非综合征型听神经病患者OTOF基因筛查研究
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摘要
目的对非综合征型听神经病患者进行OTOF基因突变筛查,了解中国听神经病患者中OTOF基因的突变频谱。方法选取西京医院耳鼻咽喉头颈外科门诊2009-2018年确诊的听神经病患者248名以及听力正常的健康志愿者100名,采集外周静脉血提取全基因组DNA。针对OTOF基因全部48个外显子设计引物,采用聚合酶链式反应扩增外显子及其侧翼序列进行Sanger测序,应用Seqman软件与标准序列进行比对。结果 248例听神经病患者中共检出9种可能致病的变异方式,包括7种错义突变和2种剪切位置的变化。同时,检出4种已有文献报道的多态性改变。所有变异方式在健康志愿者中均未发现。结论可能致病突变集中于Exon40、Intron32、Exon12、Ex?on18、Exon24、Intron28和Exon7中,在门诊耳聋患者或者听力正常人群中,优先开展以上外显子的筛查。OTOF基因突变并不是患者唯一的致病原因,还应进行其他相关基因的筛查。
Objective To investigate mutation spectrum of the OTOF gene among Chinese patients with auditory neuropathy. Methods Genomic DNA samples were extracted from peripheral venous blood in 248 outpatients diagnosed with non-syndromic auditory neuropathy at Xijing Hospital from 2009 to 2018 and 100 healthy unrelated individuals with normal hearing. PCR primers were designed to amplify the whole 48 exons and their flanking sequences. The amplified target fragments were sequenced directly. The Seqman software was used analyze the sequence data. Results In this study, we found 9 probable pathogenic mutations, including 7 missense mutations and 2 splice-site changes. Meanwhile, 4 reported polymorphic changes were also detected. None of the variants was found in the 100 healthy subjects.Conclusion The probable pathogenic mutations appear to be concentrated in Exon40, Intron32, Exon12, Exon18, Exon24, Intron28 and Exon7. OTOF screening of these exons are therefore preferred in patients with auditory neuropathy or other types of hearing loss. Towards the findings, OTOF singe allele mutations are not the only cause of auditory neuropathy and other related genes should also be screened.
Objective To investigate mutation spectrum of the OTOF gene among Chinese patients with auditory neuropathy. Methods Genomic DNA samples were extracted from peripheral venous blood in 248 outpatients diagnosed with non-syndromic auditory neuropathy at Xijing Hospital from 2009 to 2018 and 100 healthy unrelated individuals with normal hearing. PCR primers were designed to amplify the whole 48 exons and their flanking sequences. The amplified target fragments were sequenced directly. The Seqman software was used analyze the sequence data. Results In this study, we found 9 probable pathogenic mutations, including 7 missense mutations and 2 splice-site changes. Meanwhile, 4 reported polymorphic changes were also detected. None of the variants was found in the 100 healthy subjects.Conclusion The probable pathogenic mutations appear to be concentrated in Exon40, Intron32, Exon12, Exon18, Exon24, Intron28 and Exon7. OTOF screening of these exons are therefore preferred in patients with auditory neuropathy or other types of hearing loss. Towards the findings, OTOF singe allele mutations are not the only cause of auditory neuropathy and other related genes should also be screened.
引文
1 Starr A,Picton T W,Sininger Y,et al.Auditory neuropathy[J].Brain,1996,119(Pt 3):741-753.
2王锦玲,石力,薛飞,等.听神经病听力学特征及病损部位分析[J].听力学及言语疾病杂志,2007(2):89-97.Wang JL,Shi L,Xue F,et al.Differential audiological assessment of patients with auditory neuropathy[J].Journal of Audiology and Speech Pathology,2007(2):89-97.
3王秋菊.听神经病的临床及基础研究[J].国际耳鼻咽喉头颈外科杂志,2006(06):394-397.Wang QJ.Clinical and basic research of auditory neuropathy[J].Int J Otolaryngol Head Neck Surg,2006,30(06):394-397.
4 Varga R,Kelley P M,Keats B J,et al.Non-syndromic recessive auditory neuropathy is the result of mutations in the otoferlin(OTOF)gene[J].J Med Genet,2003,40(1):45-50.
5王锦玲,高磊,薛飞,等.听神经病的临床与听功能特征[J].临床耳鼻咽喉科杂志,2002(10):518-520.Wang JL,Gao L,Xue F,et al.Auditory neuropathy[J].Journal of Clinical Otorhinolaryngology Head and Neck Surgery,2002(10):518-520.
6 Houseman M J,Jackson A P,Al-Gazali L I,et al.A novel mutation in a family with non-syndromic sensorineural hearing loss that disrupts the newly characterised OTOF long isoforms[J].JMed Genet,2001,38(8):E25.
7 Wang J,Fan Y Y,Wang S J,et al.Variants of OTOF and PJVKgenes in Chinese patients with auditory neuropathy spectrum disorder[J].PLoS One,2011,6(9):e24000.
8 Romanos J,Kimura L,Favero M L,et al.Novel OTOF mutations in Brazilian patients with auditory neuropathy[J].J Hum Genet,2009,54(7):382-385.
9 Wang D Y,Wang Y C,Weil D,et al.Screening mutations of OTOF gene in Chinese patients with auditory neuropathy,including a familial case of temperature-sensitive auditory neuropathy[J].BMC Med Genet,2010,11:79.
10 Chiu Y H,Wu C C,Lu Y C,et al.Mutations in the OTOF gene in Taiwanese patients with auditory neuropathy[J].Audiol Neurootol,2010,15(6):364-374.
11 Varga R,Avenarius M R,Kelley P M,et al.OTOF mutations revealed by genetic analysis of hearing loss families including a potential temperature sensitive auditory neuropathy allele[J].JMed Genet,2006,43(7):576-581.
12 Yasunaga S,Grati M,Cohen-Salmon M,et al.A mutation in OTOF,encoding otoferlin,a FER-1-like protein,causes DFNB9,a nonsyndromic form of deafness[J].Nat Genet,1999,21(4):363-369.
13 Roux I,Safieddine S,Nouvian R,et al.Otoferlin,defective in a human deafness form,is essential for exocytosis at the auditory ribbon synapse[J].Cell,2006,127(2):277-289.
14齐悦,张秋静,王大勇,等.听神经病谱系障碍患者SLC17A8基因筛查研究[J].中华耳科学杂志,2015(02):224-228.Qi Y,Zhang QJ,Wang DY,et al.Mutation screening of SLC17A8 gene in patients with auditory neuropathy spectrum disorder[J].Chinese Journal of Otology.2015(02):224-228.
15 Wang Q J,Li Q Z,Rao S Q,et al.AUNX1,a novel locus responsible for X linked recessive auditory and peripheral neuropathy,maps to Xq23-27.3[J].J Med Genet,2006,43(7):e33.
16 Zong L,Guan J,Ealy M,et al.Mutations in apoptosis-inducing factor cause X-linked recessive auditory neuropathy spectrum disorder[J].J Med Genet,2015,52(8):523-531.
2王锦玲,石力,薛飞,等.听神经病听力学特征及病损部位分析[J].听力学及言语疾病杂志,2007(2):89-97.Wang JL,Shi L,Xue F,et al.Differential audiological assessment of patients with auditory neuropathy[J].Journal of Audiology and Speech Pathology,2007(2):89-97.
3王秋菊.听神经病的临床及基础研究[J].国际耳鼻咽喉头颈外科杂志,2006(06):394-397.Wang QJ.Clinical and basic research of auditory neuropathy[J].Int J Otolaryngol Head Neck Surg,2006,30(06):394-397.
4 Varga R,Kelley P M,Keats B J,et al.Non-syndromic recessive auditory neuropathy is the result of mutations in the otoferlin(OTOF)gene[J].J Med Genet,2003,40(1):45-50.
5王锦玲,高磊,薛飞,等.听神经病的临床与听功能特征[J].临床耳鼻咽喉科杂志,2002(10):518-520.Wang JL,Gao L,Xue F,et al.Auditory neuropathy[J].Journal of Clinical Otorhinolaryngology Head and Neck Surgery,2002(10):518-520.
6 Houseman M J,Jackson A P,Al-Gazali L I,et al.A novel mutation in a family with non-syndromic sensorineural hearing loss that disrupts the newly characterised OTOF long isoforms[J].JMed Genet,2001,38(8):E25.
7 Wang J,Fan Y Y,Wang S J,et al.Variants of OTOF and PJVKgenes in Chinese patients with auditory neuropathy spectrum disorder[J].PLoS One,2011,6(9):e24000.
8 Romanos J,Kimura L,Favero M L,et al.Novel OTOF mutations in Brazilian patients with auditory neuropathy[J].J Hum Genet,2009,54(7):382-385.
9 Wang D Y,Wang Y C,Weil D,et al.Screening mutations of OTOF gene in Chinese patients with auditory neuropathy,including a familial case of temperature-sensitive auditory neuropathy[J].BMC Med Genet,2010,11:79.
10 Chiu Y H,Wu C C,Lu Y C,et al.Mutations in the OTOF gene in Taiwanese patients with auditory neuropathy[J].Audiol Neurootol,2010,15(6):364-374.
11 Varga R,Avenarius M R,Kelley P M,et al.OTOF mutations revealed by genetic analysis of hearing loss families including a potential temperature sensitive auditory neuropathy allele[J].JMed Genet,2006,43(7):576-581.
12 Yasunaga S,Grati M,Cohen-Salmon M,et al.A mutation in OTOF,encoding otoferlin,a FER-1-like protein,causes DFNB9,a nonsyndromic form of deafness[J].Nat Genet,1999,21(4):363-369.
13 Roux I,Safieddine S,Nouvian R,et al.Otoferlin,defective in a human deafness form,is essential for exocytosis at the auditory ribbon synapse[J].Cell,2006,127(2):277-289.
14齐悦,张秋静,王大勇,等.听神经病谱系障碍患者SLC17A8基因筛查研究[J].中华耳科学杂志,2015(02):224-228.Qi Y,Zhang QJ,Wang DY,et al.Mutation screening of SLC17A8 gene in patients with auditory neuropathy spectrum disorder[J].Chinese Journal of Otology.2015(02):224-228.
15 Wang Q J,Li Q Z,Rao S Q,et al.AUNX1,a novel locus responsible for X linked recessive auditory and peripheral neuropathy,maps to Xq23-27.3[J].J Med Genet,2006,43(7):e33.
16 Zong L,Guan J,Ealy M,et al.Mutations in apoptosis-inducing factor cause X-linked recessive auditory neuropathy spectrum disorder[J].J Med Genet,2015,52(8):523-531.