Kyu-Hee Han

Diagnostic Application of Targeted Resequencing for Familial Nonsyndromic Hearing Loss

Identification of causative genes for hereditary nonsyndromic hearing loss (NSHL) is important to decide treatment modalities and to counsel the patients. Due to the genetic heterogeneity in sensorineural genetic disorders, the high-throughput method can be adapted for the efficient diagnosis. To this end, we designed a new diagnostic pipeline to screen all the reported candidate genes for NSHL. For validation of the diagnostic pipeline, we focused upon familial NSHL cases that are most likely to be genetic, rather than to be infectious or environmental. Among the 32 familial NSHL cases, we were able to make a molecular genetic diagnosis from 12 probands (37.5%) in the first stage by their clinical features, characteristic inheritance pattern and further candidate gene sequencing of GJB2, SLC26A4, POU3F4 or mitochondrial DNA. Next we applied targeted resequencing on 80 NSHL genes in the remaining 20 probands. Each proband carried 4.8 variants that were not synonymous and had the occurring frequency of less than three among the 20 probands. These variants were then filtered out with the inheritance pattern of the family, allele frequency in normal hearing 80 control subjects, clinical features. Finally NSHL-causing candidate mutations were identified in 13(65%) of the 20 probands of multiplex families, bringing the total solve rate (or detection rate) in our familial cases to be 78.1% (25/32) Damaging mutations discovered by the targeted resequencing were distributed in nine genes such as WFS1, COCH, EYA4, MYO6, GJB3, COL11A2, OTOF, STRC and MYO3A, most of which were private. Despite the advent of whole genome and whole exome sequencing, we propose targeted resequencing and filtering strategy as a screening and diagnostic tool at least for familial NSHL to find mutations based upon its efficacy and cost-effectiveness.

Prevalence of p.V37I Variant of GJB2 in Mild or Moderate Hearing Loss in a Pediatric Population and the Interpretation of Its Pathogenicity

A p.V37I variant of GJB2 has been reported from subjects with moderate or slight hearing loss especially in East Asian populations. This study aimed to estimate the prevalence of the p.V37I variant among such subjects and prove, epidemiologically, its pathogenic potential to cause mild hearing loss. A total of 380 subjects from 201 families with hearing loss were enrolled. From them, 103 families were selected who had autosomal recessive inheritance or sporadic occurrence of hearing loss and who were younger than 15 years old. GJB2 sequencing was carried out for the probands of all 103 families. The prevalence of the p.V37I variant was compared between the subtle, mild or moderate hearing loss (group I) and the severe or profound hearing loss (group II) groups. Where possible, a targeted next generation sequencing of 82 deafness genes was performed from the p.V37I carrier to exclude the existence of other pathogenic genes. Five (4.8%) of 103 probands were found to carry p.V37I. The carrier frequency of p.V37I among group I (18.2%) was significantly higher than that of group II (1.2%) or the reported Korean normal hearing control group (1.0%). Detection of the p.V37I variant of GJB2 in 18.2% of Koreans with mild hearing loss strongly suggests its contribution to the pathogenesis of milder hearing loss, which might justify sequencing of GJB2 from these subjects in the Korean population.

Multiphasic analysis of whole exome sequencing data identifies a novel mutation of ACTG1 in a nonsyndromic hearing loss family

BackgroundThe genetic heterogeneity of sensorineural hearing loss is a major hurdle to the efficient discovery of disease-causing genes. We designed a multiphasic analysis of copy number variation (CNV), linkage, and single nucleotide variation (SNV) of whole exome sequencing (WES) data for the efficient discovery of mutations causing nonsyndromic hearing loss (NSHL).ResultsFrom WES data, we identified five distinct CNV loci from a NSHL family, but they were not co-segregated among patients. Linkage analysis based on SNVs identified six candidate loci (logarithm of odds [LOD] >1.5). We selected 15 SNVs that co-segregated with NSHL in the family, which were located in six linkage candidate loci. Finally, the novel variant p.M305T in ACTG1 (DFNA20/26) was selected as a disease-causing variant.ConclusionsHere, we present a multiphasic CNV, linkage, and SNV analysis of WES data for the identification of a candidate mutation causing NSHL. Our stepwise, multiphasic approach enabled us to expedite the discovery of disease-causing variants from a large number of patient variants.

High-Resolution, Parallel Patterning of Nanoparticles via an Ion-Induced Focusing Mask

An ion-induced focusing mask under the simultaneous injection of ions and charged aerosols generates invisible electrostatic lenses around each opening, through which charged nanoparticles are convergently guided without depositing on the mask surface. The sizes of the created features become significantly smaller than those of the mask openings due to the focusing capability. It is not only demonstrated that material-independent nanoparticles including proteins can be patterned as an ordered array on any surface regardless of the conductive, nonconductive, or flexible nature of the substrate, but also that the array density can be increased. Highly sensitive gas sensors based on these focused nanoparticle patterns are fabricated via the concept.

Malleus neck-anchoring malleostapedotomy: preliminary results.

Objective To compare the hearing outcomes between 2 malleostapedotomy (MS) procedures, handle-MS, connecting the prosthesis with the malleus handle and neck-MS, connecting the prosthesis with the malleus neck. Patients Fourteen individuals having undergone MS in the setting of otosclerosis or congenital ossicular fixation from January 1983 through December 2009. Intervention Review of preoperative and postoperative audiometric data, ossicular abnormalities, and postoperative complications. Main Outcome Measures Postoperative air-bone gap (ABG), closure of ABG, and postoperative changes in bone conduction thresholds. Results Of 14 patients, 7 underwent handle-MS, and 7 underwent neck-MS. Morphologic or functional abnormalities of the incus were identified in all cases. There was no significant sensorineural hearing loss. The mean postoperative ABGs were 19.8 ± 11.9 dB in the handle-MS group and 14.7 ± 5.5 dB in the neck-MS group. The postoperative ABGs for single frequencies revealed better results for neck-MS at all frequencies (0.25, 0.5, 1, 2, 3, and 4 kHz) without statistical significance. The functional success rate (ABG closure, ⩽10 dB) was 28.6% for the handle-MS group and 42.9% for the neck-MS group (p > 0.05). Conclusion Inasmuch as neck-MS is easy to perform and yields comparable results to those of handle-MS, it may be an alternative procedure of use in selected cases of otosclerosis or stapes fixation with incus anomaly.

Large-area assembly of three-dimensional nanoparticle structures via ion assisted aerosol lithography with a multi-pin spark discharge generator

We present an approach utilizing ion assisted aerosol lithography (IAAL) with a newly designed multi-pin spark discharge generator (SDG) for fabricating large-area three-dimensional (3D) nanoparticle-structure (NPS) arrays. The design of the multi-pin SDG allows us to uniformly construct 3D NPSs on a large area of 50 mm × 50 mm in a parallel fashion at atmospheric pressure. The ion-induced focusing capability of IAAL significantly reduces the feature size of 3D NPSs compared to that of the original pre-patterns formed on a substrate. The spatial uniformity of 3D NPSs is above 95% using the present multi-pin SDG, which is far superior to that of the previous single-pin SDG with less than 32% uniformity. The effect of size distributions of nanoparticles generated via the multi-pin SDG on the 3D NPSs also has been studied. In addition, we measured spectral reflectance for the present 3D NPSs coated with Ag, demonstrating enhanced diffuse reflectance.

Distinct vestibular phenotypes in DFNA9 families with COCH variants

Mutations of COCH can cause hearing loss and less frequently vestibular symptoms. However, vestibular phenotypes, especially in terms of the location of specific variants are not well documented yet. In this study, a retrospective and prospective cohort survey was performed in two tertiary referral hospitals to demonstrate vestibular phenotypes of DFNA9 subjects with a focus on the relationship with the location of COCH mutations. Two DFNA9 subjects were recruited from the previously collected cohort, each segregating p.G38D and p.C162Y of the COCH gene. Another two DFNA9 families were newly detected by targeted resequencing of known 129 deafness genes (TRS-129). These two families segregated the p.G38D variant of the COCH gene as the causative mutation, making p.G38D the most frequent COCH mutation in our Korean cohorts. Regarding the detailed clinical phenotype of the four DFNA9 families with documented vestibular phenotypes, we were able to classify them into two groups: one (p.C162Y variant) with a Meniere’s disease (MD)-like phenotype and the other three (p.G38D variant) with significant bilateral vestibular loss without any definite MD symptoms. Distinct vestibular phenotypes depending on the location of COCH mutations were demonstrated, and this study correlates a genotype of p.G38D in COCH to the phenotype of bilateral total vestibular loss, therefore expanding the vestibular phenotypic spectrum of DFNA9 to range from bilateral vestibular loss without episodic vertigo to MD-like features with devastating episodic vertigo. In addition, the p.G38D variant of the COCH gene is suggested to be a frequent cause of progressive audiovestibular dysfunction in Koreans eventually requiring cochlear implantation.

ATP1A3 mutations can cause progressive auditory neuropathy: a new gene of auditory synaptopathy

The etiologies and prevalence of sporadic, postlingual-onset, progressive auditory neuropathy spectrum disorder (ANSD) have rarely been documented. Thus, we aimed to evaluate the prevalence and molecular etiologies of these cases. Three out of 106 sporadic progressive hearing losses turned out to manifest ANSD. Through whole exome sequencing and subsequent bioinformatics analysis, two out of the three were found to share a de novo variant, p.E818K of ATP1A3, which had been reported to cause exclusively CAPOS (cerebellar ataxia, areflexia, pes cavus, optic atrophy, and sensorineural hearing loss) syndrome. However, hearing loss induced by CAPOS has never been characterized to date. Interestingly, the first proband did not manifest any features of CAPOS, except subclinical areflexia; however, the phenotypes of second proband was compatible with that of CAPOS, making this the first reported CAPOS allele in Koreans. This ANSD phenotype was compatible with known expression of ATP1A3 mainly in the synapse between afferent nerve and inner hair cells. Based on this, cochlear implantation (CI) was performed in the first proband, leading to remarkable benefits. Collectively, the de novo ATP1A3 variant can cause postlingual-onset auditory synaptopathy, making this gene a significant contributor to sporadic progressive ANSD and a biomarker ensuring favorable short-term CI outcomes.

Benign paroxysmal positional vertigo after surgical drilling of the temporal bone.

Objective To present the clinical characteristics of secondary BPPV after surgical drilling of the temporal bone. Study Design Retrospective study. Setting Tertiary referral academic medical center. Patients Ten patients who developed BPPV after surgical procedure of temporal bone drilling were identified from 965 subjects who underwent surgical drilling of the temporal bone at Seoul National University Bundang Hospital. The localization and lateralization of BPPV were based on positional test using video eye movement recording system and videonystagmography. Main Outcome Measures Onset of BPPV after surgery, distributions of involved semicircular canals, response to particle repositioning maneuver and factors that may influence the development of secondary BPPV after surgical drilling of the temporal bone. Results Onset of positional vertigo was mostly within 3 days except 1 case (sixth postoperative day). Postoperative BPPV was usually in the contralateral ear in 9 cases (90%), which occurred predominantly on the contralateral horizontal canal in 8 patients (80%). Positional vertigo was resolved after repositioning maneuvers in every case. None of them showed aggravation of bone conduction threshold. Conclusion The incidence of BPPV after surgical drilling of the temporal bone was around 1%, and the horizontal semicircular canal of the contralateral ear was predominantly involved. Head position during surgery (head restriction to contralateral ear down) as well as limitation of head movement due to compressive mastoid bandage after surgery seems to be responsible for such predominance.

Usefulness of 1000-Hz probe tone in tympanometry according to age in Korean infants

OBJECTIVE Numerous studies have shown the superiority of a 1000-Hz frequency probe tone for evaluating the middle ear status of infants. However, most of these studies examined Caucasian populations. This study validated the 1000-Hz probe tone and evaluated the age at which it should be used in Korean infants. METHODS Data from 83 infants (43 males, 40 females; mean age 9.2±6.2 (range 1-30) months, 165 ears) were analyzed. Tympanograms were classified according to Baldwins modification of the method of Marchant et al. and correlated with results based on combined diagnostic tests, including an endoscopic examination of the tympanic membrane, myringotomy findings, and the air and bone conduction auditory brainstem response (ABR) thresholds. Data were analyzed in five age groups, each covering a 3-month range. The traces were measured for both 226- and 1000-Hz probe tones. The sensitivity and specificity for the different age groups were also determined. RESULTS For the 226-Hz probe tone, the tympanograms showed normal traces for most ears with otitis media effusions in infants younger than 12 months. By contrast, the tympanograms using the 1000-Hz probe tone showed abnormal traces in most of the infants with otitis media effusions in all age groups. In infants with no otitis media effusion, the tympanograms using both 226- and 1000-Hz probe tones were interpreted as normal in most cases in all age groups. In infants younger than 12 months, the sensitivity of the 226-Hz probe tone was very low (0-6.6%), whereas that of the 1000-Hz probe tone was very high (90-100%). In infants older than 13 months, however, the sensitivities of the 226- and 1000-Hz probe tones were 76.2% and 85.7%, respectively. Regarding specificity, the difference between the two probe tones was not significant for any age group. CONCLUSIONS This study confirmed the superiority of the 1000-Hz probe tone for evaluating the middle ear in infants. We recommend using a 1000-Hz probe tone at least up to the age of 12 months for Korean infants.