Mutations in the connexin 26 gene in patients with nonsyndromic hearing impairment]

OBJECTIVE: To determine the prevalence and characteristics of deafness-causing mutations in Connexin 26(Cx26, GJB2) gene in Chinese with nonsyndromic hearing impairment(NSHI). METHODS: Study subjects are all Chinese including 16 infants with sporadic congenital deaf-mutism, 39 patients with autosomal recessive hereditary hearing loss, 30 patients with autosomal dominant hereditary hearing loss and 100 normal adults. The subjects were screened for base variations by single-strand conformational polymorphism (SSCP) analysis of the amplified products of polymerase chain reaction (PCR). Those who were found have abnormal conformational band were sequenced. RESULTS: Five kinds of polymorphism were found in 15 cases of controls and six kinds of polymorphism in 10 patients. No mutation was found in Cx26 gene in Chinese with autosomal recessive NSHI. Heterozygous deletion AT at position 299-300 of Cx26 cDNA, which results in premature chain termination, was found in a pedigree with autosomal dominant hereditary nonsyndromic hearing loss. CONCLUSION: The prevalence of deafness-causing mutations in Cx26 gene in Chinese with autosomal recessive NSHI maybe is lower than that of other ethnic groups. Heterozygous deletion AT at position 299-300 of Cx26 cDNA can lead to autosomal dominant hereditary hearing loss (DFNA3).     

非综合征性耳聋患者连接蛋白26基因突变的研究

目的　探讨中国人非综合征性耳聋患者连接蛋白 2 6 (connexin 2 6 ,Cx2 6 )基因突变频率和特性。方法　收集中国散发先天性聋哑儿童 16例 ,常染色体隐性遗传性聋 39例 (39个家系 ) ,10岁前开始听力下降的常染色体显性遗传性聋 30例 (30个家系 )和健康对照组 10 0例。聚合酶链反应 单链构象多态 (singlestrandconformationalpolymorphismanalysisofpolymerasechainreaction ,PCR SSCP)分析初筛可疑突变者 ,SSCP分析发现异常构象带后再行DNA测序。结果　健康对照组中 15例发现 5种多态性改变 ,耳聋患者中 10例发现 6种多态性改变。散发先天性聋哑和常染色体隐性遗传非综合征性耳聋中未发现致病突变。 1个常染色体显性遗传性聋家系发现所有患者 (3例 )Cx2 6基因的编码区2 99 30 0位碱基AT杂合性缺失 ,导致移码突变 ,翻译的蛋白质截短 ,该家系听力正常者无此突变。结论　蒙古人种中常染色体隐性遗传性非综合征性耳聋的Cx2 6基因突变率可能低于其他人种。Cx2 6基因编码区 2 99 30 0位碱基AT杂合性缺失可致常染色体显性遗传性聋DFNA3型     

Study on mutations in the connexin 26 gene among Chinese with nonsyndromic hearing loss]

OBJECTIVE: To study the relation between nonsyndromic hearing loss in Chinese and mutations in connexin 26 (Cx 26) gene and to explore the pathogenic mechanism. METHODS: One hundred and thirty-eight individuals from thirty-five pedigrees with nonsyndromic hearing loss, 99 children with sporadic nonsyndromic hearing loss and 100 normal adults as control were collected in present studies. The Cx 26 coding sequence was screened by single strand conformational polymorphism (SSCP) and analyzed by direct sequencing when SSCP shifts were observed. RESULTS: Five SSCP shifts in 2 pedigrees were observed. Homozygous deletion C at position 233-235 of Cx 26 cDNA, which resulted in frameshift mutation, was found in 2 pedigrees with nonsyndromic hearing loss. CONCLUSION: The hot-spot mutations of Cx 26 gene in Chinese with nonsyndromic hearing loss may be different from other ethnic groups. The 233-235 delC homozygous mutation of Cx 26 cDNA can result in autosomal recessive nonsyndromic hearing loss in Chinese population.     

connexin 26基因突变与国人遗传性无综合征耳聋相关性分析

目的分析国人遗传性无综合征耳聋与缝隙连接蛋白26（connexin 26,Cx 26）基因突变相关性,从分子水平探讨该病的发病机理。方法收集国人35个无综合征耳聋家系中138名成员,99例散发的无综合征耳聋患者以及100份健康对照个体的外周血DNA样本共337份;采用聚合酶链反应-单链构像多态性（polymerase chain reaction-single stand conformational     

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。     

Lack of association between Connexin 31 (GJB3) alterations and sensorineural deafness in Austria

Mutations in the gap junction protein beta 3 (GJB3) gene encoding Connexin 31 (Cx31) are known to cause autosomal inherited sensorineural deafness, erythrokeratodermia and neuropathy. The role of Cx31 mutations has not been described in familial cases of non-syndromic hearing impairment (NSHI) in central European populations. To identify mutations in the Austrian population, highly selected familial (n=24) and sporadic (n=21) cases of isolated NSHI were screened by analysis of the complete coding sequence of Cx31, after exclusion of a common Cx26 causing deafness. Three different variations occurring in a total of 37% of all cases were identified. A C94T (R32W) missense mutation was seen in 4.4% of cases and two silent alterations C357T and C798T were detected in 8.9% and 24.4% of cases exclusively in a heterozygous pattern. No correlation between Cx31 alterations and deafness was found. To investigate the role of heterozygous Cx31 variations for a possibly combination allelic disease inheritance with Cx26 mutations as shown for Connexin 30 and Connexin 26, patients with Cx26 variations were tested. Our data suggest that Cx31 alterations are common but have no or a low genetic relevance in the Austrian hearing impaired population with or without Cx26 alterations.     

中国人非综合征型听力损失患者Cx26基因的突变分析

目的 分析中国人遗传性非综合征型听力损失（nonsyndromic hearing impairmetn,NSHI)患者缝隙连接蛋白（connexin 26,Cx26)基因编码区的突变。方法 对天津市聋哑学校的8个聋哑学生的家系中29例以及健康对照2例和有家族史但本人听力正常的遗传咨询者2例共33例取外周血提取DNA，经聚合酶链反应（olymerase chain reaction,PCR)扩增Cx26基因编码区片段，通过限制酶切指纹－单链构像多态性（restriction endonucleases fingerprinting-single strand conformation polymorphism,REF-SSCP)分析法进行突变筛选，经DNA测序判断多态性改变或致病突变。结果 33例中有30例Cx26基因发生改变，改变率为90．91％（30／33），共发现8种不同形式的改变，包括79G→A，109G→A，161A→T，235delC,240G→A,341A→G,571T→Ct 608T→C,其中161A→t,240G→A和571T→C为新发现的突变。22例耳聋患者中有3例为235delC，突变率为13．64％（3／22）。结论 Cx26基因235delC是中国人NSHI患者中主要的突变方式，NSHI患者中存在较多的多态性改变。     

CX31.1基因在遗传性聋家系中的突变分析

目的：通过对非综合征型聋家系进行CX31．1基因的突变分析,以鉴定CX31．1基因是否为遗传性聋的致病基因。方法：通过对从全国10多个省收集到的遗传性聋家系61个,其中常染色体隐性非综合征型聋家系37个,常染色体显性非综合征型聋家系24个的106例成员及50例正常人进行聚合酶链反应(polymerasc chain reaction,PCR)及直接测序,对遗传性聋个体进行CX31．1基因的突变检测。结果：发现CX31．1的同义突变1种,多态1种,内含子缺失2种。结论：我们目前的检测虽未能证明CX31．1基冈突变是上述聋家系的致病基因,但根据基因结构分析该基因与遗传性聋的关系不能排除,有待收集更多的家系做进一步的研究。     

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.     

安阳市特教学校重度感音性聋分子病因学分析--GJB2 235delC突变和线粒体DNA 12SrRNA A1555G突变筛查报告

目的 调查河南省安阳地区重度感音神经性耳聋聋病分子病因学情况。方法对安阳市聋哑学校160名学生进行耳聋病因问卷调查、纯音听阈测试。对其中154名非综合征性感音神经性耳聋患者进行线粒体DNA 12SrDNA A1555G点突变检测和G朋12基因突变检测。结果 8例（5．19％）存在线粒体DNA 12SrDNA A1555G点突变；11例（7．14％）存在GJB2 235delC纯合突变；13例（8．44％）存在GJB2 235delC杂合突变。在分子水平能够明确诊断者占20．77％。结论 安阳地区耳聋患者存在较高的遗传性耳聋发生率。特别是线粒体DNA A1555G突变发生率高于全国平均水平，通过聋病分子诊断，可达到防聋、指导聋儿康复及评估耳聋预后等积极效果。