CX26基因在非综合征型耳聋中的产前诊断及早期干预

目的：对非综合征型耳聋家系进行CX26基因的突变检测,对检测出CX26基因突变的家系进行产前诊断并实施早期干预。方法：对来自国内十多个省份的100个非综合征型耳聋家系中的先证者通过聚合酶链反应、单链构像多态性分析以及直接测序法进行CX26基因的突变检测,对确诊为CX26基因所致的遗传性耳聋家系中的一成员在妊娠时通过脐静脉穿刺术抽取脐胎血进行产前诊断及早期干预。结果：①发现CX26基因的致病性突变1种：cDNA编码区233～235位点c的纯合缺失;多态6种：G79A、G109A、A341G、G442A、G506A和T608C;②对一个确诊为CX26基因233~235delC的遗传性聋家系的成员于第2次妊娠时进行产前诊断,发现胎儿具有同种致病性突变。结论：①CX26基因cDNA编码区233～235位点C的杂合缺失不致聋,纯合缺失可导致非综合征型常染色体隐性遗传性聋;②产前诊断和早期干预可预防遗传性聋。这是我国首次确诊携带耳聋致病基因的胎儿并实施早期干预。     

Connexin26 mutations associated with nonsyndromic hearing loss

OBJECTIVE: Mutations in the GJB2 gene are a major cause of autosomal recessive and sporadic types of congenital deafness. The 35delG mutation is the most frequent type of mutation in white populations. However, several other forms were reported, such as 167delT among Ashkenazi Jews and R143W in Africans. The present study investigated the mutations of connexin26 (Cx26) found in patients with nonsyndromic hearing loss (NSHL) and newborns in the Korean population. STUDY DESIGN: The sequencing data for 147 unrelated patients with congenital NSHL and 100 audiologically screened newborns were included in this prospective study. METHODS: Genomic DNA samples from all patients and newborns were sequenced in both directions for detection of Cx26 mutations. RESULTS: Thirteen different types of mutations were found in the patients and newborns. V27I and E114G are the popular types of polymorphic mutations in both groups. 235delC-deletion and frameshift--was detected in patients (15 in 294 alleles) and newborns (1 in 200 alleles). 35delG was rarely found in both group. In addition to above mutations, several types of mutations--S85P, K41R, S72C, V84A, 176-191del, and 299-300del-were identified. The family study of the 235delC showed a typical autosomal recessive trait of NSHL in their audiological evaluation of hearing threshold. CONCLUSION: The frequency of 235delC allele showed much higher in the patients (5%) than in newborns (0.5%). We rarely found 35delC mutant in both groups. These results suggest that the different types of Cx26 mutations affect autosomal recessive NSHL according to ethnic background.     

Identification of novel variants in the Myosin VIIA gene of patients with nonsyndromic hearing loss from Taiwan

Objective

To determine whether variants of exons 7, 11, 22 and 28 of the MYO7A gene are causes of nonsyndromic deafness in Taiwanese.

Methods

We screened a total of 331 unrelated Taiwanese individuals (age range, 4-22 years), including 231 patients with severe to profound nonsyndromic hearing loss and 100 individuals with normal hearing. Genomic DNA was extracted from peripheral blood leukocytes and then subjected to PCR to amplify selected exons and flanking introns of the MYO7A gene; the amplified products were screened for base mutations by autosequence. Data from the two groups were then compared using the chi-square (χ²) test.

Results

The analysis revealed six variants in 3 out of 4 screened exons and flanking intronic sequences of the MYO7A gene (exons 7, 11, and 22). Three missense variants were found only in patients with hearing loss and were heterozygous, including Arg206Cys, Arg206His and Thr381Met. A variant, c.IVS22+58G>A, was found in intron 22 of the MYO7A gene from both patients and control group. Allele frequencies of c.IVS22+58G>A were shown to be significant between the two groups using χ² test (P < 0.05).

Conclusion

Our results indicate that Arg206 and Thr381 residues in the motor head region of MYO7A protein are critical sites and the mutations of these residues may lead to the development of nonsyndromic deafness.     

Identification of novel variants in the TMIE gene of patients with nonsyndromic hearing loss

Objective

To determine whether variants of the TMIE gene are causes of nonsyndromic deafness in Taiwan.

Methods

A genetic survey was made from 370 individuals, with 250 nonsyndromic hearing loss and 120 normal hearing individuals. Genomic DNA was extracted from peripheral blood leukocytes and then subjected to PCR to amplify selected exons and flanking introns of the TMIE gene; the amplified products were screened for base variants by autosequence. Data from the two groups were then compared using Fisher's two-tailed exact test and Armitage's trend test.

Results

The analysis revealed 7 novel variants in the TMIE gene. Of the 7 variants, 5 variants were found in both nonsyndromic hearing loss and normal hearing group. Both allelic and genotype frequencies of these sequence changes did not differ significantly between patients and controls (P > 0.05). However, a missense variant (c.257G > A) and one promoter variant (g.1-219A > T) were found in two patients with nonsyndromic hearing loss. Family study and microsatellite analysis found that c.257G > A variant is not inherited from his parents. The c.257G > A variant encodes a protein with glutamine at position 86 instead of arginine (p.R86Q), a residue that is conserved in mammals but different in fish, and predicted to be extracellular.

Conclusions

Despite the fact that the frequency of TMIE variants in our study subjects was low, we suggested that c.257G > A (p.R86Q) variant is a de novo and may be as a risk factor for the development of hearing loss in Taiwanese.     

Connexin 26 and connexin 30 mutations in children with nonsyndromic hearing loss

OBJECTIVES/HYPOTHESIS: Mutations in the connexin 26 (Cx26) or gap junction beta 2 gene are the leading cause of hereditary nonsyndromic sensorineural hearing loss in Caucasians. The Cx26 coding region of 68 children with nonsyndromic sensorineural hearing loss was sequenced to determine the frequency and type of Cx26 mutations in this population. Screening was also performed for a common connexin 30 (Cx30) or gap junction beta 6 mutation (del [GJB6-D13S1830]). Children also underwent audiological testing to determine whether any correlation exists between Cx26 mutations and severity of hearing loss. STUDY DESIGN: In all, 68 children with nonsyndromic sensorineural hearing loss were screened for Cx26 and Cx30 mutations by polymerase chain reaction and direct sequencing. METHODS: Genomic DNA was amplified by polymerase chain reaction using primers that flank the entire Cx26 coding region. Screening for the 342-kb Cx30 deletion was performed using primers that amplified the breakpoint junction of the deletion. The amplicons were then sequenced in both directions and analyzed for mutations. Audiometric testing, including pure-tone audiometry and auditory evoked brainstem response, was also performed to determine the degree of hearing loss. RESULTS: Twenty-seven of 68 children tested had mutations in Cx26 with 35delG being the most prevalent. Ten additional Cx26 mutations were detected including a novel compound heterozygote. Two children were heterozygous for the Cx30 del (GJB6-D13S1830) mutation. CONCLUSION: Cx26 and Cx30 mutations were present in 41.2% of children tested in the study population. Audiometric data supported previous studies demonstrating a greater degree of hearing loss in subjects who are homozygous for the 35delG mutation.     

Hearing loss in enlarged vestibular aqueduct and incomplete partition type II

Purpose

The purpose of this work is to identify the role of incomplete partition type II on hearing loss among patients with enlarged vestibular aqueduct (EVA).

中国人非综合征型学语前聋患者Taperin基因突变筛查研究

目的：应用耳聋基因芯片联合DNA测序法对134例非综合征型聋患者进行静纤毛表达的Taperin基因的突变检测,分析该基因在中国人遗传性聋患者中的突变率及类型特点。方法：采集134例非综合征型聋患者和100例听力正常者的外周血,提取基因组DNA。用遗传性耳聋基因芯片排除常见的4个致聋基因突变患者,对未携带或仅带有单个杂合GJB2或SLC26A4突变的患者应用PCR—DNA测序法对Taperin基因序列进行测定,分析有无突变。结果：在134例非综合征型聋患者组中,基因芯片方法排除出19个患者携带常见致聋基因突变;剩下115例患者应用DNA测序法对Taperin基因进一步检测,结果在2例患者中发现A187S杂合突变。经同源性分析,A187S发生在保守的氨基酸残基。在家族成员中也检出携带了上述突变的杂合子。此外,在患者和对照组中发现2种多态157C〉T和318C〉T。结论：在中国人非综合征聋患者中发现Taperin基因1种新突变A187S,可能与耳聋有关。还在中国人中发现Tapetin基因的2种多态157C〉T和318C〉T。在中国人非综合征型耳聋患者中Taperin基因的突变携带率约为1．74％。     

新疆维吾尔族非综合征型遗传性聋患者线粒体DNA 12SrRNA A1555G突变分析

目的:调查新疆地区维吾尔族非综合征型遗传性聋患者的线粒体DNA 12SrRNA A1555G突变情况,为预防氨基苷类抗生素致聋提供依据。方法:收集新疆地区51例维吾尔族非综合征型遗传性聋患者,53例维吾尔族听力正常者作为对照组。抽取外周静脉血,从白细胞中提取DNA,PCR扩增线粒体DNA目的片断,Alw26I限制性内切酶检测A1555G点突变,而后对阳性患者的PCR产物进行DNA测序验证。结果:在所有样本中,2例存在线粒体DNA A1555G点突变,均为维吾尔族非综合征型遗传性聋患者,且均有明确氨基苷类抗生素用药史。结论:新疆地区维吾尔族耳聋患者及维吾尔族正常人线粒体DNA A1555G检出率比较差异无统计学意义。携带有该突变的个体对氨基苷类抗生素的耳毒作用有高度易感性。新疆地区聋哑患者的A1555G突变检出率低于全国平均水平。     

Phenotype of DFNA11: a nonsyndromic hearing loss caused by a myosin VIIA mutation

OBJECTIVES/HYPOTHESIS: To characterize the audiovestibular phenotype of DFNA11, an autosomal dominant nonsyndromic hearing impairment caused by a mutation in the myosin VIIA gene (MYO7A), including whether DFNA11-affected subjects have retinal degeneration as is characteristic of Usher syndrome type 1B, caused by different MYO7A mutations. STUDY DESIGN: Retrospective study of audiovestibular and ophthalmological data in a Japanese family linked to DFNA11. METHODS: Otoscopic examination and pure-tone audiometry were performed in all participants in the family. Selected subjects underwent additional examinations including speech discrimination scoring, acoustic reflex measurements, Békésy audiometry, evoked and distortion-product otoacoustic emissions, auditory brainstem responses, and bithermal caloric testing; visual acuity, ocular tonometry, slit-lamp examination, ophthalmoscopy, and electroretinography; and computed tomography of the temporal bone. RESULTS: Most affected individuals had moderate cochlear hearing loss beginning in the second decade and progressing at all frequencies. Variable degrees of asymptomatic vestibular dysfunction were present. Computed tomography showed normal inner and middle ear structures. No evidence suggested retinitis pigmentosa. CONCLUSIONS: The phenotype of DFNA11 is postlingual, nonsyndromic sensorineural hearing loss with gradual progression. Showing moderate hearing loss with asymptomatic variable vestibular dysfunction and no retinal degeneration, the DFNA11 phenotype is mildest among phenotypes caused by MYO7A mutations.     

新疆哈萨克族非综合征型聋GJB2基因突变的研究分析

目的：研究新疆哈萨克族非综合征型聋患者GJB2基因突变的情况。方法：调查对象为来自新疆地区的193例哈萨克族患者,采用直接测序法对非综合征型聋患者97例和健康对照96例进行GJB2基因突变的检测。结果：在编码区耳聋组共发现8种碱基改变：其中35delG纯和12例,79G〉A纯合5例,79G〉A杂合8例,79G〉A与608T〉C复合杂合1例,79G〉A与341A〉G复合杂合5例,235delC杂合4例,341A〉G杂合2例,439T〉G杂合1例,457G〉A杂合1例,521G〉A纯合2例。对照组发现4种已明确的常见多态性碱基改变。结论：本研究提示新疆哈萨克族非综合征型聋患者GJB2基因突变具有种族和地域性特点,该地区哈萨克族耳聋人群中GJB2有较高携带率,在本研究中35 delG为其常见突变方式。     

Spectrum of GJB2 (Cx26) gene mutations in Iranian Azeri patients with nonsyndromic autosomal recessive hearing loss

Objective

Hereditary hearing impairment is a genetically heterogeneous disorder. In spite of this, mutations in the GJB2 gene, encoding connexin 26 (Cx26), are a major cause of nonsyndromic recessive hearing loss in many countries and are largely dependent on ethnic groups. The purpose of our study was to characterize the type and prevalence of GJB2 mutations among Azeri population of Iran.

Methods

Fifty families presenting autosomal recessive nonsyndromic hearing loss from Ardabil province of Iran were studied for mutations in GJB2 gene. All DNA samples were screened for c.35delG mutation by ARMS PCR. Samples from patients who were normal for c.35delG were analyzed for the other variations in GJB2 by direct sequencing. In the absence of mutation detection, GJB6 was screened for the del(GJB6-D13S1830) and del(GJB6-D13S1854).

Result

Thirteen families demonstrated alteration in the Cx26 (26%). The 35delG mutation was the most common one, accounting for 69.2% (9 out of 13 families). All the detected families were homozygous for this mutation. Two families were homozygous for delE120 and 299-300delAT mutations. We also identified a novel mutation: c.463-464 delTA in 2 families resulting in a frame shift mutation.

Conclusion

Our results suggest that c.35delG mutation in the GJB2 gene is the most important cause of GJB2 related deafness in Iranian Azeri population.     

Autosomal recessive non-syndromic hearing loss is caused by novel compound heterozygous mutations in TMC1 from a Tibetan Chinese family

ObjectivesHearing loss is the most common sensory disorder worldwide. Biallelic mutations in 42 different genes have been identified as associated with autosomal recessive non-syndromic hearing loss (ARNSHL). One of the common genes responsible for ARNSHL is TMC1. TMC1 mutations have been reported to cause non-syndromic hearing loss in a variety of populations. The current study is designed to investigate mutations prevalent among Chinese ethnic groups with ARNSHL.MethodsTargeted exome sequencing (TES) was employed to study the genetic causes of two siblings with ARNSHL in a Tibetan Chinese family. Variants identified by TES were further confirmed by Sanger sequencing.ResultsWe identified two distinct variants in the TMC1 gene in two deaf siblings of one Tibetan Chinese family using TES. Both siblings inherited a paternal allele containing a deletion of c.1396_1398AAC (p.Asn466del) and a maternal allele containing an insertion of c.2210_2211insCT (p.Glu737HisfsX2). The former disrupts a highly conserved residue in the large intracellular loop domain adjacent to the fourth transmembrane domain, and the latter causes a truncation of a portion of the C-terminal domain. These variants were compound heterzygous and segregated with the hearing impairment in this family.ConclusionThe novel compound heterozygous mutant alleles of TMC1 identified in this study were responsible for the ARNSHL in this Tibetan Chinese family. Although compound heterozygous mutations in TMC1 occurring in different TMC1 domains have been previously described in Han Chinese; this result suggests that the TMC1 variants contributing to hereditary deafness in Chinese populations may be more complex than initially assumed and that sequence-based diagnostics will be required for a comprehensive evaluation of ARNSHL.     

A mutation in Wolfram syndrome type 1 gene in a Japanese family with autosomal dominant low-frequency sensorineural hearing loss

Conclusion. Our findings suggest that Wolfram syndrome type 1 gene (WFS1) mutation is an important cause of autosomal dominant low-frequency sensorineural hearing loss (LFSNHL) in Japan. Objective. DFNA6/14 is caused by a heterozygous mutation of WFS1 and is a common cause of autosomal dominant LFSNHL among populations in both Europe and the US. The purpose of this study was to investigate WFS1 mutations among Japanese patients whose phenotypes were consistent with those of DFNA6/14. Material and methods. Using audiometry and genetic analysis, we searched for WFS1 mutations in three unrelated Japanese patients with LFSNHL and a familial history of autosomal dominant hearing loss. Results. One patient carried a heterozygous G2700A mutation at codon 844 in exon 8, resulting in substitution of a threonine for an alanine (A844T). Genetic analysis of the available members of the patient's family showed that the A844T mutation segregated with LFSNHL, but was not detected in any of 140 control chromosomes. It thus appears likely that the A844T mutation is causative for hearing loss in this group. Speech audiometry, self-recording audiometry and auditory brainstem responses showed the patient to have cochlear deafness without retrocochlear dysfunction. No mutation was found in the other two patients.     

A novel missense mutation in the Connexin 26 gene associated with autosomal recessive nonsyndromic sensorineural hearing loss in a consanguineous Tunisian family

Nonsyndromic sensorineural hearing impairment is inherited in a predominantly autosomal recessive manner in up to 70% of cases. The gene more often involved is GJB2, encoding the gap junction protein Connexin 26. We report here a novel missense mutation in the GJB2 gene found in a Tunisian family. A homozygous change C/G at nucleotide 263 was detected in the 4-year-old girl of this family, affected by congenital moderate hearing loss. This transversion leads to the replacement of a highly conserved alanine with glycine at codon 88 (A88G). The consanguineous parents of the child are healthy carriers of the mutation.     

特发性突聋患者GJB2 235delC突变分析

目的：分析GJB2 235delC突变在特发性突聋患者中的发生频率,探寻其与特发性突聋发病的可能相关性。方法：收集234例特发性突聋患者,以同期听力正常的80例体检人员为对照组。分别采外周静脉血,提取基因组DNA,经聚合酶链反应扩增GJB2基因编码区后,用限制性内切酶酶切的方法筛查235delC位点突变,同时对特发性突聋患者的临床资料汇总分析。结果：234例特发性突聋患者中,5例检测出GJB2 235delC杂合突变,突变检出率为2.1%,未检测出235delC纯合突变;对照组中未检出GJB2 235delC突变。2组人群235delC突变检出率差异无统计学意义（P〉0.05）。结论：特发性突聋患者中GJB2 235delC突变检出率低,提示该突变位点可能与特发性突聋的发病无相关性。     

扬州市特教学校耳聋学生常见耳聋突变基因调查报告

目的:调查扬州地区非综合征性聋患儿常见耳聋突变基因的发病情况。方法:选择扬州市特教学校90例中、重度非综合征性聋学生为研究对象。在苏北人民医院医学检测中心利用耳聋基因芯片诊断试剂盒筛查常见的耳聋相关基因的9个热点突变,包括GJB2(35delG、176del16、235delC及299delAT),GJB3(538C>T),SLC26A4(IVS7-2A>G、2168A>G)和mtDNA 12SrRNA(A>G、1494C>T)。结果:在90例耳聋患者中,基因芯片方法共检出携带致聋基因突变64例(71.1%)。其中,GJB2基因突变40例(44.4%),包括235delC纯合突变20例(22.2%),235delC单杂合突变4例(4.4%),235delC和299delAT复合杂合突变2例(2.2%);299de-lAT单杂合突变2例(2.2%),299delAT纯合突变2例(2.2%);176del16单杂合突变2例(2.2%),176del16纯合突变2例(2.2%),176del16和235delC复合杂合突变6例(6.7%)。SLC26A4基因突变22例(24.4%),包括IVS7-2A>G纯合突变2例,IVS7-2A>G和2168A>G复合杂合突变2例(2.2%),IVS7-2A>G单杂合突变18例(20.0%);mtDNA 12SrRNA A>G纯合突变2例(2.2%);未检出GJB3基因突变。结论:应用基因诊断技术可以在耳聋患者病因调查中进行快速筛查诊断,值得推广应用。     

Prevalence of GJB6 mutations in Chinese patients with non-syndromic hearing loss

Objective

To investigate the distribution of GJB6 mutations in Central Chinese population with non-syndromic hearing loss.

Method

Totally 655 hearing impaired patients in Hubei province of China were screened for del(GJB6-D13S1830) deletions by using multiplex PCR and sequencing of GJB6 whole coding region.

Result

The del(GJB6-D13S1830) and other mutations in GJB6 gene were not observed in our study cohort.

Conclusion

The results suggest that GJB6 mutations is not a common cause among Central Chinese population and screening for the mutations of GJB6 can be ranked as unconventional deaf gene test for this population.