The Laryngoscope C 2014 The American Laryngological, V
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Audiologic Presentation of Enlargement of the Vestibular Aqueduct According to the SLC26A4 Genotypes Yoon C. Rah, MD; Ah R. Kim, BS; Ja-Won Koo, MD; Jun H. Lee, MD; Seung-ha Oh, MD; Byung Y. Choi, MD Objectives/Hypothesis: To determine the distribution of the number and types of mutant alleles of SLC26A4 and their correlations with hearing phenotypes in Korean bilateral enlargement of vestibular aqueduct (EVA) patients. Study Design: Prospective cohort study. Methods: To determine the number and type of mutant alleles, Sanger sequencing of coding region of SLC26A4 was performed for 56 patients with bilateral EVA who were consecutively recruited. Their correlations with hearing phenotypes were analyzed based on 0.5-, 1-, 2-, and 3-kHz air conduction averages of pure-tone audiometry. Results: Most patients with bilateral EVA (83.9%) carried two mutant alleles of SLC26A4 (M2), and all others (16.1%) had only one detectable mutant allele of SLC26A4 (M1) in the Korean population. There were no cases with zero mutations. p.H723R/p.H723R was the most frequently observed mutant allelic pair (34%), followed by p.H723R/c.919-2A>G (20%). There was no significant difference in hearing threshold, progression, or fluctuation of hearing level between the M1 and M2 groups. However, focusing on the type of mutations exclusively in the M2 group, cases with p.H723R/c.919-2A>G were associated with more frequent progression of hearing loss during the follow-up period. The cases with p.H723R/c.919-2A>G tended to show slightly better hearing than p.H723R homozygotes, although the difference was not statistically significant. There appears to be a different genotype-auditory phenotype correlation among ethnicities. Conclusions: Our data suggest that the auditory phenotype of Korean bilateral EVA patients is more strongly correlated with the type rather than the number of mutations in SLC26A4. Key Words: SLC26A4, enlarged vestibular aqueduct, hearing. Level of Evidence: NA Laryngoscope, 125:E216–E222, 2015
INTRODUCTION The phenotypes of mutations in SLC26A4 are basically presented as pre- or perilingual onset sensorineural or mixed hearing loss, with enlargement of the vestibular aqueduct (EVA) as a radiological finding.1 Hearing loss usually fluctuates and is progressive, and the EVA is commonly combined with an incomplete partition of the apical turn of the cochlea resulting in Mondini dysplasia. SLC26A4 has been actively studied and has been shown to manifest either syndromic congenital hearing loss as Pendred syndrome (PS), which involves a thyroid
From the Department of Otorhinolaryngology–Head and Neck Surgery (Y.C.R., A.R.K. J.H.L., S.O.), Seoul National University Hospital, Seoul National University College of Medicine, Seoul; and the Department of Otorhinolaryngology–Head and Neck Surgery (J.K., B.Y.C.), Seoul National University Bundang Hospital, Seoul National University College of Medicine, Seongnam, Republic of Korea. Editor’s Note: This Manuscript was accepted for publication November 10, 2014. This study was supported by the Seoul National University Bundang Hospital Research Fund (No. 02-2011-024) (B.Y.C.) and the Korean Health Technology R&D Project, Ministry for Health, Welfare, and Family Affairs, Republic of Korea (No. A111377 (HI11C13310000) (B.Y.C.). The authors have no other funding, financial relationships, or conflicts of interest to disclose. Send correspondence to Byung Yoon Choi, MD, Department of Otorhinolaryngology, Seoul National University Bundang Hospital, 166, Gumi-ro, Bundang-gu, Seongnam-si, Gyeonggi-do, 463–707, Republic of Korea. E-mail: [email protected] DOI: 10.1002/lary.25079
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organification defect, or nonsyndromic bilateral sensorineural hearing loss (DFNB4).2 Interestingly, the number of detectable mutant alleles varies from none to both, although it is known to be inherited in an autosomal recessive pattern. According to previous reports in Caucasian subjects, approximately 25% of patients carried mutations in both alleles, and the remaining 75% carried none or only one mutation.3,4 Some phenotypic differences according to these variations have been reported. Thyroid organification defects, which are identified by abnormal results on perchlorate discharge test, are known to manifest almost exclusively in patients with mutations of both alleles.3,5 Unilateral EVA is more prevalent in patients with a mutation in one of the two alleles, whereas patients with mutations in both alleles almost always show bilateral EVA.3,5,6 There have been inconsistent reports regarding hearing presentations. Biallelic mutations were initially believed to cause more severe hearing loss and bilateral involvement,6,7 but recent studies indicated no correlation between the number of mutations and hearing presentation.8–10 Mutations in SLC26A4 are common causes of congenital hearing loss, accounting for 5% to 10% of cases of childhood sensorineural hearing loss,11 including those in East Asia.12 However, few studies have investigated the variation in the number of detectable mutant alleles and its impact on hearing in East Asians. The present study was performed to determine the Rah et al.: Audiologic Presentation of EVA
TABLE I. Distribution of Detailed Allelic Changes in SLC26A4 Mutations (N 5 56). Protein Change
Hearing Thresholds (dB)
Sex
Age, yr
Allele 1
Allele 2
Better
Worse
SH15–39 SH36–77
M M
2 10
p.H723R p.H723R
p.H723R p.H723R
113.33 98.75
115.00 101.25
SH112–231
F
30
p.H723R
p.H723R
98.75
108.75
SH112–232 SH125–258
F M
33 1
p.H723R p.H723R
p.H723R p.H723R
71.25 92.50
97.50 97.50
ID
SH126–259
F
8
p.H723R
p.H723R
65.00
108.75
SH131–272 SB1-1
M M
8 10
p.H723R p.H723R
p.H723R p.H723R
52.50 42.50
78.75 97.50
SB17–38
M
9
p.H723R
p.H723R
88.75
110.00
SB20–47 SB23–58
M M
1 30
p.H723R p.H723R
p.H723R p.H723R
86.67 —
86.67 —
SB58–105
M
4
p.H723R
p.H723R
95.00
105.00
SHJ7 SHJ8
F M
16 5
p.H723R p.H723R
p.H723R p.H723R
92.50 82.50
93.75 101.25
SHP1-1
F
4
p.H723R
p.H723R
97.50
105.00
SHP2-1 SHP3-1
M F
3 12
p.H723R p.H723R
p.H723R p.H723R
100.00 96.25
105.00 100.00
SHP4-1
F
7
p.H723R
p.H723R
93.75
97.50
SHP5-1 SH31–69
F F
4 3
p.H723R p.H723R
p.H723R c.919-2A>G
95.00 75.00
95.00 77.50
SH33–74
F
21
p.H723R
c.919-2A>G
55.00
65.00
SH56–123 SH100–214
M M
4 23
p.H723R p.H723R
c.919-2A>G c.919-2A>G
87.50 98.75
103.75 107.50
SB12–28
F
45
p.H723R
c.919-2A>G
102.50
120.00
SB104–196 SB129–222
F M
4 11
p.H723R p.H723R
c.919-2A>G c.919-2A>G
76.70 65.00
120.00 70.00
SB142–236
F
3
p.H723R
c.919-2A>G
—
—
SHJ17 SHP6-1
F M
6 16
p.H723R p.H723R
c.919-2A>G c.919-2A>G
66.25 78.75
91.25 80.00
SHP7-1
M
12
p.H723R
c.919-2A>G
108.75
118.75
SH29–65 SH71–162
M M
3 1
c.919-2A>G c.919-2A>G
— —
70.00 100.00
87.50 102.50
SH74–168
M
3
c.919-2A>G
—
85.00
120.00
G25 SHP8-1
F F
12 2
c.919-2A>G c.919-2A>G
— —
65.00 95.00
68.75 107.50
SB16–34
F
12
p.H723R
p.L676Q
93.75
103.75
SB16–35 SB28–61
F M
10 15
p.H723R p.H723R
p.L676Q p.L676Q
76.25 51.25
103.75 113.30
SHP9-1
M
11
p.H723R
p.L676Q
85.00
102.50
SH129–267 SH133–276
M M
3 3
p.H723R p.H723R
p.T410M p.T410M
81.25 —
93.75 —
SB23–54
F
5
p.H723R
p.T410M
75.00
81.67
G13 G18
M M
21 21
p.H723R p.H723R
— —
100 101.25
101.25 120
SHP10-1
F
8
p.H723R
—
103.75
106.25
SB26–59 SH123–253
M F
14 3
p.H723R p.H723R
c.131213A>G p.V510D
50.00 66.25
82.50 75.00
SH18–44
F
5
p.H723R
p.F572L
75.00
77.50
SHP11-1
M
9
p.H723R
p.M147V
75.00
120.00
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TABLE I. (Continued) Protein Change
Hearing Thresholds (dB)
Sex
Age, yr
Allele 1
Allele 2
Better
Worse
M M
2 27
p.V659L p.V138L
c.919-2A>G c.131213A>G
105.00 93.75
110.00 100.00
SB23–55
F
34
p.T410M
c.919-2A>G
95.00
97.50
SH73–165 SHP14-1
M F
3 7
p.M147V c.919-2A>G
c.919-2A>G c.919-2A>G
62.50 83.75
68.75 96.25
SB143–237
F
3
c.919-2A>G
c.131213A>G
—
—
SB137–230
F
7
p.M147V
—
42.5
97.5
ID
SHP12-1 SHP13-1
F 5 female; M 5 male.
distribution of the number and types of mutant alleles and its correlations with hearing phenotypes.
MATERIALS AND METHODS Patient Selection and Ethnicity Considerations Written informed consent was obtained from patients, or their guardians in the cases of pediatric patients, according to a protocol approved by the Seoul National University Hospital (IRB Y-H-0905-041-281) and Seoul National University Bundang Hospital Institutional Review Board (IRB-B-1007-105-402) prior to the study. The participants were recruited from among patients who attended Seoul National University Hospital or Seoul National University Bundang Hospital with congenital sensorineural hearing loss. To determine the clinical phenotype, pure-tone audiometry and temporal bone computed tomography were carried out. Auditory steady-state response (ASSR) or frequency-specific auditory brainstem response (ABR) was examined in cases in which pure-tone audiometry could not be performed due to the patient’s young age. EVA was defined when a diameter of the vestibular aqueduct measured midway between the operculum and the common crus on computed tomography scans was greater than or equal to 1.5 mm.1 Patients diagnosed with bilateral EVA with hearing loss were selected for inclusion in this study.
Molecular Genetic Testing and Evaluation of Allelic Changes In 56 EVA patients who agreed to undergo genotyping, Sanger sequencing of the coding regions of SLC26A4 was performed as described previously.13 The subjects were divided into two groups according to the number of mutant alleles of SLC26A4: M1, those with only one detectable mutant allele; M2, those with two mutant alleles. There were no patients with zero mutations (M0). The distribution of detailed allelic changes of SLC26A4 was summarized, and the frequencies and types of allelic changes were analyzed.
Audiometric Evaluations Hearing levels were calculated based primarily on 0.5-, 1-, 2-, and 3-kHz air conduction averages on pure-tone audiometry. They were measured at every visit, and the mean duration of hearing follow-up was 30.3 months (range, 5.1–98.8 months). The average age of patients was calculated using the age at the initial visit. As mentioned above, ASSR or ABR were examined in young patients who could not undergo pure-tone audiometry.
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If the hearing threshold was worse than measurable intensity on pure-tone audiometry, it was considered as 120 dB hearing loss in comparisons. We compared and analyzed hearing thresholds, symmetry, patterns of hearing loss (such as fluctuation and progression) according to the number of mutations (M1 and M2), or the type of mutation or aging. Asymmetric hearing loss was defined as a difference of more than 10 dB in the calculated hearing thresholds of both sides. Progression of hearing loss was defined as worsening of hearing threshold by more than 10 dB without any recovery in more than two frequencies on puretone audiometry. Fluctuation of hearing loss was defined as a more than 10 dB improvement of hearing threshold before or after worsening of the hearing threshold in more than two frequencies on pure-tone audiometry. We categorized patterns of hearing loss as stable hearing level with fluctuation, stable without fluctuation, progression of hearing loss with fluctuation, or progression without fluctuation. Average better or worse hearing thresholds were compared according to the number or types of mutant alleles. For detailed analysis, progression and fluctuation of hearing thresholds according to the number or types of mutant alleles were analyzed by v2 test. Relative risks for progression or fluctuation of hearing loss were calculated according to M1 group over M2 group and patients with p.H723R/c.919-2A>G over those with p.H723R/ p.H723R. Individual hearing thresholds were plotted according to patient age to determine the longitudinal progression of hearing.
Statistical Analysis For comparisons of hearing thresholds between the conditions described above, data were analyzed by Student t test, the v2 test, or two-tailed Fisher exact-test. In all analyses, a value of P G c.2168A>G
c.2168A>G c.919-2A>G
p.H723R p.H723R
p.H723R IVS7-2A>G
19 11
34% 20%
3
c.919-2A>G
—
IVS7-2A>G
—
5
9%
4 5
c.2168A>G c.2168A>G
c.2027T>A c.1229C>T
p.H723R p.H723R
p.L676Q p.T410M
4 3
7% 5%
6
c.2168A>G
—
p.H723R
—
3
5%
7 8
c.2168A>G c.2168A>G
c.131213A>G c.1529T>A
p.H723R p.H723R
IVS913A>G p.V510D
1 1
A
p.H723R
p.F572L
1
G c.1976G>C
c.439A>G c.919-2A>G
p.H723R p.V659L
p.M147V IVS7-2A>G
1 1
G
p.V138L
IVS913A>G
1
T c.439A>G
c.919-2A>G c.919-2A>G
p.T410M p.M147V
IVS7-2A>G IVS7-2A>G
1 1
G
IVS7-2A>G
IVS7-2A>G
1
G c.439A>G
c.131213A>G —
IVS7-2A>G p.M147V
IVS913A>G —
1 1
G (N 5 11)
81.4
p.H723R/p.H723R (N 5 19)
86.8
Any allele/c.919-2A>G (N 5 21)
83.8
PM/PM (N 5 29)
83.0
PM/SV or SV/SV (N 5 18)
82.6
Worse Hearing, dB
97.5
P
Difference, dB
.48
14.6
101.3 .45
16.5
100.2
.40
95.4 .59
100.2
.37
95.9 .94
13.4 14.0 13.4 12.1
99.1
.36
94.9
16.1 12.3
PM5 point mutation; SV 5 splice-site variant.
hearing loss (Table IV); the difference between the two groups was not significant (v2 test, P 5 1.00). The rate of fluctuation of hearing loss in the M1 group was 33% (4/ 12), whereas that in the M2 group was only 19% (13/68). However, the difference was not statistically significant (v2 test, P 5.27) (Table IV).
Hearing Phenotypes According to Principal Allelic Changes As we did not detect any significant differences in auditory phenotype according to the number of SLC26A4 mutations, we focused on the phenotypic differences in the M2 group according to the types of allelic changes. For this purpose, we compared hearing thresholds between the two most frequent pairs of allelic
changes in our cohort. Patients with homozygous p.H723R mutant alleles, which accounted for 34% (n 5 19) of total pairs of allelic changes, showed thresholds of 86.8 dB in the better ear and 100.2 dB in the worse ear. The group with the second most frequent pair, p.H723R/c.919-2A>G (n 5 11), showed thresholds of 81.4 dB in the better ear and 95.4 dB in the worse ear. The latter group carrying c.919-2A>G in a trans configuration with p.H723R seemed to have a slightly better hearing threshold than p.H723R homozygotes, but the difference was not statistically significant (Student t test, P 5.45 for the better ear and P 5.40 for the worse ear) (Table III). Interestingly, progression of hearing loss was observed more frequently in the group carrying c.919-2A>G in a trans configuration with p.H723R than in p.H723R homozygotes during the longitudinal follow-
TABLE IV. Progression and Fluctuation of Hearing Thresholds According to the Number of Mutations of SLC26A4 or the Kind of Allelic Changes Number of Mutations M1
M2
Total
P
1.000*
1.24 (0.34–4.53)
.271*
0.47 (0.12–1.81)
.041†
4.05 (1.03–16.01)
.694*
1.68 (0.37–7.64)
Progression
4 (33%)
26 (38%)
30 (38%)
Stable
8 (67%)
42 (62%)
50 (62%)
Fluctuation
4 (33%)
13 (19%)
17 (21%)
No fluctuation
8 (67%)
55 (81%)
63 (79%)
Odds Ratio (95% CI)
Allelic Changes p.H723R/c.919-2A>G
p.H723R/p.H723R
Progression
9 (64%)
8 (31%)
17 (42%)
Stable
5 (36%)
18 (69%)
23 (58%)
Fluctuation No fluctuation
4 (28%) 10 (72%)
5 (19%) 21 (81%)
9 (22%) 31 (78%)
*Fisher exact test. † 2 v test. CI 5 confidence interval.
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Fig. 2. Change of pure-tone threshold according to age. p.H723R/ p.H723R showed initially worse but stable hearing level with aging, whereas p.H723R/p.919-2A>G showed initially better hearing level, but hearing loss progressed with aging. PTA 5 pure-tone audiometry. [Color figure can be viewed in the online issue, which is available at www.laryngoscope.com.]
up period (v2 test, P 5.041, odds ratio 5 4.05) (Table IV). Hearing levels of p.H723R homozygotes remained stable over the follow-up period, whereas those of p.H723R/ c.919-2A>G tended to become worse over time (Fig. 2). We also compared hearing thresholds of p.H723R homozygotes with those carrying c.919-2A>G either as one of two mutant alleles or as a single heterozygote; the latter group, carrying at least one c.919-2A>G, had a slightly better hearing threshold than p.H723R homozygotes, but the difference was not statistically significant (Student t test, P 5.59 for the better ear and P 5.37 for the worse ear) (Table III). In addition, we compared hearing thresholds between the group solely with point mutations and the group possessing at least one splice-site variant. The results indicated no significant difference in hearing threshold between these two groups (Student t test, P 5.94 for the better ear and P 5.36 for the worse ear) (Table III).
DISCUSSION In our study of a Korean population, most cases of bilateral EVA (47/56; 83.9%) carried two mutant alleles of SLC26A4 (M2), whereas nine subjects (9/56; 16.1%) had only one detectable mutant allele of SLC26A4 (M1). None of the bilateral EVA cases had no mutation (i.e., M0). This strongly supports etiological homogeneity of bilateral EVA in this Korean population, in marked contrast to the different distribution of the number of SLC26A4 in Caucasian EVA subjects. In detail, substantial proportions of bilateral EVA subjects in Caucasian populations comprised M1 and M0 subjects.3,6,14 Based on the very low sibling recurrence risk of nonsyndromic EVA (NSEVA, DFNB4; NIM 600791), which is close to zero among M0 subjects, etiological heterogeneity of NSEVA was proposed in Caucasians.6,15 Our results were similar to the distribution in a Chinese population reported previously.12 The distribution of the Japanese population fell midway between Chinese and Caucasian populations.12 In this study, we did not discern any difference in severity of auditory phenotypes between the M1 and M2 Laryngoscope 125: June 2015
Korean EVA groups. An abnormal iodine organification defect indicative of PS (NIM 274600) was reported to be strongly associated with two mutant alleles of SLC26A4 (M2)3,6,14 in Caucasians. In addition, the severity of auditory phenotype in bilateral EVA subjects was reported to be more strongly correlated with the number of mutations of SLC26A4 (M2 vs. M1/M0) than with cochlear radiological structures in Caucasian populations.4,7 However, this genotype–phenotype correlation involving the number of SLC26A4 mutations does not seem to apply to East Asian populations. Recent studies conducted in Chinese and Japanese populations failed to detect the same genotype–phenotype correlation.8–10 Rather, correlations between the types of SLC26A4 mutations in the M2 group and the auditory phenotype were reported in a recent study in a Korean EVA population16; subjects carrying p.T410M or c.919-2A>G showed better residual hearing than p.H723R homozygotes.16 It was hypothesized that the better residual hearing could be due to the occurrence of normal splicing mRNA production, because c.919-2A>G may not abolish the original splice acceptor site but may instead make it leaky.16 Our study again observed the previously reported tendency and supported the hypothesis that the auditory phenotype is more strongly correlated with the type of SLC26A4 mutation than with the number of mutations, at least in the Korean population. In the present study, two mutant alleles of SLC26A4 were not associated with worse hearing thresholds, less/frequent progression of hearing loss, or less/frequent fluctuation of hearing thresholds. Rather, the M1 group showed a slightly poorer hearing threshold than the M2 group, but the difference was not statistically significant. In contrast, the presence of the most frequent splice-site variant, c.919-2A>G, in the M2 group was significantly associated with progression of hearing loss compared to p.H723R homozygotes (P 5.05). The residual hearing of subjects carrying c.919-2A>G tended to be slightly better than that of p.H723R homozygotes, but the difference did not reach statistical significance due to the small sample size. The significantly more frequent progression of hearing loss in EVA subjects carrying c.919Rah et al.: Audiologic Presentation of EVA
E221
2A>G may be associated with slightly better residual hearing or initial hearing thresholds in this group, although the residual hearing did not differ significantly between the two groups (p.H723R/c.919-2A>G vs. p.H723R homozygotes). This was compatible with the findings of longitudinal traces of hearing thresholds for the two groups in this study (Fig. 2). A number of hypotheses have been proposed regarding the mechanisms underlying the recessive phenotype even with only one mutation of SLC26A4 (M1). One of the hypotheses was the possibility of occult mutation in the regulatory region or intron of SLC26A4, which cannot be easily detected using current sequencing techniques.15 The less-severe phenotype in the M1 group than in the M2 group in Caucasians suggested that the occult mutant allele of SLC26A4 may encode some residual pendrin. Digenic inheritance involving FOXI1 or KCNJ10 in trans with one mutant allele of SLC26A4 was also proposed,17,18 but this does not seem to contribute significantly to the M1 group either in Caucasians or East Asians.9,19 The lack of differences in auditory phenotype between the M1 and M2 groups in this study, along with other reports in East Asian populations, suggested that an undetected occult mutant allele of SLC26A4 or other as-yet-undiscovered recessive gene in the East Asian M1 group may have comparable pathogenic potential to the known pathogenic mutant allele of SLC26A4, at least in terms of development of the auditory system. This is contrasted with a proposed milder pathogenic potential of the occult mutant allele of SLC26A4 in the Caucasian M1 group.15 It has been reported that noncoding regions of SLC26A4 in the Chinese M1 group had nine divergent haplotypes among the 21 no-mutation-detected SLC26A4 alleles, suggesting that there may be no common but diverse occult mutant alleles of SLC26A4.9 Alternatively, epigenetic mechanisms rather than genetic alterations may be involved in the development of EVA in this enigmatic M1 group in East Asian populations. Unfortunately, we did not perform the perchlorate discharge test in many subjects due to their reluctance; therefore, we could not address possible differences in thyroid phenotype between the M1 and M2 groups.
CONCLUSION Our data suggest that the auditory phenotype of Korean bilateral EVA patients is more correlated with the type rather than the number of mutations in
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SLC26A4. There appears to be a different genotype– auditory phenotype correlation among ethnicities, which may suggest that the molecular etiology, especially of the M1 EVA group, differs between Caucasians and Koreans.
BIBLIOGRAPHY 1. Valvassori GE, Clemis JD. The large vestibular aqueduct syndrome. Laryngoscope 1978;88:723–728. 2. Phelps PD, Coffey RA, Trembath RC, et al. Radiological malformations of the ear in Pendred syndrome. Clin Radiol 1998;53:268–273. 3. Pryor SP, Madeo AC, Reynolds JC, et al. SLC26A4/PDS genotype-phenotype correlation in hearing loss with enlargement of the vestibular aqueduct (EVA): evidence that Pendred syndrome and non-syndromic EVA are distinct clinical and genetic entities. J Med Genet 2005;42:159–165. 4. Albert S, Blons H, Jonard L, et al. SLC26A4 gene is frequently involved in nonsyndromic hearing impairment with enlarged vestibular aqueduct in Caucasian populations. Eur J Hum Genet 2006;14:773–779. 5. Madeo AC, Manichaikul A, Reynolds JC, et al. Evaluation of the thyroid in patients with hearing loss and enlarged vestibular aqueducts. Arch Otolaryngol Head Neck Surg 2009;135:670–676. 6. Azaiez H, Yang T, Prasad S, et al. Genotype-phenotype correlations for SLC26A4-related deafness. Hum Genet 2007;122:451–457. 7. King KA, Choi BY, Zalewski C, et al. SLC26A4 genotype, but not cochlear radiologic structure, is correlated with hearing loss in ears with an enlarged vestibular aqueduct. Laryngoscope 2010;120:384–389. 8. Reyes S, Wang G, Ouyang X, et al. Mutation analysis of SLC26A4 in mainland Chinese patients with enlarged vestibular aqueduct. Otolaryngol Head Neck Surg 2009;141:502–508. 9. Wu CC, Lu YC, Chen PJ, et al. Phenotypic analyses and mutation screening of the SLC26A4 and FOXI1 genes in 101 Taiwanese families with bilateral nonsyndromic enlarged vestibular aqueduct (DFNB4) or Pendred syndrome. Audiol Neurootol 2010;15:57–66. 10. Miyazawa M, Nishio SY, Usami S, et al.; Deafness Gene Study Consortium. Mutation spectrum and genotype-phenotype correlation of hearing loss patients caused by SLC26A4 mutations in the Japanese: a large cohort study. J Hum Genet 2014;59:262–268. 11. Park HJ, Shaukat S, Liu XZ, et al. Origins and frequencies of SLC26A4 (PDS) mutations in east and south Asians: global implications for the epidemiology of deafness. J Med Genet 2003;40:242–248. 12. Wang QJ, Zhao YL, Rao SQ, et al. A distinct spectrum of SLC26A4 mutations in patients with enlarged vestibular aqueduct in China. Clin Genet 2007;72:245–254. 13. Choi BY, Stewart AK, Nishimura KK, et al. Efficient molecular genetic diagnosis of enlarged vestibular aqueducts in East Asians. Genet Test Mol Biomarkers 2009;13:679–687. 14. Choi BY, Stewart AK, Madeo AC, et al. Hypo-functional SLC26A4 variants associated with nonsyndromic hearing loss and enlargement of the vestibular aqueduct: genotype-phenotype correlation or coincidental polymorphisms? Hum Mutat 2009;30:599–608. 15. Choi BY, Madeo AC, King KA, et al. Segregation of enlarged vestibular aqueducts in families with non-diagnostic SLC26A4 genotypes. J Med Genet 2009;46:856–861. 16. Lee HJ, Jung J, Shin JW, et al. Correlation between genotype and phenotype in patients with bi-allelic SLC26A4 mutations. Clin Genet 2014;86: 270–275. 17. Yang T, Gurrola JG, Wu H, et al. Mutations of KCNJ10 together with mutations of SLC26A4 cause digenic nonsyndromic hearing loss associated with enlarged vestibular aqueduct syndrome. Am J Hum Genet 2009;84:651–657. 18. Yang T, Vidarsson H, Rodrigo-Blomqvist S, et al. Transcriptional control of SLC26A4 is involved in Pendred syndrome and nonsyndromic enlargement of vestibular aqueduct (DFNB4). Am J Hum Genet 2007;80:1055–1063. 19. Ito T, Choi BY, King KA, et al. SLC26A4 genotypes and phenotypes associated with enlargement of the vestibular aqueduct. Cell Physiol Biochem 2011;28:545–552.
Rah et al.: Audiologic Presentation of EVA
Rhinological and Otological Society, Inc.
Audiologic Presentation of Enlargement of the Vestibular Aqueduct According to the SLC26A4 Genotypes Yoon C. Rah, MD; Ah R. Kim, BS; Ja-Won Koo, MD; Jun H. Lee, MD; Seung-ha Oh, MD; Byung Y. Choi, MD Objectives/Hypothesis: To determine the distribution of the number and types of mutant alleles of SLC26A4 and their correlations with hearing phenotypes in Korean bilateral enlargement of vestibular aqueduct (EVA) patients. Study Design: Prospective cohort study. Methods: To determine the number and type of mutant alleles, Sanger sequencing of coding region of SLC26A4 was performed for 56 patients with bilateral EVA who were consecutively recruited. Their correlations with hearing phenotypes were analyzed based on 0.5-, 1-, 2-, and 3-kHz air conduction averages of pure-tone audiometry. Results: Most patients with bilateral EVA (83.9%) carried two mutant alleles of SLC26A4 (M2), and all others (16.1%) had only one detectable mutant allele of SLC26A4 (M1) in the Korean population. There were no cases with zero mutations. p.H723R/p.H723R was the most frequently observed mutant allelic pair (34%), followed by p.H723R/c.919-2A>G (20%). There was no significant difference in hearing threshold, progression, or fluctuation of hearing level between the M1 and M2 groups. However, focusing on the type of mutations exclusively in the M2 group, cases with p.H723R/c.919-2A>G were associated with more frequent progression of hearing loss during the follow-up period. The cases with p.H723R/c.919-2A>G tended to show slightly better hearing than p.H723R homozygotes, although the difference was not statistically significant. There appears to be a different genotype-auditory phenotype correlation among ethnicities. Conclusions: Our data suggest that the auditory phenotype of Korean bilateral EVA patients is more strongly correlated with the type rather than the number of mutations in SLC26A4. Key Words: SLC26A4, enlarged vestibular aqueduct, hearing. Level of Evidence: NA Laryngoscope, 125:E216–E222, 2015
INTRODUCTION The phenotypes of mutations in SLC26A4 are basically presented as pre- or perilingual onset sensorineural or mixed hearing loss, with enlargement of the vestibular aqueduct (EVA) as a radiological finding.1 Hearing loss usually fluctuates and is progressive, and the EVA is commonly combined with an incomplete partition of the apical turn of the cochlea resulting in Mondini dysplasia. SLC26A4 has been actively studied and has been shown to manifest either syndromic congenital hearing loss as Pendred syndrome (PS), which involves a thyroid
From the Department of Otorhinolaryngology–Head and Neck Surgery (Y.C.R., A.R.K. J.H.L., S.O.), Seoul National University Hospital, Seoul National University College of Medicine, Seoul; and the Department of Otorhinolaryngology–Head and Neck Surgery (J.K., B.Y.C.), Seoul National University Bundang Hospital, Seoul National University College of Medicine, Seongnam, Republic of Korea. Editor’s Note: This Manuscript was accepted for publication November 10, 2014. This study was supported by the Seoul National University Bundang Hospital Research Fund (No. 02-2011-024) (B.Y.C.) and the Korean Health Technology R&D Project, Ministry for Health, Welfare, and Family Affairs, Republic of Korea (No. A111377 (HI11C13310000) (B.Y.C.). The authors have no other funding, financial relationships, or conflicts of interest to disclose. Send correspondence to Byung Yoon Choi, MD, Department of Otorhinolaryngology, Seoul National University Bundang Hospital, 166, Gumi-ro, Bundang-gu, Seongnam-si, Gyeonggi-do, 463–707, Republic of Korea. E-mail: [email protected] DOI: 10.1002/lary.25079
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organification defect, or nonsyndromic bilateral sensorineural hearing loss (DFNB4).2 Interestingly, the number of detectable mutant alleles varies from none to both, although it is known to be inherited in an autosomal recessive pattern. According to previous reports in Caucasian subjects, approximately 25% of patients carried mutations in both alleles, and the remaining 75% carried none or only one mutation.3,4 Some phenotypic differences according to these variations have been reported. Thyroid organification defects, which are identified by abnormal results on perchlorate discharge test, are known to manifest almost exclusively in patients with mutations of both alleles.3,5 Unilateral EVA is more prevalent in patients with a mutation in one of the two alleles, whereas patients with mutations in both alleles almost always show bilateral EVA.3,5,6 There have been inconsistent reports regarding hearing presentations. Biallelic mutations were initially believed to cause more severe hearing loss and bilateral involvement,6,7 but recent studies indicated no correlation between the number of mutations and hearing presentation.8–10 Mutations in SLC26A4 are common causes of congenital hearing loss, accounting for 5% to 10% of cases of childhood sensorineural hearing loss,11 including those in East Asia.12 However, few studies have investigated the variation in the number of detectable mutant alleles and its impact on hearing in East Asians. The present study was performed to determine the Rah et al.: Audiologic Presentation of EVA
TABLE I. Distribution of Detailed Allelic Changes in SLC26A4 Mutations (N 5 56). Protein Change
Hearing Thresholds (dB)
Sex
Age, yr
Allele 1
Allele 2
Better
Worse
SH15–39 SH36–77
M M
2 10
p.H723R p.H723R
p.H723R p.H723R
113.33 98.75
115.00 101.25
SH112–231
F
30
p.H723R
p.H723R
98.75
108.75
SH112–232 SH125–258
F M
33 1
p.H723R p.H723R
p.H723R p.H723R
71.25 92.50
97.50 97.50
ID
SH126–259
F
8
p.H723R
p.H723R
65.00
108.75
SH131–272 SB1-1
M M
8 10
p.H723R p.H723R
p.H723R p.H723R
52.50 42.50
78.75 97.50
SB17–38
M
9
p.H723R
p.H723R
88.75
110.00
SB20–47 SB23–58
M M
1 30
p.H723R p.H723R
p.H723R p.H723R
86.67 —
86.67 —
SB58–105
M
4
p.H723R
p.H723R
95.00
105.00
SHJ7 SHJ8
F M
16 5
p.H723R p.H723R
p.H723R p.H723R
92.50 82.50
93.75 101.25
SHP1-1
F
4
p.H723R
p.H723R
97.50
105.00
SHP2-1 SHP3-1
M F
3 12
p.H723R p.H723R
p.H723R p.H723R
100.00 96.25
105.00 100.00
SHP4-1
F
7
p.H723R
p.H723R
93.75
97.50
SHP5-1 SH31–69
F F
4 3
p.H723R p.H723R
p.H723R c.919-2A>G
95.00 75.00
95.00 77.50
SH33–74
F
21
p.H723R
c.919-2A>G
55.00
65.00
SH56–123 SH100–214
M M
4 23
p.H723R p.H723R
c.919-2A>G c.919-2A>G
87.50 98.75
103.75 107.50
SB12–28
F
45
p.H723R
c.919-2A>G
102.50
120.00
SB104–196 SB129–222
F M
4 11
p.H723R p.H723R
c.919-2A>G c.919-2A>G
76.70 65.00
120.00 70.00
SB142–236
F
3
p.H723R
c.919-2A>G
—
—
SHJ17 SHP6-1
F M
6 16
p.H723R p.H723R
c.919-2A>G c.919-2A>G
66.25 78.75
91.25 80.00
SHP7-1
M
12
p.H723R
c.919-2A>G
108.75
118.75
SH29–65 SH71–162
M M
3 1
c.919-2A>G c.919-2A>G
— —
70.00 100.00
87.50 102.50
SH74–168
M
3
c.919-2A>G
—
85.00
120.00
G25 SHP8-1
F F
12 2
c.919-2A>G c.919-2A>G
— —
65.00 95.00
68.75 107.50
SB16–34
F
12
p.H723R
p.L676Q
93.75
103.75
SB16–35 SB28–61
F M
10 15
p.H723R p.H723R
p.L676Q p.L676Q
76.25 51.25
103.75 113.30
SHP9-1
M
11
p.H723R
p.L676Q
85.00
102.50
SH129–267 SH133–276
M M
3 3
p.H723R p.H723R
p.T410M p.T410M
81.25 —
93.75 —
SB23–54
F
5
p.H723R
p.T410M
75.00
81.67
G13 G18
M M
21 21
p.H723R p.H723R
— —
100 101.25
101.25 120
SHP10-1
F
8
p.H723R
—
103.75
106.25
SB26–59 SH123–253
M F
14 3
p.H723R p.H723R
c.131213A>G p.V510D
50.00 66.25
82.50 75.00
SH18–44
F
5
p.H723R
p.F572L
75.00
77.50
SHP11-1
M
9
p.H723R
p.M147V
75.00
120.00
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TABLE I. (Continued) Protein Change
Hearing Thresholds (dB)
Sex
Age, yr
Allele 1
Allele 2
Better
Worse
M M
2 27
p.V659L p.V138L
c.919-2A>G c.131213A>G
105.00 93.75
110.00 100.00
SB23–55
F
34
p.T410M
c.919-2A>G
95.00
97.50
SH73–165 SHP14-1
M F
3 7
p.M147V c.919-2A>G
c.919-2A>G c.919-2A>G
62.50 83.75
68.75 96.25
SB143–237
F
3
c.919-2A>G
c.131213A>G
—
—
SB137–230
F
7
p.M147V
—
42.5
97.5
ID
SHP12-1 SHP13-1
F 5 female; M 5 male.
distribution of the number and types of mutant alleles and its correlations with hearing phenotypes.
MATERIALS AND METHODS Patient Selection and Ethnicity Considerations Written informed consent was obtained from patients, or their guardians in the cases of pediatric patients, according to a protocol approved by the Seoul National University Hospital (IRB Y-H-0905-041-281) and Seoul National University Bundang Hospital Institutional Review Board (IRB-B-1007-105-402) prior to the study. The participants were recruited from among patients who attended Seoul National University Hospital or Seoul National University Bundang Hospital with congenital sensorineural hearing loss. To determine the clinical phenotype, pure-tone audiometry and temporal bone computed tomography were carried out. Auditory steady-state response (ASSR) or frequency-specific auditory brainstem response (ABR) was examined in cases in which pure-tone audiometry could not be performed due to the patient’s young age. EVA was defined when a diameter of the vestibular aqueduct measured midway between the operculum and the common crus on computed tomography scans was greater than or equal to 1.5 mm.1 Patients diagnosed with bilateral EVA with hearing loss were selected for inclusion in this study.
Molecular Genetic Testing and Evaluation of Allelic Changes In 56 EVA patients who agreed to undergo genotyping, Sanger sequencing of the coding regions of SLC26A4 was performed as described previously.13 The subjects were divided into two groups according to the number of mutant alleles of SLC26A4: M1, those with only one detectable mutant allele; M2, those with two mutant alleles. There were no patients with zero mutations (M0). The distribution of detailed allelic changes of SLC26A4 was summarized, and the frequencies and types of allelic changes were analyzed.
Audiometric Evaluations Hearing levels were calculated based primarily on 0.5-, 1-, 2-, and 3-kHz air conduction averages on pure-tone audiometry. They were measured at every visit, and the mean duration of hearing follow-up was 30.3 months (range, 5.1–98.8 months). The average age of patients was calculated using the age at the initial visit. As mentioned above, ASSR or ABR were examined in young patients who could not undergo pure-tone audiometry.
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If the hearing threshold was worse than measurable intensity on pure-tone audiometry, it was considered as 120 dB hearing loss in comparisons. We compared and analyzed hearing thresholds, symmetry, patterns of hearing loss (such as fluctuation and progression) according to the number of mutations (M1 and M2), or the type of mutation or aging. Asymmetric hearing loss was defined as a difference of more than 10 dB in the calculated hearing thresholds of both sides. Progression of hearing loss was defined as worsening of hearing threshold by more than 10 dB without any recovery in more than two frequencies on puretone audiometry. Fluctuation of hearing loss was defined as a more than 10 dB improvement of hearing threshold before or after worsening of the hearing threshold in more than two frequencies on pure-tone audiometry. We categorized patterns of hearing loss as stable hearing level with fluctuation, stable without fluctuation, progression of hearing loss with fluctuation, or progression without fluctuation. Average better or worse hearing thresholds were compared according to the number or types of mutant alleles. For detailed analysis, progression and fluctuation of hearing thresholds according to the number or types of mutant alleles were analyzed by v2 test. Relative risks for progression or fluctuation of hearing loss were calculated according to M1 group over M2 group and patients with p.H723R/c.919-2A>G over those with p.H723R/ p.H723R. Individual hearing thresholds were plotted according to patient age to determine the longitudinal progression of hearing.
Statistical Analysis For comparisons of hearing thresholds between the conditions described above, data were analyzed by Student t test, the v2 test, or two-tailed Fisher exact-test. In all analyses, a value of P G c.2168A>G
c.2168A>G c.919-2A>G
p.H723R p.H723R
p.H723R IVS7-2A>G
19 11
34% 20%
3
c.919-2A>G
—
IVS7-2A>G
—
5
9%
4 5
c.2168A>G c.2168A>G
c.2027T>A c.1229C>T
p.H723R p.H723R
p.L676Q p.T410M
4 3
7% 5%
6
c.2168A>G
—
p.H723R
—
3
5%
7 8
c.2168A>G c.2168A>G
c.131213A>G c.1529T>A
p.H723R p.H723R
IVS913A>G p.V510D
1 1
A
p.H723R
p.F572L
1
G c.1976G>C
c.439A>G c.919-2A>G
p.H723R p.V659L
p.M147V IVS7-2A>G
1 1
G
p.V138L
IVS913A>G
1
T c.439A>G
c.919-2A>G c.919-2A>G
p.T410M p.M147V
IVS7-2A>G IVS7-2A>G
1 1
G
IVS7-2A>G
IVS7-2A>G
1
G c.439A>G
c.131213A>G —
IVS7-2A>G p.M147V
IVS913A>G —
1 1
G (N 5 11)
81.4
p.H723R/p.H723R (N 5 19)
86.8
Any allele/c.919-2A>G (N 5 21)
83.8
PM/PM (N 5 29)
83.0
PM/SV or SV/SV (N 5 18)
82.6
Worse Hearing, dB
97.5
P
Difference, dB
.48
14.6
101.3 .45
16.5
100.2
.40
95.4 .59
100.2
.37
95.9 .94
13.4 14.0 13.4 12.1
99.1
.36
94.9
16.1 12.3
PM5 point mutation; SV 5 splice-site variant.
hearing loss (Table IV); the difference between the two groups was not significant (v2 test, P 5 1.00). The rate of fluctuation of hearing loss in the M1 group was 33% (4/ 12), whereas that in the M2 group was only 19% (13/68). However, the difference was not statistically significant (v2 test, P 5.27) (Table IV).
Hearing Phenotypes According to Principal Allelic Changes As we did not detect any significant differences in auditory phenotype according to the number of SLC26A4 mutations, we focused on the phenotypic differences in the M2 group according to the types of allelic changes. For this purpose, we compared hearing thresholds between the two most frequent pairs of allelic
changes in our cohort. Patients with homozygous p.H723R mutant alleles, which accounted for 34% (n 5 19) of total pairs of allelic changes, showed thresholds of 86.8 dB in the better ear and 100.2 dB in the worse ear. The group with the second most frequent pair, p.H723R/c.919-2A>G (n 5 11), showed thresholds of 81.4 dB in the better ear and 95.4 dB in the worse ear. The latter group carrying c.919-2A>G in a trans configuration with p.H723R seemed to have a slightly better hearing threshold than p.H723R homozygotes, but the difference was not statistically significant (Student t test, P 5.45 for the better ear and P 5.40 for the worse ear) (Table III). Interestingly, progression of hearing loss was observed more frequently in the group carrying c.919-2A>G in a trans configuration with p.H723R than in p.H723R homozygotes during the longitudinal follow-
TABLE IV. Progression and Fluctuation of Hearing Thresholds According to the Number of Mutations of SLC26A4 or the Kind of Allelic Changes Number of Mutations M1
M2
Total
P
1.000*
1.24 (0.34–4.53)
.271*
0.47 (0.12–1.81)
.041†
4.05 (1.03–16.01)
.694*
1.68 (0.37–7.64)
Progression
4 (33%)
26 (38%)
30 (38%)
Stable
8 (67%)
42 (62%)
50 (62%)
Fluctuation
4 (33%)
13 (19%)
17 (21%)
No fluctuation
8 (67%)
55 (81%)
63 (79%)
Odds Ratio (95% CI)
Allelic Changes p.H723R/c.919-2A>G
p.H723R/p.H723R
Progression
9 (64%)
8 (31%)
17 (42%)
Stable
5 (36%)
18 (69%)
23 (58%)
Fluctuation No fluctuation
4 (28%) 10 (72%)
5 (19%) 21 (81%)
9 (22%) 31 (78%)
*Fisher exact test. † 2 v test. CI 5 confidence interval.
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Fig. 2. Change of pure-tone threshold according to age. p.H723R/ p.H723R showed initially worse but stable hearing level with aging, whereas p.H723R/p.919-2A>G showed initially better hearing level, but hearing loss progressed with aging. PTA 5 pure-tone audiometry. [Color figure can be viewed in the online issue, which is available at www.laryngoscope.com.]
up period (v2 test, P 5.041, odds ratio 5 4.05) (Table IV). Hearing levels of p.H723R homozygotes remained stable over the follow-up period, whereas those of p.H723R/ c.919-2A>G tended to become worse over time (Fig. 2). We also compared hearing thresholds of p.H723R homozygotes with those carrying c.919-2A>G either as one of two mutant alleles or as a single heterozygote; the latter group, carrying at least one c.919-2A>G, had a slightly better hearing threshold than p.H723R homozygotes, but the difference was not statistically significant (Student t test, P 5.59 for the better ear and P 5.37 for the worse ear) (Table III). In addition, we compared hearing thresholds between the group solely with point mutations and the group possessing at least one splice-site variant. The results indicated no significant difference in hearing threshold between these two groups (Student t test, P 5.94 for the better ear and P 5.36 for the worse ear) (Table III).
DISCUSSION In our study of a Korean population, most cases of bilateral EVA (47/56; 83.9%) carried two mutant alleles of SLC26A4 (M2), whereas nine subjects (9/56; 16.1%) had only one detectable mutant allele of SLC26A4 (M1). None of the bilateral EVA cases had no mutation (i.e., M0). This strongly supports etiological homogeneity of bilateral EVA in this Korean population, in marked contrast to the different distribution of the number of SLC26A4 in Caucasian EVA subjects. In detail, substantial proportions of bilateral EVA subjects in Caucasian populations comprised M1 and M0 subjects.3,6,14 Based on the very low sibling recurrence risk of nonsyndromic EVA (NSEVA, DFNB4; NIM 600791), which is close to zero among M0 subjects, etiological heterogeneity of NSEVA was proposed in Caucasians.6,15 Our results were similar to the distribution in a Chinese population reported previously.12 The distribution of the Japanese population fell midway between Chinese and Caucasian populations.12 In this study, we did not discern any difference in severity of auditory phenotypes between the M1 and M2 Laryngoscope 125: June 2015
Korean EVA groups. An abnormal iodine organification defect indicative of PS (NIM 274600) was reported to be strongly associated with two mutant alleles of SLC26A4 (M2)3,6,14 in Caucasians. In addition, the severity of auditory phenotype in bilateral EVA subjects was reported to be more strongly correlated with the number of mutations of SLC26A4 (M2 vs. M1/M0) than with cochlear radiological structures in Caucasian populations.4,7 However, this genotype–phenotype correlation involving the number of SLC26A4 mutations does not seem to apply to East Asian populations. Recent studies conducted in Chinese and Japanese populations failed to detect the same genotype–phenotype correlation.8–10 Rather, correlations between the types of SLC26A4 mutations in the M2 group and the auditory phenotype were reported in a recent study in a Korean EVA population16; subjects carrying p.T410M or c.919-2A>G showed better residual hearing than p.H723R homozygotes.16 It was hypothesized that the better residual hearing could be due to the occurrence of normal splicing mRNA production, because c.919-2A>G may not abolish the original splice acceptor site but may instead make it leaky.16 Our study again observed the previously reported tendency and supported the hypothesis that the auditory phenotype is more strongly correlated with the type of SLC26A4 mutation than with the number of mutations, at least in the Korean population. In the present study, two mutant alleles of SLC26A4 were not associated with worse hearing thresholds, less/frequent progression of hearing loss, or less/frequent fluctuation of hearing thresholds. Rather, the M1 group showed a slightly poorer hearing threshold than the M2 group, but the difference was not statistically significant. In contrast, the presence of the most frequent splice-site variant, c.919-2A>G, in the M2 group was significantly associated with progression of hearing loss compared to p.H723R homozygotes (P 5.05). The residual hearing of subjects carrying c.919-2A>G tended to be slightly better than that of p.H723R homozygotes, but the difference did not reach statistical significance due to the small sample size. The significantly more frequent progression of hearing loss in EVA subjects carrying c.919Rah et al.: Audiologic Presentation of EVA
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2A>G may be associated with slightly better residual hearing or initial hearing thresholds in this group, although the residual hearing did not differ significantly between the two groups (p.H723R/c.919-2A>G vs. p.H723R homozygotes). This was compatible with the findings of longitudinal traces of hearing thresholds for the two groups in this study (Fig. 2). A number of hypotheses have been proposed regarding the mechanisms underlying the recessive phenotype even with only one mutation of SLC26A4 (M1). One of the hypotheses was the possibility of occult mutation in the regulatory region or intron of SLC26A4, which cannot be easily detected using current sequencing techniques.15 The less-severe phenotype in the M1 group than in the M2 group in Caucasians suggested that the occult mutant allele of SLC26A4 may encode some residual pendrin. Digenic inheritance involving FOXI1 or KCNJ10 in trans with one mutant allele of SLC26A4 was also proposed,17,18 but this does not seem to contribute significantly to the M1 group either in Caucasians or East Asians.9,19 The lack of differences in auditory phenotype between the M1 and M2 groups in this study, along with other reports in East Asian populations, suggested that an undetected occult mutant allele of SLC26A4 or other as-yet-undiscovered recessive gene in the East Asian M1 group may have comparable pathogenic potential to the known pathogenic mutant allele of SLC26A4, at least in terms of development of the auditory system. This is contrasted with a proposed milder pathogenic potential of the occult mutant allele of SLC26A4 in the Caucasian M1 group.15 It has been reported that noncoding regions of SLC26A4 in the Chinese M1 group had nine divergent haplotypes among the 21 no-mutation-detected SLC26A4 alleles, suggesting that there may be no common but diverse occult mutant alleles of SLC26A4.9 Alternatively, epigenetic mechanisms rather than genetic alterations may be involved in the development of EVA in this enigmatic M1 group in East Asian populations. Unfortunately, we did not perform the perchlorate discharge test in many subjects due to their reluctance; therefore, we could not address possible differences in thyroid phenotype between the M1 and M2 groups.
CONCLUSION Our data suggest that the auditory phenotype of Korean bilateral EVA patients is more correlated with the type rather than the number of mutations in
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SLC26A4. There appears to be a different genotype– auditory phenotype correlation among ethnicities, which may suggest that the molecular etiology, especially of the M1 EVA group, differs between Caucasians and Koreans.
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