Cochlin-Tomoprotein: A Novel Perilymph-Specific Protein and a Potential Marker for the Diagnosis of Perilymphatic Fistula

Ikezono, Tetsuo; Shindo, Susumu; Sekiguchi, Satomi; Hanprasertpong, Charuk; Li, Lishu; Pawankar, Ruby; Morizane, Toshio; Baba, Shunkichi; Koizumi, Yasuo; Sekine, Kuwon; Watanabe, Atsushi; Komatsuzaki, Atsushi; Murakami, Shingo; Kobayashi, Toshimitsu; Miura, Masakazu; Yagi, Toshiaki

doi:10.1159/000212113

Cited by 40 publications

(34 citation statements)

References 59 publications

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“…In rats, the COCH gene is expressed preferentially in the inner ear, and the expression of full-length cochlin p63 is specific to the inner ear . We also found CTP to be expressed only in the perilymph, not in the serum or cerebrospinal fluid in humans [Ikezono et al, 2009]. These results suggest that mechanisms exist to ensure these tissue-specific expression patterns.…”

Section: Introductionmentioning

confidence: 52%

Expression of Cochlin mRNA Splice Variants in the Inner Ear

Sekine¹,

Ikezono²,

Matsumura³

et al. 2009

Audiol Neurotol

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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.

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Section: Introductionmentioning

confidence: 52%

Expression of Cochlin mRNA Splice Variants in the Inner Ear

Sekine¹,

Ikezono²,

Matsumura³

et al. 2009

Audiol Neurotol

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“…We previously analyzed the expression of CTP in various human bodily fluids, including the serum, CSF, saliva and perilymph [Ikezono et al, 2009]. All bodily fluid samples, except the perilymph, were negative for CTP.…”

Section: Discussionmentioning

confidence: 99%

“…We detected 3 cochlin isoforms, p63s, p44s and p40s, in the inner ear tissue and a short 16-kDa isoform named Cochlin-tomoprotein (CTP) in the perilymph [Ikezono et al, 2001[Ikezono et al, , 2004. Since cochlin was found to be highly specific to the inner ear [Robertson et al, 1994;Abe et al, 2003;Li et al, 2005], we tested the expression specificity of CTP in perilymph; CTP was indeed selectively expressed only in the perilymph, and not in CSF, saliva or serum [Ikezono et al, 2009]. In addition, we reported the molecular mechanisms that regulate the perilymph-specific expression of CTP [Sekine et al, 2009].…”

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confidence: 99%

The Performance of Cochlin-Tomoprotein Detection Test in the Diagnosis of Perilymphatic Fistula

et al. 2009

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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°C) or 4°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 µl/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.

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“…Its characteristics have already been reported in detail [1][2][3]8]. We initially developed a Western blotting-based CTP detection test for PLF diagnosis, and improved the throughput and sensitivity of this test by using an ELISA-based CTP detection kit.…”

Section: Fistula Signmentioning

confidence: 99%

“…Based on proteomic analysis, we have identified an isoform of Cochlin, Cochlin tomo-protein (CTP), [1] as a perilymph-specific protein that is not expressed in the blood, CSF or saliva [2]. The detection of CTP in the middle ear is indicative of the presence of a PLF and perilymph leakage.…”

Section: Introductionmentioning

confidence: 99%

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

Matsuda

Sakamoto

Matsumura

et al. 2017

Acta Oto-Laryngologica

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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.ARTICLE HISTORY

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Cochlin-Tomoprotein: A Novel Perilymph-Specific Protein and a Potential Marker for the Diagnosis of Perilymphatic Fistula

Cited by 40 publications

References 59 publications

Expression of Cochlin mRNA Splice Variants in the Inner Ear

Expression of Cochlin mRNA Splice Variants in the Inner Ear

The Performance of Cochlin-Tomoprotein Detection Test in the Diagnosis of Perilymphatic Fistula

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

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