Association between Uncoupling Protein 2 Gene Ala55val Polymorphism and Sudden Sensorineural Hearing Loss.
The pathology underlying sudden sensorineural hearing loss, which is known as sudden deafness (SD), remains unknown. It affects the unilateral ear in most cases and is often accompanied by tinnitus and vertigo . The hearing level can recover within the first 1 or 2 months, and after this period, the hearing level will be fixed. Several underlying pathologies, such as vascular disturbance and viral infection, are taken into consideration [2, 3].
Sudden deafness (SD) is thought to have multiple causes, including genetic and environmental factors. To date, some gene polymorphisms have been identified for the vasculature- or inflammation-related pathogenesis of SD, such as protein kinase C-eta 1425G/A, matrix metalloproteinase-1 1607G/2G, methylenetetrahydrofolate reductase 677C/T, prothrombin 20210G/A, platelet Gly IIIaA1/A2, factor V Leiden 1691G/A, interleukin-1A-889C/T, interleukin-6C 572C/G, complement factor H 402Y/H, and nitric oxide synthase 3 894G/T polymorphisms [4-13]. Environmental factors related to lifestyle, including short sleeping times, heavy smoking, alcohol abuse, and fatigue, have been found to be risk factors for SD [14, 15].
Uncoupling proteins (UCPs) are mitochondrial transporters located in the inner mitochondrial membrane and are able to uncouple adenosine triphosphate (ATP) from mitochondrial respiration. Among the five UCPs, UCP2 has a wide expression in almost all mammalian tissues, such as white adipose tissue, kidney tissue, and liver tissue . The UCP2 mRNA transcripts have an abundant expression in the spiral and vestibular ganglia in the inner ear of rats [17, 18]. Sugiura et al.  have described that the UCP2 Ala55Val polymorphism (rs660339) was significantly associated with age-related hearing loss. The objective of the present study was to investigate the association between UCP2 polymorphism and SD.
MATERIALS and METHODS
In total, 83 patients (39 males and 44 females; mean age, 58.0[+ or -]14.2 years; range, 22-86 years) with SD were enrolled at the Nagoya University Hospital between November 2007 and March 2011, and a genetic analysis was performed on these patients. SD was defined as a type of deafness or hearing loss with an unknown etiology and with a sudden onset complicated by no cranial nerve palsy other than the auditory nerve. We used the criteria established by the Sudden Deafness Research Committee Study Group of the Ministry of Health and Welfare (1973), Japan.
The controls population were recruited from the Longitudinal Study of Aging at the National Institute for Longevity Sciences (NILS-LSA). Entrants in the NILS-LSA hospital were sampled at random from resident registrations, and they were stratified by age and sex. The area for the NILS-LSA study was located within approximately 30 km of the Nagoya University hospital. The details of the NILS-LSA have been described previously . The participants responded to a series of surveys helpful in collecting population statistics and clinical data, such as comorbidities. Those with a history of SD in the surveys were excluded. In total, 2048 participants (1033 males and 1015 females; mean age, 59.2[+ or -]10.9 years) were included as controls. These subjects completed the first examination of NILS-LSA between November 1997 and April 2000, and the analyses of UCP2 rs660339 gene were sampled.
The study protocol was reviewed and approved by the ethics committees of Nagoya University (370-4) and by the National Center for Geriatrics and Gerontology (#14, #52, and #74), and written informed consent was obtained from all subjects.
Genomic DNA was extracted from the lymphocytes isolated form peripheral blood using standard protocols, and polymerase chain reaction (PCR) amplification was performed. Genotyping using an allele-specific primer (ASP) method was also performed (Toyobo Gene Analysis, Tsuruga, Japan), as described previously . The data of the primer sequences and PCR conditions are presented in Table 1.
We evaluated the hearing levels in patients with SD using the audiometer (Model AA-79S; Rion, Tokyo, Japan) in a silent chamber. The average hearing level was expressed as the average score recorded at five frequencies (250, 500, 1000, 2000, and 4000 Hz, respectively). The hearing level results of patients with SD were assessed against the criteria of the Ministry of Health and Welfare, Japan . The recovery rate was divided into four classes: complete recovery (all frequencies on the final audiogram were .20 dB or improvement to the same degree of hearing level as in the other side of ear), remarkable improvement (improvement in the hearing level of .30 dB on an average), slight improvement (improvement in the hearing level of .10 dB but <30 dB on an average), and no change (improvement in the hearing level of <10 dB on an average). A good recovery comprises complete recovery and remarkable improvement. A poor recovery comprises slight improvement and no recovery. Because the possibility of recovery of hearing is extremely low at 1 month after the disease onset, to analyze the hearing recovery, SD patients who first visited our hospital within 1 month of disease onset were selected.
Statistical analyses were conducted using the Statistical Analysis System software, version 9.1.3 (SAS institute, Cary, NC, USA), with significance achieved at a p value of <0.05. Univariate analyses of the categorical variables were performed using the chi-squared test. Student's t test was used to evaluate the differences in the continuous variables between the two groups. Multiple logistic regression was performed to calculate the odds ratios (ORs) for SD risk concerning the UCP2 polymorphism for the multivariate analysis. Genotypes were defined as major allele homozygotes (CC), heterozygotes (CT), and minor allele homozygotes (TT) in UCP polymorphism. The major allele was determined for practical reasons.
We used the additive genetic model for the analyses, and the minor allele frequency was compared between patients with SD and controls by allocating scores of 0, 1, and 2 to major allele homozygotes, heterozygotes, and minor allele homozygotes, respectively. We used two models in this analysis, in the presence and absence of moderator variables, respectively; the former is the crude model, while the latter is the adjusted model in which age; sex; and a history of hypertension, diabetes, or dyslipidemia, were taken as moderator variables.
The characteristics of patients with SD and controls are summarized in Table 2. No significant difference in sex or age was found between patients with SD and controls. The genotype distributions are shown in Table 3. No significant difference in the genotype distributions was observed between the patients with SD and controls based on the chi-squared test.
The results of multiple logistic regression are shown in Table 4. A significant difference in SD risk was observed between patients with SD and controls in terms of UCP2 (rs660339) polymorphisms following adjustments for age, sex, and diseases, and the OR for SD risk was 1.468 (95% confidence interval, 1.056-2.040).
Allele frequency was compared between the good recovery group and poor recovery groups among patients with SD. No significant difference was found between the good and poor recovery groups (Table 5).
We conducted a case-control retrospective study in which the OR analysis was mainly used to predict disease susceptibility. This study demonstrated that the UCP2 polymorphism (rs 660339) has a significant association with SD risk. UCPs are the members of the anion carrier protein family and are located in the inner mitochondrial membrane. UCP inhibits insulin secretion and may be related to obesity, a cell damage, and diabetes. UCP exerts a protective effect against free radicals, and it is related to thermogenesis . Previous studies have investigated the association between UCP2 gene polymorphisms and the presence of diabetic complications. The polymorphisms of the UCP2 gene have been reported to be significantly associated with a risk of proliferative diabetic retinopathy in Brazilian patients with type 1 and type 2 diabetes mellitus . The mRNA expression of UCP3, UCP4, and especially UCP2 is upregulated after a unilateral labyrinthectomy in the inner ear of rats. Furthermore, the expression of UCP2, 3, and 4 is upregulated after a systemic administration of kanamycin in the mouse inner ear [17, 18]. Sugiura et al.  have described that the UCP2 T allele (rs660339) was also associated with presbycusis as in SD. The neuroprotective part against oxidative stress may be associated with the hearing function. Conversely, it is assumed that the UCP2 C allele may have a protective role to prevent sensorineural hearing loss by strengthening the antioxidative function. This was suggested in the present study of SD as well as another study of presbycusis. Manche et al.  have also reported that UCP2 (G-866 A)--another common polymorphism of this gene--was associated with presbycusis.
The present study demonstrates that UCP2 polymorphisms may be associated with the risk of developing SD, as suggested by the results of our multivariate analysis with moderating variables. Future studies including more cases are warranted.
Ethics Committee Approval: Ethics committee approval was received for this study from the ethics committees of Nagoya University (370-4) and by the National Center for Geriatrics and Gerontology (nos.14, 52, and 74).
Informed Consent: Written informed consent was obtained from all subjects who participated in this study.
Peer-review: Externally peer-reviewed.
Author contributions: Concept--Y.U., H.S., T.N.; Design--Y.K, M.T., S.S., Y.U., T.N.; Supervision- M.S.; Data Collection and/or Processing--M.T., N.N., K.K., H.O., T.Y., R.O., F.A., H.S.; Analysis and/or Interpretation--Y.K., M.T., S.S., Y.U., T.N.; Literature Search--Y.K., M.T., S.S.; Writing- Y.K., M.T., T.N.; Critical Review--F.A., H.S., Y.H, T.N.
Acknowledgements: We would like to give our sincere thanks to the researchers from Section of National Institute for Longevity Sciences, Longitudinal Study of Aging (NILS-LSA), National Center for Geriatrics and Gerontology who were involved in data collection and analyses.
Conflict of Interest: The authors have no conflict of interest to declare.
Financial Disclosure: This work was supported in part by Research Funding for Longevity Sciences (28-2) from National Center for Geriatrics and Gerontology (NCGG) and a Grant-in-Aid for Scientific Research (18K09376) from the Ministry of Education, Culture, Sports, Science and Technology, Japan
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Yusuke Koide [iD], Masaaki Teranishi [iD], Saiko Sugiura [iD], Yasue Uchida [iD], Naoki Nishio [iD]
Department of Otorhinolaryngology, Nagoya University Graduate School of Medicine, Nagoya, Japan (YK, MT, NN, KK, HO, TY, MS)
Department of Head and Neck Surgery, Aichi Cancer Center Hospital, Nagoya, Japan (YK, YH)
Toyota Josui Mental Clinic, Aichi, Japan (SS)
Department of Otorhinolaryngology, National Center for Geriatrics and Gerontology, Aichi, Japan (SS, YU)
Department of Otorhinolaryngology, Aichi Medical University School of Medicine, Aichi, Japan (YU)
Section of National Institute for Longevity Sciences, Longitudinal Study of Aging (NISL-LSA), National Center for Geriatrics and Gerontology, Aichi, Japan (RO, FA, HS)
Faculty of Health and Medical Sciences, Aichi Shukutoku University, Aichi, Japan (FA)
Graduate School of Nutritional Sciences, Nagoya University of Arts and Sciences, Aichi, Japan (HS)
Ichinomiya Medical Treatment and Habilitation Center, Aichi, Japan (TN)
ORCID IDs of the authors: Y.K. 0000-0002-4437-048X; M.T. 0000-0002-7730-0445; S.S. 0000-0001-5419-0813; Y.U. 0000-0002-3731-3890; N.N. 0000-0003-1495-0376; K.K. 0000-0001-8619-857X; H.O. 0000-0001-6776-8290; T.Y. 0000-0001-9993-0766; R.O. 0000-0001-6184-570X; F.A. 0000-0002-5444-6388; H.S. 0000-0002-0830-2041; Y.H. 0000-0002-6635-3955; T.N. 0000-0003-3930-9120; M.S. 0000-0001-7380-610X
Cite this article as: Koide Y, Teranishi M, Sugiura S, Uchida Y, Nishio N, Kato K, et al. Association between Uncoupling Protein 2 Gene Ala55val Polymorphism and Sudden Sensorineural Hearing Loss. J Int Adv Otol 2018; 14(2): 166-9.
Corresponding Address: Yusuke Koide E-mail: firstname.lastname@example.org
Submitted: 06.05.2018 * Revision Received: 11.06.2018 * Accepted: 12.06.2018 * Available Online Date: 03.08.2018
Table 1. PCR condition used in genotyping the gene ASP method Gene rs No Labeled primers Sequence (5'[right arrow]3') UCP2 rs660339 F1 (FITC) CCAGTGCGCGCTACAxCC F2 (Texas Red) CCAGTGCGCGCTACAxTC R (Biotin) TCAGAATGGTGCCCATCACA Gene rs No Amplicon (F1/R)/(F2/R) (bp) Annealing temp. ([degrees]C) Mg (mM) UCP2 rs660339 65/72 65 2.5 55 67.5 4.5 Table 2. Characteristics of case and control groups SD group Control group p No. 83 (3.9%) 2,048 (96.1%) Sex, % male 47.0 50.4 0.5375 Age (years) 58.0[+ or -]14.2 59.2[+ or -]10.9 0.3365 SD: sudden deafness Table 3. Genotype distribution in SD cases and controls Controls SD patients Total n=2048 Total n=83 Genotype n (%) n (%) (vs controls) p UCP2 (rs660339) CC 549 (26.81) 15 (18.07) 0.1571 CT 1032 (50.39) 44 (53.01) TT 467 (22.80) 24 (28.92) SD: sudden deafness; UCP2: uncoupling protein 2 Table 4. Risk of SD associated with the additive model Crude: model 1 Adjusted: model 2 Mode of 95% p 95% inheritance OR CI value OR CI p UCP2(rs660339) 1.352 0.987-1.851 0.0601 1.468 1.056-2.040 0.0222 SD: sudden deafness; UCP2: uncoupling protein 2; OR: odds ratio; CI: confidence interval Table 5. Allele frequency in SD case with good and poor recovery SD cases with SD cases with good recovery poor recovery p Number of cases 22 38 0.8289 UCP2 (rs660339) C allele, T allele 20, 24 33, 43 0.5701 Age (years) 57.2[+ or -]12.1 59.5[+ or -]15.8 0.3071 Average hearing level (dB) 74.7[+ or -]19.6 68.2[+ or -]25.7 0.1178 Period from onset to first visit (days) 4.6[+ or -]4.6 7.8[+ or -]8.8 SD: sudden deafness; UCP2: uncoupling protein 2
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|Title Annotation:||Original Article|
|Author:||Koide, Yusuke; Teranishi, Masaaki; Sugiura, Saiko; Uchida, Yasue; Nishio, Naoki; Kato, Ken; Otake, H|
|Publication:||The Journal of the International Advanced Otology|
|Article Type:||Clinical report|
|Date:||Aug 1, 2018|
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