一个同时携带线粒体DNA A1555G突变和GJB2235delC单杂合突变家系的基因型与听力表型

目的 调查一个同时携带线粒体DNA A1555G突变和GJB2 235delC突变的非综合征型耳聋家系,分析其基因型和听力表型的关系.方法 对家系成员进行临床听力测试,收集家系中8名成员的外周静脉血样本,从白细胞中提取DNA,聚合酶链反应扩增GJB2基因和线粒体DNA(mitochondric DNA,mtDNA)目的 片段,对扩增片段直接测序进行GJB2基因、mtDNA 12S rRNA及tRNASer(UCN)基因突变分析.结果 此家系先证者存在mtDNA A1555G突变和GJB2 235delC杂合突变,听力表型为极重度感音神经性耳聋.其他母系成员携带mtDNA A1555G突变,未发现tRNASer(UCN)基因突变,家系中其他母系成员听力表型为双侧对称高频下降或听力正常.结论 GJB2 235delC单杂合突变可能参与了mtDNA A1555G的听力损害.     

一个同时携带线粒体DNA A1555G突变和GJB2235delC单杂合突变家系的基因型与听力表型

目的 调查一个同时携带线粒体DNA A1555G突变和GJB2 235delC突变的非综合征型耳聋家系,分析其基因型和听力表型的关系.方法 对家系成员进行临床听力测试,收集家系中8名成员的外周静脉血样本,从白细胞中提取DNA,聚合酶链反应扩增GJB2基因和线粒体DNA(mitochondric DNA,mtDNA)目的 片段,对扩增片段直接测序进行GJB2基因、mtDNA 12S rRNA及tRNASer(UCN)基因突变分析.结果 此家系先证者存在mtDNA A1555G突变和GJB2 235delC杂合突变,听力表型为极重度感音神经性耳聋.其他母系成员携带mtDNA A1555G突变,未发现tRNASer(UCN)基因突变,家系中其他母系成员听力表型为双侧对称高频下降或听力正常.结论 GJB2 235delC单杂合突变可能参与了mtDNA A1555G的听力损害.     

线粒体DNA A1555G突变在新生儿大规模筛查的临床意义

目的 探讨在新生儿中进行线粒体DNA(mitochondria DNA,mtDNA)A1555G突变基因大规模筛查在预防药物性耳聋的必要性.方法 随机取2008年在深圳市出生的1000名新生儿的血滤纸标本,用Chelex-100树脂提取DNA,PCR扩增,变性高效液相色谱法(denaturing hig-performance liquid chromatography,DHPLC)进行mtDNA A1555G突变基因筛查,计算出阳性突变频率.结果 1000名新生儿血滤纸样本中,共检测出2例样本存在mtDNA A1555G突变,突变率为0.2%.结论 mtDNA A1555G突变在新生儿中出现的频率较高,对其进行mtDNA A1555G突变大规模筛查发现氨基甙类抗生素敏感个体,能有效地对新生儿及其家族高危人群进行合理性指导用药,从而更好地预防药物性耳聋.     

线粒体DNA A1555G突变的异质性研究进展

线粒体DNA 12S rRNA基因A1555G突变是最早发现的、最常见的与非综合征性耳聋相关的人类线粒体DNA突变。线粒体DNA A1555G突变大多以均质性突变的形式出现,但近来有报道指在不同的家系中也存在异质性的突变形式,且突变负荷与耳聋程度相关。鉴于线粒体DNA A1555G的临床表型在不同个体间存在显著差异,该突变异质性的存在为解释其表型多样性提供了依据。     

线粒体肌病患者线粒体DNA的突变分析

目的分析线粒体肌病患者线粒体DNA的突变情况，为疾病诊断提供依据。方法用常规HE、酶组化染色和电镜检查等病理形态学方法对3例线粒体肌病疑似患者进行诊断，并用聚合酶链反应-单链构象多态和DNA测序等方法对患者线粒体DNA中全部22个tRNA基因进行突变筛查。结果3例患者均被确诊为线粒体肌病，其中例1tRNA—VaI基因发生A1627G纯合突变，例2tRNA—Val基因发生A1627G／A杂合突变，例3tRNA—Trp基因发生T5554C突变、tRNA—Arg基因发生A10412C／A杂合突变。结论线粒体DNA中的tRNA基因突变是线粒体肌病的重要病因之一。     

120例非综合征性耳聋患者线粒体A1555G和Cl494T突变分析

目的分析温州地区120例耳聋患者的致聋原因,并探讨线粒体ONA（mitochondrial DNA,mtDNA）12S rRNA基因A1555G和C1494T突变与耳聋之间的关系。方法对我院收集的120例耳聋患者进行分子流行病学的调查,并针对线粒体1555和1494位点进行引物设计,通过PCR扩增,产物Sanger测序后比对标准序列,检测A1555G和C1494T突变的频率以及和患者使用氨基糖苷类抗生素的相关性。结果在120例重度耳聋患者中具有氨基糖苷类抗生素用药史的有66例（55％）,家族性遗传耳聋有22例（18．3％）,近亲结婚可能致聋有10例（8．3％）,不明原因的耳聋有22例（18．3％）;我们还发现,有9例患者携带线粒体A1555G突变,突变的阳性率为7．5％,1例携带C1494T突变,阳性率为0．83％,这10例患者均有用药史。结论遗传因素和氨基糖苷类用药史是导致耳聋的重要原因,其中Al555G突变和C1494T突变是耳聋较为常见的线粒体DNA突变,这对早期诊断和预防药物性耳聋具有一定的临床意义。     

7个非综合征型耳聋家系患者的mtDNA A1555G突变分析

目的探讨非综合征型耳聋家系患者mtDNA A1555G突变及其临床特征。方法应用聚合酶链反应、限制性核酸内切酶酶切和DNA测序技术对7个非综合征型耳聋家系112个成员的mtDNA A1555G突变进行检测，并分析听力临床资料。结果7个家系中所有受检的母系成员mtDNA A1555G突变均为阳性，突变性质含同质性和异质性二种；非母系成员及配偶该突变为阴性。突变的性质与临床表型的有关。结论mtDNA A1555G突变可导致非综合征型耳聋和氨基糖苷类抗生素致聋，其突变性质含均质性和异质性两种，且与临床表型相关。     

长治地区耳聋患者常见耳聋基因GJB2、GJB3和线粒体DNA 12S rRNA 1555A〉G相关性分析

目的通过对长治地区115例耳聋患者常见基因GJB2、GJB3和线粒体DIqA 12S rRNA 1555A〉G测序分析,研究该地区耳聋基因的突变情况及热点突变位点。方法收集长治地区特殊教育学校的115q,J耳聋患者的外周血样本,提取其DblA后对它的目的基因进行扩增并测序。结果115例耳聋患者中GJB2基因检测到81例发生突变,并发现了10个突变位点,其中C．7657T〉c是主要的多态位点,其携带率为22．6％（52／115）。另外,长治地区发现2例线粒体DNAl555A〉G位点突变,未检测出GJB3基因突变位点。结论通过对长治地区常见耳聋基因突变位点的研究,了解该地区的耳聋基因突变谱,为后续国内耳聋基因型分布提供数据支持,同时也为耳聋的早期诊断,治疗提供理论依据。     

七个非综合征型耳聋家系患者的mtDNA A1555G突变性质与特点

目的 探讨非综合征型耳聋家系患者mtDNA A1555G突变性质及其特点,探索临床表型多样性的分子遗传学基础.方法 应用聚合酶链反应-限制性片段长度多态和实时荧光-扩增阻碍突变系统-定量PCR(real time-amplification refractory mutation system-quantitative PCR,RT-ARMS-qPCR)检测7个非综合征型耳聋家系71个成员的mtDNA A1555G突变,并收集、分析其临床资料.结果 7个家系中所有受检的母系成员mtDNA A1555G突变均为阳性,突变性质含同质性和异质性两种;非母系成员及配偶该突变为阴性.7个家系mtDNA A1555G同质性突变的拷贝数与耳聋轻重程度相关(R=0.341,P=0.022);mtDNA A1555G异质性突变的拷贝数与耳聋轻重程度相关(R=0.85,P=0.015).结论 mtDNA A1555G突变可导致非综合征型耳聋和氨基糖甙类抗生素致聋,其突变性质含同质性和异质性两种,且含mtDNA A1555G位点的突变型与野生型的比例与耳聋的严重程度密切相关.     

忻州地区耳聋患者常见耳聋基因GJB2、GJB3和线粒体DNA 12S rRNA 1555A〉G相关性分析

目的通过对忻州地区83例耳聋患者常见基因GJB2、GJB3和线粒体DNA 12S rRNA 1555A〉G测序分析,从分子水平研究该地区人群聋病的遗传病因和特点,为临床防聋治聋提供策略、依据。方法收集忻州地区83例耳聋患者外周血样本,提取DNA后对目的基因扩增并进行测序分析。结果83例耳聋患者中GJB2基因检测到57例发生突变,9个突变位点,与编码连接蛋白的非综合症耳聋突变数据库（http：／／davinci．crg．OS／deafneSS／index．php？Seccion=mut_db＆db=nonsynd）比对,8个位点已见报道,其中包括3个多态位点c．79G〉A、c．341A〉G、c．368C〉A和5个致病位点c．235delC、c．30-35delC、c．109G〉A、c．176-c．191dell6和c．299-c．300delAT,其中,c．79G〉A和c．341A〉G是主要突变方式,携带率为30．12％（42／174）和23．49％（39／166）。新发现1例未见报道的突变位点c．186C〉T;患者均未检测出GJB3和线粒体DNA12SrRNA1555A〉G基因突变位点。结论通过对忻州地区常见耳聋基因突变位点的研究,了解忻州地区该基因突变谱,为后续国内耳聋基因型分布提供数据支持,同时也为耳聋的早期诊断、治疗提供理论依据。     

Connexin 26 gene (GJB2) mutation modulates the severity of hearing loss associated with the 1555A→G mitochondrial mutation

We report a high prevalence of GJB2 heterozygous mutations in patients bearing the 1555A→G mitochondrial mutation, and describe a family in which potential interaction between GJB2 and a mitochondrial gene appears to be the cause of hearing impairment. Patients who are heterozygotes for the GJB2 mutant allele show hearing loss more severe than that seen in sibs lacking a mutant GJB2 allele, suggesting that heterozygous GJB2 mutations may synergistically cause hearing loss when in the presence of a 1555A→G mutation. The present findings indicate that GJB2 mutations may sometimes be an aggravating factor, in addition to aminoglycoside antibiotics, in the phenotypic expression of the non‐syndromic hearing loss associated with the 1555A→G mitochondrial mutation. © 2001 Wiley‐Liss, Inc.     

耳聋患者及正常人GJB2基因的突变筛查

目的 检测常染色体隐性遗传耳聋患者GJB2基因突变情况,并分析其与临床表型的关系.方法 收集42例耳聋患者的临床资料,对患者进行纯音电测听检查、声阻抗检测、脑干听觉诱发电位检查;应用聚合酶链反应和直接测序法,对患者和9例患者的父母以及105名正常对照进行GJB2基因检测.结果 两例患者具有235delC纯合性突变,其中1例系感音神经性耳聋,另1例系混合性耳聋;1对混合性耳聋的双生子患者同时携带176de116bp杂合性突变.109G→A、79G→A和341A→G的纯合及杂合突变在患者及正常对照中均有出现.结论 235delC纯合性突变为致病突变,该突变可出现在混合性耳聋中;双生子患者的176de116bp杂合性突变考虑为宫内受到外界环境影响所致,或者由其它基因突变所致.109G→A、79G→A和341A→G考虑为是该基因的多态性,其临床意义仍需进一步探索.     

Factors that affect hearing level in individuals with the mitochondrial 1555A>G mutation

The mitochondrial 1555A>G mutation is one of the most common mutations responsible for hearing loss in Asians. Although the association with aminoglycoside exposure is well known, there is great variation in the severity of hearing loss. We analyzed hearing levels in 221 Japanese individuals with this mutation and attempted to identify relevant covariants including (i) age, (ii) aminoglycoside exposure, (iii) heteroplasmy ratio, and (iv) other gene mutations. At every age, average hearing levels were worse than those in normal subjects, suggesting that mitochondrial function itself may affect the severity of hearing loss. Although the hearing loss in individuals with the 1555A>G mutation progressed with age, the rate did not differ from that of the normal subjects. Those who had reported aminoglycoside exposure had moderate-to-severe hearing impairment regardless of age, confirming that such exposure is the most important environmental variable. We also confirmed the presence of heteroplasmy, which is known to modify the expression of other mitochondrial diseases, but found no evidence for a significant correlation with hearing impairment. A high prevalence of GJB2 heterozygous mutations was noted, indicating that these mutations may exhibit epistatic interaction with the 1555A>G mutation.     

Level of heteroplasmy for the mitochondrial mutation A3243G correlates with age at onset of diabetes and deafness

The mitochondrial mutation A3243G has been shown to be associated with a syndrome of diabetes mellitus and sensorineural hearing loss. Using a solid-phase-based sequencing method we have investigated the relation between the proportion of mutant mitochondrial genomes and the time of disease onset among members of three families where the mutation segregates. A striking association was observed between the level of heteroplasmy and time of onset of disease, particularly hearing loss. Accordingly, this syndrome shares features of diseases caused by dynamic mutations in that variable transmission of the level of heteroplasmy between generations influences disease severity. Hum Mutat 12:52–58, 1998. © 1998 Wiley-Liss, Inc.     

遗传性非综合征型耳聋患者GJB2基因编码序列分析

目的 对6个遗传性非综合征型耳聋家系成员的GJB2基因编码序列进行分析,寻找耳聋患者的致病基因突变,探讨GJB2基因突变致病的遗传模式.方法 提取患者及家系成员的外周血基因组DNA,扩增GJB2基因的编码序列,然后对扩增产物进行DNA测序,对出现重叠峰形的扩增产物进行TA克隆后再测序,确定基因突变是否存在于同一拷贝.结果 6个遗传性非综合征型耳聋家系中,4个家系是GJB2基因突变所致.患者的GJB2基因突变包括235delC、299-300delAT、79G→A+341A→G和109G→A.非致聋突变79G→A与341A→G组合具有致聋效应,109G→A和235delC的杂合突变可能也有致聋效应.结论 GJB2基因突变致聋具有明显异质性,非致聋突变并非完全不致聋,环境因素或其它基因可能参与GJB2基因突变所致耳聋.     

Update of the Spectrum of GJB2 Mutations in 107 Patients with Nonsyndromic Hearing Loss in the Fujian Population of China

Mutations in the GJB2 gene, encoding connexin26, which is expressed in the inner ear, have been shown to be responsible for the majority of nonsyndromic hearing loss (NSHL) cases. To update and evaluate the spectrum and prevalence of GJB2 mutations in the Fujian population, we screened exon 2 (coding), exon 1, and flanking introns of GJB2 in 107 NSHL probands and 61 individuals with normal hearing. Twelve different variants were identified, including three pathogenic mutations (c.235delC, c.299_300delAT, and c.508insAACG), one hypomorphic allele (p.V37I), three polymorphic variants (p.V27I, p.E114G, and p.I230T), and five rare variants (p.N62N, p.F115C, p.T123N, p.G21E, and p.F142I). The p.G21E and p.F142I variants were potentially pathogenic as predicted by PolyPhen‐2, SIFT, and PROVEAN. The most common mutation was c.235delC with allele frequency 12.6% (27/214). The most common polymorphisms in the Fujian population were p.V27I and p.E114G, both detected at high frequency in probands and controls. The p.E114G variant was always in cis with p.V27I, and formed the haplotype, p.[V27I; E114G] in the Fujian population. Interestingly, only 17.76% (19/107) of NSHL probands had clearly defined pathogenic mutations in GJB2, indicating that the pathogenesis of NSHL in the Fujian population is heterogenous, and that further analysis of other NSHL genes is necessary.     

大同地区耳聋基因GJB2和mtDNA1555突变分析

目的通过筛查大同地区87例耳聋患者常见基因GJB2和mtDNA1555的突变频率,研究该地区CJB2和mtD-NA1555基因突变情况及热点突变位点。方法采集大同地区87例耳聋患者外周血,对目的基因扩增并进行测序分析。结果所有患者中有58例GJB2基因检测到11个突变位点,与编码连接蛋白的非综合征耳聋突变数据库（http：／／davinci．crg．es／deafness／index．php？section=mut_db＆db=nonsynd）比对,9个位点已见报道,其中包括5个多肽位点c．79G〉A、c．341A〉G、c．608T〉C、c．368C〉A、c．765T〉C和4个致病住点c．235delc、c．109G〉A、c．176-c．191del16和c．299-c．300delAT,其中,c．79G〉A是主要突变方式,携带率为24．14％（42／174）。新发现两例未见报道的突变位点c．277A〉G和c．558G〉A;患者中只有1例检测到mtDNA1555A〉G位点突变。结论通过对大同地区GJB2基因和mtD-NA1555突变位点的研究,了解大同地区该基因突变谱,为后续国内耳聋基因型分布提供数据支持,同时也为耳聋的早期诊断,治疗提供理论依据。