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Several lines of evidence have suggested that steeply sloping audiometric losses are caused by hair cell degeneration, while flat audiometric losses are caused by strial atrophy, but this concept has never been rigorously tested in human specimens. Here, we systematically compare audiograms and cochlear histopathology in 160 human cases from the archival collection of celloidin-embedded temporal bones at the Massachusetts Eye and Ear. The dataset included 106 cases from a prior study of normal-aging ears, and an additional 54 cases selected by combing the database for flat audiograms. Audiogram shapes were classified algorithmically into five groups according to the relation between flatness (i.e., SD of hearing levels across all frequencies) and low-frequency pure-tone average (i.e., mean at 0.25, 0.5, and 1.0 kHz). Outer and inner hair cell losses, neural degeneration, and strial atrophy were all quantified as a function of cochlear location in each case. Results showed that strial atrophy was worse in the apical than the basal half of the cochlea and was worse in females than in males. The degree of strial atrophy was uncorrelated with audiogram flatness. Apical atrophy was correlated with low-frequency thresholds and basal atrophy with high-frequency thresholds, and the former correlation was higher. However, a multivariable regression with all histopathological measures as predictors and audiometric thresholds as the outcome showed that strial atrophy was a significant predictor of threshold shift only in the low-frequency region, and, even there, the contribution of outer hair cell damage was larger.SIGNIFICANCE STATEMENT Cochlear pathology can only be assessed postmortem; thus, human cochlear histopathology is critical to our understanding of the mechanisms of hearing loss. Dogma holds that relative damage to sensory cells, which transduce mechanical vibration into electrical signals, versus the stria vascularis, the cellular battery that powers transduction, can be inferred by the shape of the audiogram, that is, down-sloping (hair cell damage) versus flat (strial atrophy). Here we quantified hair cell and strial atrophy in 160 human specimens to show that it is the degree of low-frequency hearing loss, rather than the audiogram slope, that predicts strial atrophy. Results are critical to the design of clinical trials for hearing-loss therapeutics, as current drugs target only hair cell, not strial, regeneration.
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Sordera , Estría Vascular , Masculino , Femenino , Humanos , Estría Vascular/patología , Cóclea/patología , Sordera/patología , Atrofia/patología , Células Ciliadas Auditivas Externas/patologíaRESUMEN
Animal studies suggest that cochlear nerve degeneration precedes sensory cell degeneration in both noise-induced hearing loss (NIHL) and age-related hearing loss (ARHL), producing a hearing impairment that is not reflected in audiometric thresholds. Here, we investigated the histopathology of human ARHL and NIHL by comparing loss of auditory nerve fibers (ANFs), cochlear hair cells and the stria vascularis in a group of 52 cases with noise-exposure history against an age-matched control group. Although strial atrophy increased with age, there was no effect of noise history. Outer hair cell (OHC) loss also increased with age throughout the cochlea but was unaffected by noise history in the low-frequency region (<2 kHz), while greatly exacerbated at high frequencies (≥2 kHz). Inner hair cell (IHC) loss was primarily seen at high frequencies but was unaffected by noise at either low or high frequencies. ANF loss was substantial at all cochlear frequencies and was exacerbated by noise throughout. According to a multivariable regression model, this loss of neural channels contributes to poor word discrimination among those with similar audiometric threshold losses. The histopathological patterns observed also suggest that, whereas the low-frequency OHC loss may be an unavoidable consequence of aging, the high-frequency loss, which produces the classic down-sloping audiogram of ARHL, may be partially because of avoidable ear abuse, even among those without a documented history of acoustic overexposure.SIGNIFICANCE STATEMENT As regenerative therapeutics in sensorineural hearing loss enter clinical trials, it becomes critical to infer which cochlear pathologies are present in addition to hair cell loss. Here, by analyzing human autopsy material, we show that acoustic injury accelerates age-related primary neural degeneration, but not strial degeneration, neither of which can be inferred from audiometric thresholds. It exacerbates outer hair cell (OHC) loss only in the high-frequency half of the cochlea, suggesting that the apical loss is age-related, whereas the basal loss is partially noise induced, and therefore avoidable. Statistical analysis suggests that neural loss helps explain differences in word-recognition ability among individuals with similar audiometric thresholds. The surprising correlation between neural loss and OHC loss in the cochlea's speech region also implicates neural loss in the well-known decline in word scores as thresholds deteriorate with age.
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Cóclea/patología , Células Ciliadas Auditivas Externas/patología , Pérdida Auditiva Provocada por Ruido/patología , Degeneración Nerviosa/patología , Ruido/efectos adversos , Adulto , Anciano , Anciano de 80 o más Años , Envejecimiento/patología , Umbral Auditivo/fisiología , Nervio Coclear/patología , Femenino , Células Ciliadas Auditivas Internas , Humanos , Masculino , Persona de Mediana Edad , Degeneración Nerviosa/etiologíaRESUMEN
Age-related hearing loss arises from irreversible damage in the inner ear, where sound is transduced into electrical signals. Prior human studies suggested that sensory-cell loss is rarely the cause; correspondingly, animal work has implicated the stria vascularis, the cellular "battery" driving the amplification of sound by hair cell "motors." Here, quantitative microscopic analysis of hair cells, auditory nerve fibers, and strial tissues in 120 human inner ears obtained at autopsy, most of whom had recent audiograms in their medical records, shows that the degree of hearing loss is well predicted from the amount of hair cell loss and that inclusion of strial damage does not improve the prediction. Although many aging ears showed significant strial degeneration throughout the cochlea, our statistical models suggest that, by the time strial tissues are lost, hair cell death is so extensive that the loss of battery is no longer important to pure-tone thresholds and that audiogram slope is not diagnostic for strial degeneration. These data comprise the first quantitative survey of hair cell death in normal-aging human cochleas, and reveal unexpectedly severe hair cell loss in low-frequency cochlear regions, and dramatically greater loss in high-frequency regions than seen in any aging animal model. Comparison of normal-aging ears to an age-matched group with acoustic-overexposure history suggests that a lifetime of acoustic overexposure is to blame.SIGNIFICANCE STATEMENT This report upends dogma about the causes of age-related hearing loss. Our analysis of over 120 autopsy specimens shows that inner-ear sensory cell loss can largely explain the audiometric patterns in aging, with minimal contribution from the stria vascularis, the "battery" that powers the inner ear, previously viewed as the major locus of age-related hearing dysfunction. Predicting inner ear damage from the audiogram is critical, now that clinical trials of therapeutics designed to regrow hair cells are underway. Our data also show that hair cell degeneration in aging humans is dramatically worse than that in aging animals, suggesting that the high-frequency hearing losses that define human presbycusis reflect avoidable contributions of chronic ear abuse to which aging animals are not exposed.
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Células Ciliadas Auditivas Internas/patología , Presbiacusia/patología , Estría Vascular/patología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Audiometría , Vías Auditivas/patología , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Persona de Mediana Edad , Presbiacusia/etiología , Adulto JovenRESUMEN
Meniere's disease (MD), a syndromal inner ear disease, is commonly associated with a pathological accumulation of endolymphatic fluid in the inner ear, termed "idiopathic" endolymphatic hydrops (iEH). Although numerous precipitating/exacerbating factors have been proposed for MD, its etiology remains elusive. Here, using immunohistochemistry and in situ protein-protein interaction detection assays, we demonstrate mineralocorticoid-controlled sodium transport mechanisms in the epithelium of the extraosseous portion of the endolymphatic sac (eES) in the murine and human inner ears. Histological analysis of the eES in an extensive series of human temporal bones consistently revealed pathological changes in the eES in cases with iEH and a clinical history of MD, but no such changes were found in cases with "secondary" EH due to other otological diseases or in healthy controls. Notably, two etiologically different pathologies-degeneration and developmental hypoplasia-that selectively affect the eES in MD were distinguished. Clinical records from MD cases with degenerative and hypoplastic eES pathology revealed distinct intergroup differences in clinical disease presentation. Overall, we have identified for the first time two inner ear pathologies that are consistently present in MD and can be directly linked to the pathogenesis of EH, and which potentially affect the phenotypical presentation of MD.
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Oído Interno/patología , Transporte Iónico/fisiología , Enfermedad de Meniere/metabolismo , Enfermedad de Meniere/patología , Sodio/metabolismo , Animales , Oído Interno/metabolismo , Hidropesía Endolinfática/metabolismo , Hidropesía Endolinfática/patología , Saco Endolinfático/metabolismo , Saco Endolinfático/patología , Humanos , Masculino , Ratones , Hueso Temporal/metabolismo , Hueso Temporal/patologíaRESUMEN
The histopathology of the inner ear in a patient with hearing loss caused by the p.L114P COCH mutation and its correlation with the clinical phenotype are presented. To date, 23 COCH mutations causative of DFNA9 autosomal dominant sensorineural hearing loss and vestibular disorder have been reported, and the histopathology of the human inner ear has been described in 4 of these. The p.L114P COCH mutation was first described in a Korean family. We have identified the same mutation in a family of non-Asian ancestry in the USA, and the temporal bone histopathology and clinical findings are presented herein. The histopathology found in the inner ear was similar to that shown in the 4 other COCH mutations and included degeneration of the spiral ligament with deposition of an eosinophilic acellular material, which was also found in the distal osseous spiral lamina, at the base of the spiral limbus, and in mesenchymal tissue at the base of the vestibular neuroepithelium. This is the first description of human otopathology of the COCH p.L114P mutation. In addition, it is the only case with otopathology characterization in an individual with any COCH mutation and residual hearing, thus allowing assessment of primary histopathological events in DFNA9, before progression to more profound hearing loss. A quantitative cytologic analysis of atrophy in this specimen and immunostaining using anti-neurofilament and anti-myelin protein zero antibodies confirmed that the principal histopathologic correlate of hearing loss was degeneration of the dendritic fibers of spiral ganglion cells in the osseous spiral lamina. The implications for cochlear implantation in this disorder are discussed.
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Oído Interno/patología , Proteínas de la Matriz Extracelular/genética , Pérdida Auditiva Sensorineural/genética , Pérdida Auditiva Sensorineural/patología , Adulto , Humanos , Masculino , Mutación , Hueso Temporal/patologíaRESUMEN
In our laboratory, human temporal bone specimens from patients who in life have undergone cochlear implantation are routinely processed with the implant in situ, embedded in Araldite, sectioned at 20 µm and serially photographed during cutting, stained with toluidine blue and mounted on glass slides. From the images, two-dimensional and three-dimensional reconstructions can be made and a very accurate implant insertion depth can be calculated from the three-dimensional reconstructions. However, this method precludes subsequent special stains and further molecular investigations of the tissue including proteomics and immunostaining, which is now possible with celloidin-embedded tissue. In this study, we correlated measurement of the implant array insertion depth calculated from histologic three-dimensional reconstruction with that measured from three-dimensional radiologic multiplanar reconstruction. Four human temporal bones with cochlear implants underwent postfixation preprocessing CT imaging with a Siemens Somatom Sensation Scanner. The CT scans from these four bones were downloaded into the Voxar software application, reformatted using the multiplanar reconstruction tool, viewed in three dimensions and measurements of intracochlear insertion lengths of the implants were obtained. The bones were processed routinely for in situ Araldite embedding, serial images were made of the block during sectioning, postprocessed using PV-Wave® software, aligned with Amira® software, and used to create histologic three-dimensional reconstructions. From these three-dimensional reconstructions, the insertion depth of the electrode array was mathematically calculated. The range of insertion depths was 15.9 mm (case 1) to 26.6 mm (case 4). The two methods, radiographic multiplanar reconstruction and three-dimensional reconstruction, differed by 0.4-0.9%. This provides confidence that important localization information about the electrode in situ can be gleaned from CT scans, thereby allowing us to extract the implants prior to processing for celloidin embedment and allow further techniques such as special stains and immunostaining to be accomplished in order to evaluate molecular mechanisms involved in cochlear implantation.
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Implantación Coclear , Implantes Cocleares , Sordera/cirugía , Hueso Temporal/cirugía , Sordera/diagnóstico por imagen , Sordera/patología , Humanos , Procesamiento de Imagen Asistido por Computador , Radiografía , Hueso Temporal/diagnóstico por imagen , Hueso Temporal/patologíaRESUMEN
Delayed loss of residual acoustic hearing after cochlear implantation is a common but poorly understood phenomenon due to the scarcity of relevant temporal bone tissues. Prior histopathological analysis of one case of post-implantation hearing loss suggested there were no interaural differences in hair cell or neural degeneration to explain the profound loss of low-frequency hearing on the implanted side (Quesnel et al., 2016) and attributed the threshold elevation to neo-ossification and fibrosis around the implant. Here we re-evaluated the histopathology in this case, applying immunostaining and improved microscopic techniques for differentiating surviving hair cells from supporting cells. The new analysis revealed dramatic interaural differences, with a > 80 % loss of inner hair cells in the cochlear apex on the implanted side, which can account for the post-implantation loss of residual hearing. Apical degeneration of the stria further contributed to threshold elevation on the implanted side. In contrast, spiral ganglion cell survival was reduced in the region of the electrode on the implanted side, but apical counts in the two ears were similar to that seen in age-matched unimplanted control ears. Almost none of the surviving auditory neurons retained peripheral axons throughout the basal half of the cochlea. Relevance to cochlear implant performance is discussed.
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Umbral Auditivo , Implantación Coclear , Implantes Cocleares , Ganglio Espiral de la Cóclea , Implantación Coclear/instrumentación , Implantación Coclear/efectos adversos , Humanos , Ganglio Espiral de la Cóclea/patología , Ganglio Espiral de la Cóclea/fisiopatología , Células Ciliadas Auditivas Internas/patología , Factores de Tiempo , Supervivencia Celular , Masculino , Audición , Pérdida Auditiva/fisiopatología , Pérdida Auditiva/patología , Pérdida Auditiva/cirugía , Pérdida Auditiva/etiología , Femenino , Células Ciliadas Auditivas/patología , Anciano , Degeneración Nerviosa , Persona de Mediana Edad , Hueso Temporal/patología , Hueso Temporal/cirugíaRESUMEN
Pathology repositories worldwide store millions of celloidin-processed human brain and temporal bone (TB) sections vital for studying central nervous system diseases and sensory organs. However, accessing these sections for modern molecular-pathological research, like immunohistochemistry, is hindered by the challenge of removing celloidin without damaging tissue. In this study, we explored the use of polyethylene glycols (PEGs), a class of non-hazardous, ethylene glycol oligomers, combined with an improved section mounting technique, to gently and effectively dissolve celloidin from sections archived for up to 40 years. Optimizing our protocol involved exploring celloidin dissolution kinetics in PEGs of varying molecular weights and terminations, as well as different temperatures. Low molecular weight PEGs, particularly PEG 200, were the most efficient celloidin solvent. Nuclear magnetic resonance (NMR) spectroscopy of celloidin-PEG 200 dissolution products revealed no chemical alterations, suggesting pure solvation without chemical modification. Because the solvation of celloidin in PEG was inhibited by proteins, we further developed a protein-free mounting protocol allowing complete celloidin removal in 30 to 60 minutes by immersing in PEG 200. In summary, our approach overcomes major methodological hurdles, rendering decades-old archival celloidin sections viable for immunohistochemical and other molecular biological techniques, while enhancing safety and workflow efficiency.
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Encéfalo , Inmunohistoquímica , Polietilenglicoles , Humanos , Polietilenglicoles/química , Inmunohistoquímica/métodos , Encéfalo/metabolismo , Encéfalo/citologíaRESUMEN
Quantifying the survival patterns of spiral ganglion cells (SGCs), the cell bodies of auditory-nerve fibers, is critical to studies of sensorineural hearing loss, especially in human temporal bones. The classic method of manual counting is tedious, and, although stereology approaches can be faster, they can only be used to estimate total cell numbers per cochlea. Here, a machine-learning algorithm that automatically identifies, counts, and maps the SGCs in digitized images of semi-serial human temporal-bone sections not only speeds the analysis, with no loss of accuracy, but also allows 3D visualization of the SGCs and fine-grained mapping to cochlear frequency. Applying the algorithm to 62 normal-aging human ears shows significantly faster degeneration of SGCs in the basal than the apical half of the cochlea. Comparison to fiber counts in the same ears shows that the fraction of surviving SGCs lacking a peripheral axon steadily increases with age, reaching more than 50% in the apical cochlea and almost 66% in basal regions.
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Cóclea , Pérdida Auditiva Sensorineural , Humanos , Envejecimiento , Ganglio Espiral de la Cóclea , Hueso TemporalRESUMEN
Animal studies have shown that the supporting-cells surviving in the organ of Corti after cochlear insult can be transdifferentiated into hair cells as a treatment for sensorineural hearing loss. Clinical trials of small-molecule therapeutics have been undertaken, but little is known about how to predict the pattern and degree of supporting-cell survival based on audiogram, hearing loss etiology or any other metric obtainable pre-mortem. To address this, we systematically assessed supporting-cell and hair cell survival, as a function of cochlear location in 274 temporal bone cases from the archives at the Massachusetts Eye and Ear and compared the histopathology with the audiograms and hearing-loss etiologies. Results showed that supporting-cell survival was always significantly greater in the apical half than the basal half of the cochlea, that inner pillars were more robust than outer pillars or Deiters' cells, and that total replacement of all supporting cells with a flat epithelium was rare outside of the extreme basal 20% of the cochlea. Supporting cell survival in the basal half of the cochlea was better correlated with the slope of the audiogram than with the mean high-frequency threshold per se: i.e. survival was better with flatter audiograms than with steeply down-sloping audiograms. Cochlear regions with extensive hair cell loss and exceptional supporting cell survival were most common in cases with hearing loss due to ototoxic drugs. Such cases also tended to have less pathology in other functionally critical structures, i.e. spiral ganglion neurons and the stria vascularis.
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Sordera , Pérdida Auditiva , Humanos , Supervivencia Celular , Cóclea/patología , Células Ciliadas Auditivas/patología , Estría Vascular/patología , Sordera/patología , Pérdida Auditiva/patologíaRESUMEN
Animal studies have shown that the supporting-cells surviving in the organ of Corti after cochlear insult can be transdifferentiated into hair cells as a treatment for sensorineural hearing loss. Clinical trials of small-molecule therapeutics have been undertaken, but little is known about how to predict the pattern and degree of supporting-cell survival based on audiogram, hearing loss etiology or any other metric obtainable pre-mortem. To address this, we systematically assessed supporting-cell and hair cell survival, as a function of cochlear location in 274 temporal bone cases from the archives at the Massachusetts Eye and Ear and compared the histopathology with the audiograms and hearing-loss etiologies. Results showed that supporting-cell survival was always significantly greater in the apical half than the basal half of the cochlea, that inner pillars were more robust than outer pillars or Deiters' cells, and that total replacement of all supporting cells with a flat epithelium was rare outside of the extreme basal 20% of the cochlea. Supporting cell survival in the basal half of the cochlea was better correlated with the slope of the audiogram than with the mean high-frequency threshold per se: i.e. survival was better with flatter audiograms than with steeply down-sloping audiograms. Cochlear regions with extensive hair cell loss and exceptional supporting cell survival were most common in cases with hearing loss due to ototoxic drugs. Such cases also tended to have less pathology in other functionally critical structures, i.e. spiral ganglion neurons and the stria vascularis. Highlights: Supporting cell survival was systematically assessed in 274 human cochleasSupporting cell survival was better with flat than with down-sloping audiogramsSupporting cell survival was most robust when hearing loss was from ototoxic drugsOtotoxic cases also showed less pathology in other critical cochlear structuresThe data can inform clinical trials for regeneration via supporting cell conversion.
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HYPOTHESIS: There are clinically relevant differences in scalae anatomy and spiral ganglion neuron (SGN) quantity between incomplete partition type II (IP-II) and normal cochleae. BACKGROUND: IP-II is a commonly implanted cochlear malformation. Detailed knowledge of intracochlear three-dimensional (3D) morphology may assist with cochlear implant (CI) electrode selection/design and enable optimization of audiologic programming based on SGN maps. METHODS: IP-II (n = 11) human temporal bone histological specimens were identified from the National Institute on Deafness and Other Communication Disorders National Temporal Bone Registry and digitized. The cochlear duct, scalae, and surgically relevant anatomy were reconstructed in 3D. A machine learning algorithm was applied to map the location and number of SGNs. RESULTS: 3D scalae morphology of the basal turn was normal. Scala tympani (ST) remained isolated for 540 degrees before fusing with scala vestibuli. Mean ST volume reduced below 1 mm 2 after the first 340 degrees. Scala media was a distinct endolymphatic compartment throughout; mean ± standard deviation cochlear duct length was 28 ± 3 mm. SGNs were reduced compared with age-matched norms (mean, 48%; range, 5-90%). In some cases, SGNs failed to ascend Rosenthal's canal, remaining in an abnormal basalward modiolar location. Two forms of IP-II were seen: type A and type B. A majority (98-100%) of SGNs were located in the basal modiolus in type B IP-II, compared with 76 to 85% in type A. CONCLUSION: Hallmark features of IP-II cochleae include the following: 1) fusion of the ST and scala vestibuli at a mean of 540 degrees, 2) highly variable and overall reduced SGN quantity compared with normative controls, and 3) abnormal SGN distribution with cell bodies failing to ascend Rosenthal's canal.
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Cóclea , Implantes Cocleares , Humanos , Cóclea/diagnóstico por imagen , Rampa Timpánica , Escala Vestibular , Conducto CoclearRESUMEN
Hearing loss in patients with vestibular schwannoma (VS) is commonly attributed to mechanical compression of the auditory nerve, though recent studies suggest that this retrocochlear pathology may be augmented by cochlear damage. Although VS-associated loss of inner hair cells, outer hair cells, and spiral ganglion cells has been reported, it is unclear to what extent auditory-nerve peripheral axons are damaged in VS patients. Understanding the degree of damage VSs cause to auditory nerve fibers (ANFs) is important for accurately modeling clinical outcomes of cochlear implantation, which is a therapeutic option to rehabilitate hearing in VS-affected ears. A retrospective analysis of human temporal-bone histopathology was performed on archival specimens from the Massachusetts Eye and Ear collection. Seven patients met our inclusion criteria based on the presence of sporadic, unilateral, untreated VS. Tangential sections of five cochlear regions were stained with hematoxylin and eosin, and adjacent sections were stained to visualize myelinated ANFs and efferent fibers. Following confocal microscopy, peripheral axons of ANFs within the osseous spiral lamina were quantified manually, where feasible, and with a "pixel counting" method, applicable to all sections. ANF density was substantially reduced on the VS side compared to the unaffected contralateral side. In the upper basal turn, a significant difference between the VS side and unaffected contralateral side was found using both counting methods, corresponding to the region tuned to 2000 Hz. Even spiral ganglion cells (SGCs) contralateral to VS were affected by the tumor as the majority of contralateral SGC counts were below average for age. This observation provides histological insight into the clinical observation that unilateral vestibular schwannomas pose a long-term risk of progression of hearing loss in the contralateral ear as well. Our pixel counting method for ANF quantification in the osseous spiral lamina is applicable to other pathologies involving sensorineural hearing loss. Future research is needed to classify ANFs into morphological categories, accurately predict their electrical properties, and use this knowledge to inform optimal cochlear implant programming strategies.
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Sordera , Pérdida Auditiva , Neuroma Acústico , Humanos , Nervio Coclear/patología , Sordera/patología , Pérdida Auditiva/patología , Neuroma Acústico/patología , Estudios Retrospectivos , Ganglio Espiral de la Cóclea/patología , Lámina EspiralRESUMEN
HYPOTHESIS: Computed tomography (CT) density measurement can be used to objectively distinguish otosclerosis from normal bone and to determine histologic grades of otosclerosis. BACKGROUND: Otosclerosis can be seen on CT as subtle radiolucent areas. An objective radiologic measurement that corresponds to known otosclerosis pathology may improve diagnostic accuracy, and could be used as a radiologic biomarker for otosclerosis grade. METHODS: A blinded, randomized evaluation of both histologic grade on histopathology slides and CT density measurement was performed on 78 human temporal bone specimens (31 with otosclerosis and 47 controls) that had undergone high-resolution multi-detector CT before histologic processing. Assessments were performed at 11 regions of interest (ROIs) in the otic capsule for each specimen. RESULTS: The CT density measurement mean (Hounsfield Units) ± standard deviation for all ROIs (Nos. 1-9) was 2245 ± 854 for grade 0 (no otosclerosis, n = 711), 1896 ± 317 for grade 1 (inactive otosclerosis, n = 109), and 1632 ± 255 for grades 2 and 3 combined (mixed/active otosclerosis, n 35). There was a strong inverse correlation of CT density to histologic grade at ROIs Nos. 1-5 (ANOVA, p < 0.0001). The inter-rater reliability for CT density was very good (correlation coefficient 0.87, p < 0.05). ROC curves suggested a cut-off of 2,150HU to distinguish otosclerosis from normal bone, and 1,811HU to distinguish low grade from mixed/high grade otosclerosis. CONCLUSIONS: In human temporal bone specimens, CT density may be used to distinguish normal bone from bone involved by otosclerosis. A higher histologic grade (i.e., indicating a more active otosclerotic focus) correlated with lower density.
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Otosclerosis , Biomarcadores , Humanos , Otosclerosis/patología , Reproducibilidad de los Resultados , Hueso Temporal/diagnóstico por imagen , Hueso Temporal/patología , Tomografía Computarizada por Rayos X/métodosRESUMEN
HYPOTHESIS: The prevalence of monocyte-derived macrophages within cochlear vessels may increase following cochlear implantation. BACKGROUND: Recently, we reported an increase in the number of ionized calcium-binding adaptor molecule 1 (Iba1)-positive macrophages in selected cochlear sites such as the osseous spiral lamina and Rosenthal's canal following cochlear implantation. Activation of the immune system induces the recruitment of monocyte-derived macrophages. The prevalence of monocyte-derived macrophages within cochlear vessels may increase following cochlear implantation. However, the delivery system of macrophages to the human cochlea is incompletely understood. METHODS: The prevalence of macrophages and monocytes within cochlear blood vessels in 10 human subjects who had undergone unilateral cochlear implantation was studied by light microscopy using anti-Iba1 immunostaining. The densities of Iba1-positve monocytes per area of lumen of cochlear vessels in the sections near the round window in implanted ears were compared with the contralateral unimplanted ears. The correlation between the densities of Iba1-positve monocytes and the duration (months after the cochlear implantation) was also evaluated. RESULTS: The prevalence of Iba1-positive macrophages/monocytes in vessels near the round window in implanted ears (mean 26%, median 21%) was greater than in opposite unimplanted ears (mean 5.2%, median 2.5%: pâ<â0.01). The density of Iba1-positive monocytes in implanted ears (mean 32, median 16âcells/105âµm2) tended to be greater than that in unimplanted ears (mean 6.6, median 0.93âcells/105âµm2: pâ=â0.08). The density of Iba1-positive monocytes was significantly correlated with duration of implantation but not in the unimplanted ears. CONCLUSION: An increase in prevalence of Iba1-positive macrophages/monocytes within cochlear blood vessels after cochlear implantation was demonstrated. These findings suggest a delivery system of Iba1-positive macrophages through cochlear vessels in human that is ongoing for long duration.
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Implantación Coclear , Implantes Cocleares , Cóclea/cirugía , Humanos , Macrófagos , PrevalenciaRESUMEN
OBJECTIVES/HYPOTHESIS: Histopathological analysis of hair cell survival in human temporal bone sections has historically been binarized such that each hair cell row is rated as either present or absent, thereby greatly underestimating the amount of hair cell loss. Here, we describe and validate a technique to reliably assess fractional hair cell survival in archival sections stained with hematoxylin and eosin (H&E) using high-resolution light microscopy and optical sectioning. STUDY DESIGN: Technique validation. METHODS: Hair cell counts in archival temporal bone slide sets were performed by several observers using either differential interference contrast (DIC) or confocal microscopy of the endogenous eosin fluorescence in hair cells. As a further cross-check, additional decelloidinized sections were immunostained with hair cell markers myosin VI and VIIa. RESULTS: Cuticular plates and stereocilia bundles are routinely resolvable in DIC imaging of archival H&E-stained human material using standard research-grade microscopes, allowing highly accurate counts of fractional hair cell survival that are reproducible across observer and can be verified by confocal microscopy. CONCLUSIONS: Reanalysis of cases from the classic temporal bone literature on presbycusis suggests that, contrary to prior reports, differences in audiometric patterns may be well explained by the patterns of hair cell loss. LEVEL OF EVIDENCE: NA Laryngoscope, 130:487-495, 2020.
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Células Ciliadas Auditivas/patología , Presbiacusia/patología , Hueso Temporal/patología , Cadáver , Supervivencia Celular , Humanos , Microscopía Confocal , Cadenas Pesadas de Miosina , Miosina Tipo IIA no Muscular , Reproducibilidad de los Resultados , Coloración y EtiquetadoRESUMEN
HYPOTHESIS: Cochlear implantation may cause an increase in the number of macrophages in the human cochlea similar to previous findings in the vestibular endorgans. BACKGROUND: Macrophages play a key role in both an inflammatory response and homeostatic maintenance. Recently, an increase in the prevalence of macrophages was demonstrated in the human vestibular endorgans after implantation. However, the prevalence of macrophages in the cochlea after implantation is unclear. The aim of this study was to compare the distribution and prevalence of macrophages in implanted human cochleae and the contralateral unimplanted ears. METHODS: The prevalence of macrophages in the cochlea in 10 human subjects who had undergone unilateral cochlear implantation was studied by light microscopy using anti-Iba1 immunostaining. The densities of macrophages in the osseous spiral lamina (OSL) and Rosenthal's canal (RC) in implanted cochleae were compared with the contralateral unimplanted ears. The distribution of macrophage morphology (amoeboid, transitional, and ramified) was also compared. RESULTS: There were activated and phagocytosing macrophages within the fibrotic sheath surrounding the electrode track and within fibrous tissue with lymphocytic infiltration in implanted ears. The densities of macrophages in OSL and RC in implanted ears were significantly greater than in unimplanted ears in some areas. There was also a difference in the prevalence of macrophage phenotype between the OSL and RC. CONCLUSION: An increase in the density of macrophages in the cochlea after cochlear implantation was demonstrated. Both phagocytosis and anti-inflammatory activity of macrophages were suggested by the distribution and prevalence of macrophages in the implanted cochlea.
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Implantación Coclear , Implantes Cocleares , Cóclea/cirugía , Humanos , Macrófagos , PrevalenciaRESUMEN
The classic view of cochlear partition (CP) motion, generalized to be for all mammals, was derived from basal-turn measurements in laboratory animals. Recently, we reported motion of the human CP in the cochlear base that differs substantially from the classic view. We described a human soft tissue "bridge" (non-existent in the classic view) between the osseous spiral lamina (OSL) and basilar membrane (BM), and showed how OSL and bridge move in response to sound. Here, we detail relevant human anatomy to better understand the relationship between form and function. The bridge and BM have similar widths that increase linearly from base to apex, whereas the OSL width decreases from base to apex, leading to an approximately constant total CP width throughout the cochlea. The bony three-dimensional OSL microstructure, reconstructed from unconventionally thin, 2-µm histological sections, revealed thin, radially wide OSL plates with pores that vary in size, extent, and distribution with cochlear location. Polarized light microscopy revealed collagen fibers in the BM that spread out medially through the bridge to connect to the OSL. The long width and porosity of the OSL may explain its considerable bending flexibility. The similarity of BM and bridge widths along the cochlea, both containing continuous collagen fibers, may make them a functional unit and allow maximum CP motion near the bridge-BM boundary, as recently described. These anatomical findings may help us better understand the motion of the structures surrounding the organ of Corti and how they shape the input to the cochlear sensory mechanism.
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Lámina Espiral/anatomía & histología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Membrana Basilar/anatomía & histología , Niño , Cóclea/fisiología , Femenino , Colágenos Fibrilares , Humanos , Imagenología Tridimensional , Masculino , Microscopía , Persona de Mediana Edad , Lámina Espiral/diagnóstico por imagen , Adulto JovenRESUMEN
The localization of proteins by immunostaining is a powerful method to investigate otologic disorders. However, the use of fixatives and embedding media (necessary for the preservation of morphology) can obscure antigens, making it difficult to perform immunoassays. We performed a systematic investigation of the effects of fixative and embedding medium on morphology and immunostaining of the mouse cochlea. Three different fixative solutions [4% formaldehyde (F), 4% formaldehyde + 1% acetic acid (FA), and 4% formaldehyde + 1% acetic acid + 0.1% glutaraldehyde (FGA)] and 3 different embedding media (paraffin, polyester wax, and celloidin) were used. Morphology was assessed using light microscopy. Immunostaining was studied using a panel of 6 antibodies (to prostaglandin D synthase, aquaporin 1, connective tissue growth factor, 200-kDa neurofilament, tubulin and Na(+),K(+)-ATPase). Preservation of morphology was suboptimal with paraffin, adequate with polyester wax and superb with celloidin. Immunostaining was successful using all 6 antibodies in all 3 fixatives and all 3 embedding media. While there were differences in strength of signal and localization of antigen between the 3 fixatives, overall, FA and FGA gave the most uniform results. For a given fixative and antibody, there was surprisingly little difference in the quality of immunostaining between celloidin and paraffin, while results in polyester wax were not as good in some cases. These results suggest that celloidin may be the embedding medium of choice for both morphological and pathological studies, including immunostaining when morphology must be optimized.
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Cóclea/citología , Fijadores , Formaldehído , Inmunohistoquímica/métodos , Adhesión en Parafina , Animales , Cóclea/metabolismo , Colodión , Ratones , Ratones Endogámicos CBA , Poliésteres , Proteínas/metabolismo , Hueso Temporal/citología , Hueso Temporal/metabolismo , CerasRESUMEN
OBJECTIVES: We sought to determine whether the technique of celloidin removal influences the results of immunostaining in celloidin-embedded cochleae. METHODS: We compared four protocols of celloidin removal, including those using clove oil, acetone, ether-alcohol, and methanol saturated with sodium hydroxide. By optimally fixing our tissue (perfused mice), and keeping constant the fixative type (formalin plus acetic acid), fixation time (25 hours), and decalcification time (ethylenediaminetetraacetic acid for 7 days), we determined whether the technique of celloidin removal influenced the immunostaining results. Six antibodies were used with each removal method: prostaglandin D synthase, sodium, potassium adenosine triphosphatase (Na+,K(+)-ATPase), aquaporin 1, connective tissue growth factor, tubulin, and 200 kd neurofilament. RESULTS: Clove oil, acetone, and ether-alcohol resulted in incomplete removal of the celloidin, thereby negatively affecting the results of immunostaining. The methanol-sodium hydroxide method was effective in completely removing the celloidin; it produced the cleanest and most reproducible immunostaining for all six antibodies. CONCLUSIONS: Freshly prepared methanol saturated with sodium hydroxide and diluted 1:2 with methanol was the best solvent for removing celloidin from mouse temporal bone sections, resulting in consistent and reproducible immunostaining with the six antibodies tested.