss, older age (above 70 years) predicts poorer hearing recovery than in younger patients in most hearing frequencies. In addition, abnormalities of brain tissue revealed by MRI predict poorer hearing recovery at high frequencies. Level of Evidence: Level III.

Development and in vivo validation of small interfering RNAs targeting NOX3 to prevent sensorineural hearing loss.

The reactive oxygen species (ROS)-generating enzyme NOX3 has recently been implicated in the pathophysiology of several acquired forms of sensorineural hearing loss, including cisplatin-, noise- and age-related hearing loss. NOX3 is highly and specifically expressed in the inner ear and therefore represents an attractive target for specific intervention aiming at otoprotection. Despite the strong rationale to inhibit NOX3, there is currently no specific pharmacological inhibitor available. Molecular therapy may represent a powerful alternative. In this study, we developed and tested a collection of small interfering (si) RNA constructs to establish a proof of concept of NOX3 inhibition through local delivery in the mouse inner ear. The inhibitory potential of 10 different siRNA constructs was first assessed in three different cells lines expressing the NOX3 complex. Efficacy of the most promising siRNA construct to knock-down NOX3 was then further assessed in vivo, comparing middle ear delivery and direct intracochlear delivery through the posterior semi-circular canal. While hearing was completely preserved through the intervention, a significant downregulation of NOX3 expression in the mouse inner ear and particularly in the spiral ganglion area at clinically relevant levels (>60%) was observed 48 h after treatment. In contrast to successful intracochlear delivery, middle ear administration of siRNA failed to significantly inhibit Nox3 mRNA expression. In conclusion, intracochlear delivery of NOX3-siRNAs induces a robust temporal NOX3 downregulation, which could be of relevance to prevent predictable acute insults such as cisplatin chemotherapy-mediated ototoxicity and other forms of acquired hearing loss, including post-prevention of noise-induced hearing loss immediately after trauma. Successful translation of our concept into an eventual clinical use in humans will depend on the development of atraumatic and efficient delivery routes into the cochlea without a risk to induce hearing loss through the intervention.

Mechanically evoked tinnitus after cochlear implantation with preservation of residual hearing.

In the recent past, inclusion criteria for cochlear implant surgery expanded to patients with ever more residual acoustic hearing in the low frequencies. By applying the meticulous hearing preservation surgical strategy and specifically designed atraumatic electrode arrays, residual hearing can be preserved to a meaningful extent in a large majority of patients. In this paper, we describe two female patients suffering from mechanically evoked tinnitus after hearing preservation cochlear implantation surgery with MEDEL flex electrodes. The occurrence of audible perceptions through mechanical stimulation in the region of the external ear is believed to be due to the direct transmission of movements via the electrode array to the basilar membrane of the inner ear. In both cases, the mechanically evoked tinnitus led to revision surgery with immobilization of the array in the mastoid cavity. Despite eliminating the tinnitus, the revision surgery led to a loss of residual hearing in one patient, whereas the relatively poor residual hearing in the other revision case remained unchanged. The presence of mechanically evoked tinnitus seems to be associated with increased fragility of inner ear structures and hearing function, possibly due to direct mechanical contact of the electrode array with the basilar membrane. Consequently, the electrode array needs to be carefully immobilized in the mastoid cavity at a distance from soft tissue to prevent mechanical damage of inner ear structures, particularly in female patients with fine muscular tissue.

WNT Activation and TGFß-Smad Inhibition Potentiate Stemness of Mammalian Auditory Neuroprogenitors for High-Throughput Generation of Functional Auditory Neurons In Vitro.

Hearing loss affects over 460 million people worldwide and is a major socioeconomic burden. Both genetic and environmental factors (i.e., noise overexposure, ototoxic drug treatment and ageing), promote the irreversible degeneration of cochlear hair cells and associated auditory neurons, leading to sensorineural hearing loss. In contrast to birds, fish and amphibians, the mammalian inner ear is virtually unable to regenerate due to the limited stemness of auditory progenitors, and no causal treatment is able to prevent or reverse hearing loss. As of today, a main limitation for the development of otoprotective or otoregenerative therapies is the lack of efficient preclinical models compatible with high-throughput screening of drug candidates. Currently, the research field mainly relies on primary organotypic inner ear cultures, resulting in high variability, low throughput, high associated costs and ethical concerns. We previously identified and characterized the phoenix auditory neuroprogenitors (ANPGs) as highly proliferative progenitor cells isolated from the A/J mouse cochlea. In the present study, we aim at identifying the signaling pathways responsible for the intrinsic high stemness of phoenix ANPGs. A transcriptomic comparison of traditionally low-stemness ANPGs, isolated from C57Bl/6 and A/J mice at early passages, and high-stemness phoenix ANPGs was performed, allowing the identification of several differentially expressed pathways. Based on differentially regulated pathways, we developed a reprogramming protocol to induce high stemness in presenescent ANPGs (i.e., from C57Bl6 mouse). The pharmacological combination of the WNT agonist (CHIR99021) and TGFß/Smad inhibitors (LDN193189 and SB431542) resulted in a dramatic increase in presenescent neurosphere growth, and the possibility to expand ANPGs is virtually limitless. As with the phoenix ANPGs, stemness-induced ANPGs could be frozen and thawed, enabling distribution to other laboratories. Importantly, even after 20 passages, stemness-induced ANPGs retained their ability to differentiate into electrophysiologically mature type I auditory neurons. Both stemness-induced and phoenix ANPGs resolve a main bottleneck in the field, allowing efficient, high-throughput, low-cost and 3R-compatible in vitro screening of otoprotective and otoregenerative drug candidates. This study may also add new perspectives to the field of inner ear regeneration.

NADPH Oxidase 3 Deficiency Protects From Noise-Induced Sensorineural Hearing Loss.

The reactive oxygen species (ROS)-generating NADPH oxidase NOX3 isoform is highly and specifically expressed in the inner ear. NOX3 is needed for normal vestibular development but NOX-derived ROS have also been implicated in the pathophysiology of sensorineural hearing loss. The role of NOX-derived ROS in noise-induced hearing loss, however, remains unclear and was addressed with the present study. Two different mouse strains, deficient in NOX3 or its critical subunit p22phox, were subjected to a single noise exposure of 2 h using an 8-16 kHz band noise at an intensity of 116-120 decibel sound pressure level. In the hours following noise exposure, there was a significant increase in cochlear mRNA expression of NOX3 in wild type animals. By using RNAscope in situ hybridization, NOX3 expression was primarily found in the Rosenthal canal area, colocalizing with auditory neurons. One day after the noise trauma, we observed a high frequency hearing loss in both knock-out mice, as well as their wild type littermates. At day seven after noise trauma however, NOX3 and p22phox knockout mice showed a significantly improved hearing recovery and a marked preservation of neurosensory cochlear structures compared to their wild type littermates. Based on these findings, an active role of NOX3 in the pathophysiology of noise-induced hearing loss can be demonstrated, in line with recent evidence obtained in other forms of acquired hearing loss. The present data demonstrates that the absence of functional NOX3 enhances the hearing recovery phase following noise trauma. This opens an interesting clinical window for pharmacological or molecular intervention aiming at post prevention of noise-induced hearing loss.

Phoenix auditory neurons as 3R cell model for high throughput screening of neurogenic compounds.

Auditory neurons connect the sensory hair cells from the inner ear to the brainstem. These bipolar neurons are relevant targets for pharmacological intervention aiming at protecting or improving the hearing function in various forms of sensorineural hearing loss. In the research laboratory, neurotrophic compounds are commonly used to improve survival and to promote regeneration of auditory neurons. One important roadblock delaying eventual clinical applications of these strategies in humans is the lack of powerful in vitro models allowing high throughput screening of otoprotective and regenerative compounds. The recently discovered auditory neuroprogenitors (ANPGs) derived from the A/J mouse with an unprecedented capacity to self-renew and to provide mature auditory neurons offer the possibility to overcome this bottleneck. In the present study, we further characterized the new phoenix ANPGs model and compared it to the current gold-standard spiral ganglion organotypic explant (SGE) model to assay neurite outgrowth, neurite length and glutamate-induced Ca2+ response in response to neurotrophin-3 (NT-3) and brain derived neurotrophic factor (BDNF) treatment. Whereas both, SGEs and phoenix ANPGs exhibited a robust and sensitive response to neurotrophins, the phoenix ANPGs offer a considerable range of advantages including high throughput suitability, lower experimental variability, single cell resolution and an important reduction of animal numbers. The phoenix ANPGs in vitro model therefore provides a robust high-throughput platform to screen for otoprotective and regenerative neurotrophic compounds in line with 3R principles and is of interest for the field of auditory neuroscience.

[Superior canal dehiscence syndrome]. / Syndrome de déhiscence du canal semi-circulaire supérieur.

Superior canal dehiscence syndrome is due to a defect in the bone coverage of this canal. The presence of this « third window ¼ alters the biomechanics of the inner ear and can induce cochleo-vestibular symptoms, most commonly bone conduction hyperacusis, autophony, pulsatile tinnitus, and pressure- or sound-induced vertigo. Although the etiology remains unknown, it could be congenital. Head trauma or a Valsalva maneuver may also trigger the symptoms. The diagnosis is based on the association of specific symptoms, a characteristic auditory and vestibular workup and the presence of a dehiscence on a high-resolution CT-scan. Management can be medical or surgical in selected cases.

Le syndrome de déhiscence du canal semi-circulaire supérieur est dû à un défaut de couverture osseuse de ce canal. La présence de cette « troisième fenêtre ¼ modifie la biomécanique de l'oreille interne et peut induire des symptômes cochléovestibulaires, le plus souvent une hyperacousie en conduction osseuse, une autophonie, un acouphène pulsatile et des vertiges induits par un changement de pression ou par des sons. Même si l'étiologie reste inconnue, elle pourrait être congénitale. Un traumatisme crânien ou une manœuvre de Valsalva peuvent aussi être à l'origine des symptômes. Le diagnostic est basé sur l'association de symptômes spécifiques, d'un bilan auditif et vestibulaire caractéristique et de la présence d'une déhiscence sur un CT-scan de haute résolution. La prise en charge peut, dans certains cas, être médicale ou chirurgicale.

[Treatment of single-sided deafness by cochlear implantation]. / Traitement de la surdité unilatérale par l'implant cochléaire.

Single sided deafness diminishes speech understanding in noise and sound localization and thereby globally auditory performance. Most patients also suffer from tinnitus and indicate reduced quality of life. Patients have the choice to adapt to the new situation without treatment, to restore pseudostereophonic hearing by contralateral routing of signal (CROS) hearing aids or to restore binaural hearing using a cochlear implant in the deaf ear. This article summarizes the physiological base of binaural hearing and treatment options for single sided deafness with a special emphasis on the cochlear implant.

La surdité unilatérale diminue la compréhension dans le bruit et la capacité de localiser les sources sonores affectant ainsi globalement la performance auditive. De plus, la qualité de vie est souvent impactée par la présence d'un acouphène dérangeant. Les patients ont le choix de s'adapter à la situation sans traitement ou de reconstituer une pseudo-stéréophonie à l'aide d'un appareillage qui transmet l'information auditive arrivant sur l'oreille sourde à l'oreille saine (Contralateral Routing of Signals (CROS)). L'implant cochléaire est une alternative récente qui permet de « réactiver ¼ l'oreille atteinte et de redonner ainsi une audition binaurale. Les différentes options de réhabilitation auditive en cas de surdité unilatérale en mettant l'accent sur l'implant cochléaire sont discutées à l'aide d'un cas clinique.

ExplantAnalyzer: An advanced automated neurite outgrowth analysis evaluated by means of organotypic auditory neuron explant cultures.

BACKGROUND: Neuronal outgrowth assays using organotypic explant cultures are commonly utilized to study neuroregenerative and -protective effects of drugs such as neurotrophins. While this approach offers higher organized tissue compared to single cell cultures and less experimental effort than in-vivo studies, quantitative evaluation of the neuronal network is often time consuming. Thus, we developed ExplantAnlayzer, a time-saving high-throughput evaluation method, yielding numerous metrics to objectively describe neuronal outgrowth. NEW METHOD: Spiral ganglion explants were cultured in 24-well plates, mechanically fixed in a collagen matrix and immunolabeled against beta-III-tubulin. The explants were imaged using a fluorescent tile-scan microscope and resulting images were stitched. The evaluation was developed as an open-source MATLAB routine and involves several image processing steps, including adaptive thresholding. The neurite network was eventually converted to a graph to track neurites from their terminals back to the explant body. COMPARISON WITH EXISTING METHOD(S): We compared ExplantAnlayzer quantitatively and qualitatively to common existing methods, such as Sholl analyses and manual fiber tracing, using representative explant images. ExplantAnlayzer is able to achieve similar and as detailed results as manual tracing while decreasing manual interaction and required time dramatically. RESULTS: After an initial setup phase, the explant images could be batch-processed altogether. Bright bundles as well as faint fibers were reliably detected. Several metrics describing the outgrowth morphology, including total outgrowth, neurite numbers and length estimations, as well as their growth directions, were computed. CONCLUSIONS: ExplantAnalyzer is a time-saving and objective method for an in-depth evaluation of organotypic explant outgrowth.

Local Cisplatin Delivery in Mouse Reliably Models Sensorineural Ototoxicity Without Systemic Adverse Effects.

Cisplatin is a lifesaving chemotherapeutic drug with marked ototoxic adverse effects. Cisplatin-induced hearing loss affects a significant part of cancer-surviving patients and is an unmet clinical need with important socioeconomic consequences. Unfortunately, in current preclinical animal models of cisplatin ototoxicity, which are mainly based on systemic delivery, important morbidity is observed, leading to premature death. This methodology not only raises obvious animal welfare concerns but also increases the number of animals used in ototoxicity studies to compensate for dropouts related to early death. To overcome these important limitations, we developed a local delivery model based on the application of a cisplatin solution directly into the otic bulla through a retroauricular approach. The local delivery model reliably induced significant hearing loss with a mean threshold shift ranging from 10 to 30 dB, strongly affecting the high frequencies (22 and 32 kHz). Importantly, mice did not show visible stress or distress indicators and no significant morbidity in comparison with a traditional systemic delivery control group of mice injected intraperitoneally with 10 mg/kg cisplatin, where significant weight loss >10% in all treated animals (without any recovery) led to premature abortion of experiments on day 3. Mass spectrometry confirmed the absence of relevant systemic uptake after local delivery, with platinum accumulation restricted to the cochlea, whereas important platinum concentrations were detected in the liver and kidney of the systemic cisplatin group. A clear correlation between the cochlear platinum concentration and the auditory threshold shift was observed. Immunohistochemistry revealed statistically significant loss of outer hair cells in the basal and apical turns of the cochlea and an important and statistically significant loss of auditory neurons and synapses in all cochlear regions. In conclusion, local cisplatin delivery induces robust hearing loss with minimal morbidity, thereby offering a reliable rodent model for human cisplatin ototoxicity, reducing the number of animals required and showing improved animal welfare compared with traditional systemic models.

Benefits of Exome Sequencing in Children with Suspected Isolated Hearing Loss.

PURPOSE: Hearing loss is characterized by an extensive genetic heterogeneity and remains a common disorder in children. Molecular diagnosis is of particular benefit in children, and permits the early identification of clinically-unrecognized hearing loss syndromes, which permits effective clinical management and follow-up, including genetic counselling. METHODS: We performed whole-exome sequencing with the analysis of a panel of 189 genes associated with hearing loss in a prospective cohort of 61 children and 9 adults presenting mainly with isolated hearing loss. RESULTS: The overall diagnostic rate using exome sequencing was 47.2% (52.5% in children; 22% in adults). In children with confirmed molecular results, 17/32 (53.2%) showed autosomal recessive inheritance patterns, 14/32 (43.75%) showed an autosomal dominant condition, and one case had X-linked hearing loss. In adults, the two patients showed an autosomal dominant inheritance pattern. Among the 32 children, 17 (53.1%) had nonsyndromic hearing loss and 15 (46.7%) had syndromic hearing loss. One adult was diagnosed with syndromic hearing loss and one with nonsyndromic hearing loss. The most common causative genes were STRC (5 cases), GJB2 (3 cases), COL11A1 (3 cases), and ACTG1 (3 cases). CONCLUSIONS: Exome sequencing has a high diagnostic yield in children with hearing loss and can reveal a syndromic hearing loss form before other organs/systems become involved, allowing the surveillance of unrecognized present and/or future complications associated with these syndromes.