Estimated incidence and characteristics of vestibular neuritis in Japan: A nationwide survey.

OBJECTIVE: To assess the annual incidence of vestibular neuritis (VN) in the Japanese population. METHODS: We conducted a mail-based survey targeting otolaryngologic clinics and hospitals across Japan to estimate the annual number of patients who were newly-diagnosed with VN during the one-year period of 2021. Using a stratified sampling method, we selected 1,107 departments and asked them to report the number of new patients with VN and their demographics. The total number of VN patients was estimated by multiplying the reported numbers by the reciprocal of the sampling rate and response rate. RESULTS: The overall survey response rate was 40.5 % (448 departments). The estimated number of newly-diagnosed VN patients in 2021 was 8,861 (95 % confidential interval [CI], 2,290-15,432) The annual incidence of VN was 7.05 per 100,000 population in Japan. The male-to-female ratio of VN patients was 0.96, and the mean age was 60.3 ± 16.1 years (range 11-94 years). CONCLUSIONS: The annual incidence of VN in Japan in 2021 had almost doubled and the mean age had become older compared to the previous study in 1993 (annual incidence; 3.5 per 100,000 per year; mean age: 45 years).

Estimated prevalence and characteristics of bilateral vestibulopathy diagnosed in Japan: A nationwide survey.

OBJECTIVE: To assess the prevalence and annual incidence of bilateral vestibulopathy (BV) diagnosed in the Japanese adult population. METHODS: We conducted a mail-based survey targeting otolaryngologic clinics and hospitals across Japan to estimate the annual number of patients who were diagnosed as having bilateral vestibulopathy after vestibular function tests during a 12-month period ending March 2019. Using a stratified sampling method, we selected 1,106 departments and asked them to report the number of patients with BV and their demographics. The total number of patients was estimated by multiplying the reported numbers by the reciprocal of the sampling rate and response rate. RESULTS: The overall survey response rate was 51.4% (568 departments). The estimated number of patients diagnoses with BV in 2018 was 1,063 (95% confidential interval [CI], 127-1,998) which included 407 patients (95% CI: 134-680) newly-diagnosed with BV. The prevalence and annual incidence of BV in Japan were 0.84 and 0.32, respectively per 100,000 population in Japan. The male-to-female ratio of BV patients was 1.29, and the mean age was 63.7 ± 16.4 years (range 18-84 years). The most frequent etiologies of BV were Meniere's disease (11.4%), meningitis (3.4%), and ototoxic agents (3%). CONCLUSIONS: Patients who were diagnosed as having BV were extremely rare in Japan.

Congenital Membranous Stapes Footplate Producing Episodic Pressure-Induced Perilymphatic Fistula Symptoms.

Introduction: Recent third window syndrome studies have revealed that the intact bony labyrinth and differences in the stiffness of the oval and round windows are essential for proper cochlear and vestibular function. Herein we report a patient with a congenital dehiscence of the right stapes footplate. This dehiscence caused long-standing episodic pressure-induced vertigo (Hennebert sign). At the time of presentation, her increased thoracic pressure changes induced the rupture of the membranous stapes footplate. Perilymph leakage was confirmed by imaging and a biochemical test [perilymph-specific protein Cochlin-tomoprotein (CTP) detection test]. Case Report: A 32-year-old woman presented with a sudden onset of right-sided hearing loss and severe true rotational vertigo, which occurred immediately after nose-blowing. CT scan showed a vestibule pneumolabyrinth. Perilymphatic fistula (PLF) repair surgery was performed. During the operation, a bony defect of 0.5 mm at the center of the right stapes footplate, which was covered by a membranous tissue, and a tear was found in this anomalous membrane. A perilymph-specific protein CTP detection test was positive. The fistula in the footplate was sealed. Postoperatively, the vestibular symptoms resolved, and her hearing improved. A more detailed history revealed that, for 15 years, she experienced true rotational vertigo when she would blow her nose. After she stopped blowing her nose, she would again feel normal. Discussion: There is a spectrum of anomalies that can occur in the middle ear, including the ossicles. The present case had a dehiscence of the stapes, with a small membranous layer of tissue covering a bony defect in the center of the footplate. Before her acute presentation to the hospital, this abnormal footplate with dehiscence induced pathological pressure-evoked fluid-mechanical waves in the inner ear, which resulted in Hennebert sign. When patients have susceptibility (e.g., weak structure) to rupture, such as that identified in this case, PLF can be caused by seemingly insignificant events such as nose-blowing, coughing, or straining. Conclusion: This case demonstrates that PLF is a real clinical entity. Appropriate recognition and treatment of PLF can improve a patient's condition and, hence, the quality of life.

The diagnostic performance of a novel ELISA for human CTP (Cochlin-tomoprotein) to detect perilymph leakage.

Perilymphatic fistula is defined as an abnormal communication between the perilymph-filled space and the middle ear, or cranial spaces. The manifestations include a broad spectrum of neuro-otological symptoms such as hearing loss, vertigo/dizziness, disequilibrium, aural fullness, tinnitus, and cognitive dysfunction. By sealing the fistula, perilymphatic fistula is a surgically correctable disease. Also, appropriate recognition and treatment of perilymphatic fistula can improve a patient's condition and hence the quality of life. However, the difficulty in making a definitive diagnosis due to the lack of an appropriate biomarker to detect perilymph leakage has caused a long-standing debate regarding its management. We have reported a clinical test for the diagnosis of perilymphatic fistula by detecting a perilymph specific protein, Cochlin-tomoprotein, as a diagnostic marker using a western blot. The aim of this study is to establish an ELISA-based human Cochlin-tomoprotein detection test and to evaluate its diagnostic accuracy in clinical subjects. The results of ELISA showed good dilution reproducibility. The mean concentration was 49.7±9.4 of 10 perilymph samples. The ROC curve in differentiating the perilymph leakage condition from the normal middle ear was significant (P < 0.001) with an area under the curve (AUC) of 0.918 (95% CI 0.824-0.100). We defined the diagnostic criteria as follows: CTP<0.4 negative; 0.4≦CTP<0.8 intermediate; 0.8≦CTP(ng/ml) positive in the clinical usage of the hCTP ELISA, and sensitivity and specificity were 86.4% and 100%, respectively. We further tested the expression specificity of the Cochlin-tomoprotein by testing blood and CSF samples. The concentration was below the detection limit (0.2 ng/ml) in 38 of the 40 blood, and 14 of the 19 CSF samples. We report the accuracy of this test for the diagnosis of perilymphatic fistula. Using ELISA, we can improve the throughput of the test. Furthermore, it is useful for a large-scale study to characterize the clinical picture and delineate the management of this medical condition.

Expression Profiling of MicroRNAs in the Inner Ear of Elderly People by Real-Time PCR Quantification.

The molecular mechanisms underlying age-related hearing loss are unknown, and currently, there is no treatment for this condition. Recent studies have shown that microRNAs (miRNAs) and age-related diseases are intimately linked, suggesting that some miRNAs may present attractive therapeutic targets. In this study, we obtained 8 human temporal bones from 8 elderly subjects at brain autopsy in order to investigate the expression profile of miRNAs in the inner ear with miRNA arrays. A mean of 478 different miRNAs were expressed in the samples, of which 348 were commonly expressed in all 8 samples. Of these, levels of 16 miRNAs significantly differed between young elderly and old elderly subjects. miRNAs, which play important roles in inner ear development, were detected in all samples, i.e., in both young and old elderly subjects, whether with or without hearing loss. Our results suggest that these miRNAs play important roles not only in development, but also in the maintenance of inner ear homeostasis.

A nationwide multicenter study of the Cochlin tomo-protein detection test: clinical characteristics of perilymphatic fistula cases.

OBJECTIVE: To investigate the positive rate for the Cochlin tomo-protein (CTP: an inner ear-specific protein) detection test among patients with inner ear-related clinical manifestations and evaluate the clinical characteristics of definite perilymphatic fistula (PLF). METHODS: We have performed an ELISA-based CTP detection test using middle ear lavage (MEL) samples from 497 cases of suspected PLF enrolled from 70 clinical centers nationwide between 2014 and 2015. In addition to the CTP-positive rate, audio-vestibular symptoms were compared between CTP-positive and -negative cases. RESULTS: 8-50% of patients in category 1 (trauma, middle and inner ear disease cases), and about 20% of those in categories 2, 3 and 4 (external origin antecedent events, internal origin antecedent events, and without antecedent event, respectively) were positive for CTP. In category 1 cases, the earlier tested samples showed a higher CTP-positive rate, whereas no differences were observed in categories 2, 3 or 4. The characteristic clinical features in the earlier tested cases were nystagmus and fistula sign in CTP test-positive cases in category 1, and streaming water-like tinnitus in those in categories 2, 3 and 4. CONCLUSION: The present study clarified that CTP detection test-positive patients exist at considerable rates among patients with inner ear-related manifestations.

Unidirectional abnormal eye movement without gaze nystagmus - Report of a case.

We report here a case with unidirectional abnormalities of smooth eye movements without gaze nystagmus. Abnormalities of eye movements were confined to unidirectional (leftward) horizontal pursuit and slow phase of OKN; however, horizontal VOR (slow phase of caloric nystagmus) and saccade were normal, and vertical eye movements were also normal. No lesions were detected in the central nervous system, and any history of drug intake was denied. Although the cause of the unidirectional abnormality in eye movement of this case is still not clear, a congenital origin seems to be the most probable.

Cochlin expression in the rat perilymph during postnatal development.

CONCLUSIONS: The changes in the cochlin isoforms in the perilymph may provide important insights to the understanding of cochlin function and the pathogenesis of related inner ear diseases. OBJECTIVES: Cochlin is involved in various pathologies of the inner ear. Altered levels of cochlin isoforms in developing inner ear tissue were reported previously. The purpose of this study was to elucidate the cochlin isoform expression in the perilymph of rats during postnatal development in relation to Coch gene mRNA expression. METHODS: We studied the cochlin isoforms in the rat perilymph during postnatal development by Western blot analysis. Real-time PCR was also performed to elucidate the expression level of Coch mRNA in the developing inner ear of rats. RESULTS: Western blot analysis showed that the expression of p63s in the perilymph was highest on the 12th day after birth (DAB12), the earliest age at which we could identify the perilymphatic space microscopically, and it decreased gradually as the cochlea developed. On the other hand, the expression of Cochlin-tomoprotein (CTP)was lowest on DAB12 and increased gradually up to DAB24. COCH mRNA was detected from DAB3 and gradually increased to DAB15, and then gradually decreased to DAB70.

Cochlin-tomoprotein (CTP) detection test identifies traumatic perilymphatic fistula due to penetrating middle ear injury.

CONCLUSIONS: The cochlin-tomoprotein (CTP) detection test can be used to make a definite, objective diagnosis of traumatic perilymphatic fistula (PLF), and therefore offers valuable information on patient selection for surgical treatment. OBJECTIVES: Penetrating middle ear injury can cause traumatic PLF, which is a surgically treatable otologic emergency. Recently, we have reported on CTP, a novel perilymph-specific protein. The purpose of this study was to determine if the CTP detection test is useful for the diagnosis of traumatic PLF. METHODS: This was a prospective study of CTP detection in penetrating middle ear injury cases with tympanic membrane perforation and hearing loss. RESULTS: A total of seven individuals were included in this study. CTP was detected in three of four cases with posterosuperior quadrant perforation of the tympanic membrane. In one of these three cases, even though the high resolution CT scan was not suggestive of PLF and the perilymph leakage could not be visualized intraoperatively, the CTP detection test was able to detect PLF. In two cases, the preoperative positive test results enabled us to make a diagnosis of PLF and a decision for surgical treatment. CTP was not detected in the cases with anterior or inferior tympanic membrane perforation.

Molecular cloning of the Coch gene of guinea pig inner ear and its expression analysis in cultured fibrocytes of the spiral ligament.

CONCLUSIONS: We have cloned guinea pig Coch cDNA and the sequence information will be useful for future molecular study combined with physiological experiments. Proper Coch gene expression appears to be dependent on the unique extracellular micro-environment of the inner ear in vivo. These results provide insight into the Coch gene expression and its regulation. OBJECTIVE: To characterize the guinea pig Coch gene, we performed molecular cloning and expression analysis in the inner ear and cultured fibrocytes of the spiral ligament. METHODS: The Coch cDNA was isolated using RACE. Cochlin isofoms were studied by Western blot using three different types of mammalian inner ear. The cochlear fibrocytes were cultured and characterized by immunostaining. Coch gene expression in the fibrocytes was investigated and the influence of cytokine stimulation was evaluated. RESULTS: The full-length 1991 bp Coch cDNA that encodes a 553 amino acid protein was isolated. The sequence had significant homology with other mammals, and the sizes of the Cochlin isoforms were identical. In the cultured fibrocytes, Coch mRNA was expressed in a very small amount and the isoform production was different, compared with the results in vivo. Cytokine stimulation did not alter the level of mRNA expression or isoform formation.

CTP (Cochlin-tomoprotein) detection in the profuse fluid leakage (gusher) from cochleostomy.

CONCLUSIONS: By testing 125 samples, we confirmed that Cochlin-tomoprotein (CTP) is present in the perilymph, not in cerebrospinal fluid (CSF). Perilymph and CSF exist in two distinct compartments, even in the case of a malformed inner ear with a bony defect in the lamina cribrosa, as described here. Cochleostomy might have suddenly decreased the perilymph pressure, allowing the influx of CSF into the inner ear resulting in profuse fluid leakage, first perilymph then CSF. OBJECTIVES: The first purpose of this study was to further confirm the specificity of the perilymph-specific protein CTP that we reported recently. Secondly, we assessed the nature of the fluid leakage from the cochleostomy using the CTP detection test. METHODS: A standardized CTP detection test was performed on 65 perilymph and 60 CSF samples. Samples of profuse fluid leakage collected from cochleostomy during cochlear implantation surgery of one patient with branchio-oto-renal (BOR) syndrome were also tested by the CTP detection test. RESULTS: CTP was detected in 60 of 65 perilymph samples but not in any of the CSF samples. The leaked fluid was shown to contain CTP, i.e. perilymph, at the outset, and then the CTP detection signals gradually disappeared as time elapsed.

The performance of Cochlin-tomoprotein detection test in the diagnosis of perilymphatic fistula.

BACKGROUND: Perilymphatic fistula (PLF), defined as an abnormal communication between the inner and middle ear, presents with a symptomatology of hearing loss and vestibular disorder that is indistinguishable from a number of other inner ear diseases. Methods of diagnosis remain controversial. We have previously shown that Cochlin-tomoprotein (CTP) is selectively detected in the perilymph. To establish a definite diagnostic test for PLF using CTP as a biochemical marker, we examined the diagnostic performance of the CTP detection test. METHODS: CTP detection test was performed by Western blot using recombinant human CTP (rhCTP) as a spiked standard. We evaluated the specificity of the CTP detection test by testing non-PLF cases. To describe the limitations of the test, we tested samples from patients with middle ear infection. We also studied the stability of CTP protein by storing the samples at room temperature (25 degrees C) or 4 degrees C for 55 days. The effects of repeated freezing and thawing were also evaluated. Serially diluted perilymph was tested to find out the detection limit of CTP. FINDINGS: We have established a standardized CTP detection test using high (0.27 ng) and low (0.13 ng) spiked standards of rhCTP in Western blotting. Middle ear lavages (MEL) from 54 of 55 non-PLF cases were negative in the CTP detection test, i.e. the specificity of the test is 98.2%. MEL from 43 out of 46 cases with chronic suppurative otitis media or middle ear cholesteatoma were negative for CTP. CTP is a stable protein and detection was not affected by the storage, or freezing and thawing. The detection limit of perilymph was 0.161 microl/lane in an average of 5 samples. INTERPRETATION: CTP is a stable perilymph-specific protein, and this CTP detection could be the first clinically established diagnostic tool to detect PLF with a high specificity. PLF is surgically correctable by sealing the fistula. Appropriate recognition and treatment of PLF can improve hearing and balance in afflicted patients.

Ultrastructural localization of cochlin in the rat cochlear duct.

Cochlin, a product of the COCH gene, is a major constituent of the inner ear extracellular matrix. Type II collagen, a protein that contributes to structural stability, is also a component of this extracellular matrix. In this study, using the postembedding immunogold method, we demonstrate the localization of cochlin and type II collagen in the cochlear duct at the ultrastructural level. The immunolabeling of cochlin was observed in the fibrillar substance in the spiral limbus, beneath the inner sulcus cells, and in the basilar membrane, the spiral prominence and the spiral ligament. Immunolabeling of type II collagen was observed in the same fibrillar substance in the extracellular matrix of the cochlear duct. This localization of cochlin is consistent with the expected localization of type II collagen. The localization of cochlin and type II collagen indicates the important roles played by these proteins in the hearing process.

Expression of cochlin mRNA splice variants in the inner ear.

Proteomic analysis of inner ear proteins revealed unique properties of cochlin, encoded by the COCH gene. We detected 3 cochlin isoforms, p63s, p44s and p40s, in the inner ear tissue and a short 16-kDa isoform, cochlin-tomoprotein (CTP), in the perilymph. The role of the cochlin isoforms has not been elucidated. To improve our understanding of the mechanism of cochlin isoform expression, we investigated rat cochlin mRNA expression in the inner ear and other organs. We performed RNA-ligation-mediated amplification of cDNA ends (RLM-RACE) using RNA isolated from the inner ear and spleen of rats, which are known to express abundant cochlin mRNA. We also examined the expression profile of full-length cochlin mRNA by nested RT-PCR in the cerebrum, cerebellum/brain stem, eye, inner ear, thyroid gland, thymus gland, lung, heart, liver, spleen, adrenal gland, kidney and blood. We verified CTP expression in rat perilymph by Western blot. By RLM-RACE, alternately spliced variants of cochlin mRNA with 3 different lengths were detected (2442, 2008 and 724 bp). The two longer mRNAs encode full-length cochlin with different polyadenylation signals in the 3'-untranslated region, which are expressed both in the ear and spleen. The short variant encodes the limulus factor C, cochlin, late gestation lung protein (LCCL) domain and the N-terminal sequence of the von Willebrand factor A (vWFA1) domain, and this variant was detected only in the ear. All 3 variants have the same transcriptional start site. By RT-PCR, we found that full-length cochlin was expressed in all organs examined, with a splice variant in the heart. By Western blot, we detected short isoforms (11-17 kDa) in the perilymph. Cochlin isoform formation is regulated, at least in part, by alternative splicing at the transcriptional level. The short mRNA was detected only in the inner ear, and this variant may provide a clue to understanding the formation and function of cochlin isoforms.

Cochlin-tomoprotein: a novel perilymph-specific protein and a potential marker for the diagnosis of perilymphatic fistula.

BACKGROUND: Perilymphatic fistula (PLF) is an abnormal connection between the inner and middle ear. A procedure for obtaining definite proof of a PLF remains elusive, and methods of diagnosis remain controversial. To date, there is no clinically relevant biochemical marker for perilymph leakage. Using proteomic analysis of inner ear proteins, we have previously found unique properties of cochlin, encoded by the COCH gene. We detected 3 cochlin isoforms (p63s, p44s and p40s) in the inner ear tissue and a short 16-kDa isoform of cochlin-tomoprotein (CTP) in the perilymph. Since cochlin was found to be highly specific to the inner ear, we speculated that CTP might also be specific to the perilymph. The aim of this study was to determine whether CTP, a novel perilymph-specific protein, could be used as a marker for the diagnosis of PLF. METHODS: By Western blotting, we investigated the specificity of CTP expression in a range of body fluids that included perilymph, serum, saliva and cerebrospinal fluid. To elucidate the detection limit of CTP, serially diluted recombinant human (rh)CTP as well as human perilymph was tested. RESULTS: CTP was selectively expressed in all 20 perilymph samples tested, but not in 77 samples of the other body fluids. The detection limit of rhCTP was 0.27 ng or 0.022 microl of perilymph per well on Western blot analysis. CONCLUSION: The results strongly suggest that CTP can be a specific marker of perilymph leakage. Moreover, CTP has the potential to be a biochemical marker that allows a definitive diagnosis of the etiology of PLF-related hearing loss and vestibular disorders.

Spatiotemporal expression of cochlin in the inner ear of rats during postnatal development.

Cochlin (encoded by COCH) constitutes 70% of non-collagenous protein in the inner ear, and the expression of cochlin is highly specific to the inner ear. Eleven missense mutation and one in-frame deletion have been reported in the COCH gene, causing hereditary progressive sensorineural hearing loss and vestibular dysfunction, DFNA9. These data imply that cochlin should bear an essential and crucial role in the inner ear function. However, the role of cochlin has not been fully clarified. We have investigated the spatiotemporal expression of cochlin in the inner ear of rats during postnatal development to better understand the functional role of cochlin. By immunohistochemistry, cochlin expression was faint in the cochlea and vestibule on the 6th day after birth (DAB6). At DAB70, strong expression of cochlin was detected in the spiral limbus and spiral ligament within the cochlea, and in the stromata of the maculae of otolithic organs and crista ampullaris within the vestibule. Immunoreactivity for cochlin increased during the postnatal development. Western blot analysis also showed an increase in the expression of cochlin isoforms. Furthermore, the dominant isoform of cochlin expressed changed from p63s to p40s between DAB24 and DAB70. These results suggest that the expression of cochlin may be related to the maturation of inner ear function, and the change in isoforms of cochlin expressed will provide important insight into the understanding of both cochlin function and formation of cochlin isoforms. This is the first to report about the spatiotemporal expression of cochlin in the developing rat inner ear.

Ultrastructural co-localization of cochlin and type II collagen in the rat semicircular canal.

Cochlin and type II collagen are major constituents of the inner ear extracellular matrix. To investigate the morphological relation of cochlin and type II collagen in the rat semicircular canal, immuno-electronmicroscopic analysis was performed using the post-embedding immunogold method. Immunolabeling for cochlin was detected in the fibrillar substance underlying the supporting epithelium of the sensory cells and beneath the epithelial cells facing the endolymph in the semicircular canals. Immunolabeling for type II collagen was observed in the same fibrillar substance in the subepithelial area. The co-localization of cochlin and type II collagen in the fibrillar substance in the subepithelial area indicate that cochlin may play a role in the structural homeostasis of the vestibule acting in concert with the fibrillar type II collagen bundles.

Expression of cochlin in the vestibular organ of rats.

The COCH gene mutated in autosomal dominant sensorineural deafness (DFNA9) encodes cochlin, a major constituent of the inner ear extracellular matrix. Cochlin constitutes 70% of the inner ear protein and cochlin isoforms can be classified into three subgroups, p63s, p44s and p40s. Symptoms of some DFNA9 patients are consistent with those of Ménière's disease. Here, we report the expression of cochlin in the vestibular organ of rats using isoform-specific antibodies that recognize all three isoforms. Cochlin is highly expressed in the stromata of the maculae of otolithic organs and cristae of semicircular canals, and in the channels in the bony labyrinth that transmit the dendritic innervation to the cristae and maculae. Cochlin cannot be detected in the sensory cells, dark cells, nor in the acellular structures, otolithic membrane or in the cupula. These findings support the theory that deposition of acidophilic substance in the inner ear caused by mutation of cochlin can induce a secondary retrograde dendritic degeneration of the vestibular nerves.

Expression of full-length Cochlin p63s is inner ear specific.

OBJECTIVE: The COCH gene mutated in DFNA9, murine an autosomal dominant hereditary hearing impairment, encodes Cochlin. Cochlin is also suggested to be the self-antigen of autoimmune sensorineural hearing loss. We previously reported that Cochlin constitutes 70% of the inner ear proteins and is classified into three types of isoform, p63s, p44s, and p40s. To study the specificity of expression of Cochlin isoforms in various organs, here we have investigated expression of the COCH gene at both the transcriptional and translational level. METHODS: COCH gene expression was studied by RT-PCR and Southern blot analysis. Cochlin isoforms were studied by Western blot analysis using an isoform specific antibody. RESULTS: At the transcriptional level, COCH mRNA was detected only in the inner ear by RT-PCR. Southern blot analysis of RT-PCR products detected a high level of COCH mRNA in the inner ear, lower level in spleen, and very low levels in the cerebrum, cerebellum/brain stem, eye, liver and kidney. At the translational level, Western blot analysis showed that a set of isoform, p63s, p44s, and p40s was detected at high levels only in the inner ear. In contrast, multiple proteins were detected at much lower levels in other organs tested. Notably, full-length Cochlin p63s was detected only in the inner ear. CONCLUSION: Our findings demonstrate that the COCH gene is expressed preferentially in the inner ear and that expression of full-length Cochlin p63s is specific to the inner ear. These results will be central to understanding the function of Cochlin and its role in the pathophysiology of DFNA9.

Identification of a novel Cochlin isoform in the perilymph: insights to Cochlin function and the pathogenesis of DFNA9.

The COCH gene mutated in DFNA9, an autosomal dominant hereditary sensorineural hearing loss and vestibular disorder, encodes Cochlin. Previously, we reported three bovine Cochlin isoforms, p63s, p44s, and p40s, which exhibit significant molecular heterogeneity in vivo. Here we have characterized Cochlin isoforms by generating four isoform-specific anti-Cochlin antibodies. The same three Cochlin isoforms, p63s, p44s, and p40s, were detected in human and cow inner ear tissue; however, p44s and p40s were not detected in perilymph. We identified a novel short 16kDa isoform in human perilymph and a 18-23kDa isoform in cow perilymph, named Cochlin-tomoprotein (CTP), corresponding to the N-terminus of full-length Cochlin (p63s) and the LCCL domain. Notably, CTP contains all of the known mutation sites associated with DFNA9. The pathogenesis of DFNA9 is not fully clarified as yet, and this novel perilymph-associated CTP isoform might provide mechanistic clues to how mutations in the COCH gene damage the inner ear function.