湖南郴州非综合征型聋患者GJB2、SLC26A4和线粒体DNA12SrRNA基因突变分析

目的探讨湖南郴州非综合征型聋患者的分子病因特点。方法采取湖南郴州154名非综合征型聋患者的外周血,提取DNA,采用基因芯片筛查GJB2、SLC26A4和线粒体DNA12SrRNA基因的热点突变,基因芯片法未确诊的样本则采用DNA测序法进一步检测。结果两种方法共在34例(22.08%,34/154)患者中检出7种GJB2基因突变,其中235delC(13.63%,21/154)发生率最高,其次是299delAT(9.09%,14/154);在8例伴有大前庭水管的患者中检测出7种SLC26A4基因突变,包括一种新突变Q696X;3例患者被检出线粒体DNA12SrRNA基因突变。结论湖南郴州非综合征型聋患者中GJB2、SLC26A4和线粒体DNA12SrRNA基因突变的发生率与中国大部分地区相似,Q696X为新发现的SLC26A4基因突变。     

新疆地区非综合征性聋患者线粒体12S rRNA基因突变位点分析

目的　研究新疆地区非综合征性聋患者线粒体12S rRNA基因突变情况。方法　收集2012～2016年新疆维吾尔自治区人民医院耳鼻咽喉诊疗中心就诊794例非综合征性聋患者和623名听力正常母系家庭成员样本。所有患者进行听力学相关测试,同时采集非综合征性聋患者及部分正常母系家庭成员口腔黏膜组织,将样本送至中优细胞分 子遗传学检测中心进行线粒体12S rRNA基因突变检测,对检测结果进行统计分析。结果　线粒体12S rRNA基因突变分析共发现17个突变位点,已知A1555G、C1494T、961DelT/InsC和T1095C突变在非综合征性聋患者中检出率分别为1.01%、0.13%、0.76%和0.25%,其他突变均为多态性位点。A1555G和961DelT/InsC突变在汉族非综合征性聋患者检出率显著高于维吾尔族（P =0.001）。C1494T突变在维吾尔族非综合征性聋患者检出率仍然为0,T1095C在两个民族非综合征性聋患者检出率无显著性差异。结论　此次新疆地区线粒体12S rRNA A1555G突变在聋病人群检出率有所降低,与耳毒性药物使用量降低有关。线粒体12S rRNAA1555G和961DelT/InsC在汉族中携带率显著高于维吾尔族,维吾尔族中可能存在其他相关的热点突变位点。     

新疆维吾尔族和汉族非综合征型遗传性聋患者线粒体DNA 12S rRNA A1555G、GJB2及GJB3基因突变研究

目的 分析新疆地区维吾尔族和汉族非综合征型遗传性聋患者线粒体DNA、GJB2、GJB3 基因突变情况,探讨新疆地区维吾尔族非综合征型遗传性聋人群中上述基因突变位点及突变频率与汉族耳聋人群的差异.方法 对93名新疆地区非综合征型遗传性聋患者进行耳聋病因问卷调查和纯音听阈测试,另选取110名听力正常人作为对照组.耳聋组中维吾尔族43例,汉族50例;对照组中维吾尔族56例,汉族54例.收集血样,提取DNA后经聚合酶链反应(PCR)扩增.GJB3基因PCR产物直接测序;针对线粒体DNA 12S rRNA A1555G点突变进行限制性内切酶分析,挑选阳性者进一步测序.对83例非综合征型遗传性聋患者和98例正常人行GJB2基因测序,其中耳聋组维吾尔族43例,汉族40例,对照组维吾尔族46例,汉族52例.结果 93名患者中检测到GJB3基因33C-T2例、766G-A 2例、357C-T 7例以及798C-T4例.8例患者存在线粒体DNA 12SrRNA A1555G突变,其中维吾尔族2例,汉族6例.在耳聋患者中,发现9种GJB2碱基改变:109G-A、233-235delC、79G-A、196G-A、341A-G、564G-A、380G-A、71G-A及35delG.对照组检测到GJB3基因357C-T 4例、798C-T 5例及93C-T 2例.对照组发现9种GJB2基因碱基改变:341A-G、380G-A、457G-A、79-G-A、109G-A、281A-G、21G-T、171G-T及368C-A.线粒体DNA A1555G突变频率在汉族耳聋组与对照组之间差异具有统计学意义(P＜0.05).GJB2基因79G-A突变在两个民族耳聋组和对照组的分布差异均具有统计学意义(P值均＜0.05);而341A-G突变在耳聋组两个民族之间的分布差异具有统计学意义(P＜0.05).GJB3基因798C-T的突变频率无论在耳聋组还是对照组,维吾尔族和汉族之间的差异均具有统计学意义(P值均＜0.05).结论 新疆地区线粒体DNA 12S rRNA A1555G突变检出率较高.GJB3 基因突变并非新疆地区非综合征型遗传性聋患者的主要致病原因.该地区GJB2和GJB3突变具有种族和地域性特点.     

205例先天性非综合征型聋患儿GJB2基因突变分析

目的探讨先天性非综合征型聋婴幼儿的GJB2基因突变频率、突变热点和听力学表型特点。方法对来自上海及周边地区的205例先天性非综合征型聋患儿GJB2基因PCR扩增产物行酶切鉴定以及直接测序法进行突变检测,对1例GJB2基因235delC纯合突变先证者母亲再次妊娠19周时通过羊水穿刺行产前诊断。结果205例先天性非综合征型聋儿中,共发现GJB2基因移码突变49例,占23.90%（49/205）,其中46例患儿存在GJB2基因235detc突变,占93.88%（46/49）,GJB2基因突变者多为中到极重度听力损失。1例胎儿产前诊断确诊为GJB2基因235delC纯合突变。结论GJB2基因纯合或复合杂合移码突变可导致非综合征型常染色体隐性遗传性聋,听力损失程度多为中度至极重度;235delC突变占所有突变等位基因85.88%。     

中国湖南地区非综合征性聋患者SLC26A4基因突变研究

目的:研究SLC26A4基因突变在中国湖南地区耳聋人群中的突变频率和突变热点.方法: 采集来自湖南各地区的非综合征性聋患者的血液样本96例,PCR扩增后,应用变性高效液相色谱技术对SLC26A4基因的全部21个外显子19个片段进行筛查,对变性高效液相色谱筛查发现异常的PCR扩增样本进行测序.测序结果运用DNASTAR软件进行分析.结果:在15例患者中检测到了SLC26A4基因突变,检出率为15.6%,其中3例纯合突变,10例为复合杂合突变,2例杂合突变.共发现了16种不同类型的碱基变异,包括10种已知突变(S90L、S252P、IVS7-2A>G、T410M、N392Y、IVS10-12T>A、S448X、G497S、S517fs、H723R),4种新发突变(S8X、A227P、C565fs、Y728H),1种同义突变(c.2182 T>C)和1种多肽(IVS11+47 T>C).在该研究中IVS7-2A>G 突变最多,共检测到9例,检出率为9.38%,占所有突变等位基因的5.73%.最常见的多肽为IVS11+47 T>C,共检测到20例.结论:在湖南地区非综合征性聋患者中IVS7-2A>G是SCL26A4基因最常见的突变方式;该研究所发现的4例新发突变在一定程度上丰富了中国人SLC26A4基因突变图谱.     

新疆地区941例新生儿聋病基因GJB2、SLC26A4、线粒体DNA 12S rRNA筛查分析

目的通过对新生儿进行聋病易感基因和听力筛查,探讨聋病易感基因筛查应用于新生儿筛查的必要性,为制订防聋治聋策略提供依据。方法以941例新生儿作为研究对象,所有新生儿出生时采脐带血,采用限制性内切酶酶切结合直接测序的方法对3种国人常见耳聋易感基因（线粒体DNA 12S rRNA、GJB2、SLC26A4）突变热点进行筛查,运用SPSS 13.0软件对结果进行统计分析。结果3种基因热点突变的总携带率为2.02%（19/941）,GJB2基因235delC杂合突变9例（0.96%）,SLC26A4基因IVS7-2A〉G杂合突变9例（0.96%）,线粒体DNA 12S rRNA A1555G突变3例（0.32%）,其中2例为复合突变（235delC杂合突变/IVS7-2A〉G杂合突变、1555A〉G均质突变/235delC杂合突变）。GJB2基因235delC杂合突变在维吾尔族和汉族新生儿中的携带率分别为0.36％（1/276）、1.19％（7/586）;SLC26A4基因IVS7-2A〉G杂合突变在维吾尔族和汉族新生儿中的携带率分别为0.36％（1/276）、1.37％（8/586）;线粒体DNA 12S rRNA 1555A〉G突变在维吾尔族和汉族新生儿中的携带率分别为0.72％（2/276）、0％。在维吾尔族和汉族新生儿中,以上三基因突变携带率不同,但没有统计学差异。结论聋病易感基因筛查应用于维、汉族新生儿筛查必要且可行。     

陕西省800例非综合征型聋患者常见致聋基因突变分析

目的：分析陕西省非综合征型聋患者常见耳聋基因突变方式及频率,了解其耳聋发病的分子机制。方法采集陕西省800例非综合征型聋患者外周血,提取基因组DNA ,采用聚合酶链式反应（PCR）扩增GJB2基因、GJB3基因、SLC26A4基因以及线粒体12S rRNA 1494和1555位点进行直接测序,序列与NCBI网站公布的标准序列进行比对分析。结果800例非综合征型聋患者中,共353例（44．13％）患者携带耳聋致病基因突变,其中153例（19．13％）携带GJB2基因双等位基因突变,GJB2基因最常见的突变方式为235delC ,检出率为13．5％（216／1600）;123例（15．38％）携带SLC26A4基因双等位基因突变,SLC26A4基因最常见突变方式为IVS7－2A＞G ,检出率为7．44％（119／1600）;1例携带线粒体12S rRNA1494C＞ T均质性突变,15例携带1555A＞G均质性突变;2例患者携带GJB3基因c ．538C＞ T杂合突变。294例（36．75％,294／800）患者由上述基因突变导致耳聋。结论陕西省非综合征型聋患者中,GJB2基因以及SLC26A4基因的突变携带率与全国以及西北地区平均水平较为一致,而线粒体基因突变的携带率偏低。     

新疆维吾尔族和汉族非综合征型感音神经性聋患者线粒体DNA 12SrRNA A1555G突变分析

目的研究新疆地区维吾尔族(简称维族)和汉族非综合征型感音神经性聋患者线粒体DNA(mtD-NA)12SrRNAA1555G突变频率并比较其差异。方法新疆地区非综合征型遗传性聋患者维族88例、汉族75例为患者组;正常人维族82例、汉族78例为正常对照组。抽取所有受检者外周静脉血,从白细胞中提取DNA,多聚酶链反应(PCR)扩增线粒体DNA目的片断,Alw26I限制性内切酶检测A1555G点突变,对阳性病例的PCR产物进行DNA测序验证。结果正常组中未检出DNA12SrRNAA1555G突变,患者组中检出16例mtDNAA1555G点突变,12人为汉族(16.00%,12/75),4人为维族(4.55%,4/88),汉族与维族比较差异有统计学意义(χ2=6.001,P=0.014)。结论线粒体DNA12SrRNAA1555G突变是导致新疆地区非综合征型遗传性聋患者致病的主要原因之一,汉族高于维吾尔族。     

非综合征型聋患者线粒体DNA A1555G突变频率分析

目的分析甘肃地区非综合征型聋患者线粒体DNA 12SrRNA A1555G的突变频率。方法收集甘肃地区五所聋哑学校802例聋哑学生血样，经基因组DNA提取后进行聚合酶链反应（polymerase chain reaction，PCR）扩增线粒体DNA目的片段，用A1w26I限制性内切酶检测A1555G点突变，对酶切阳性病例的PCR产物纯化后进行直接测序验证酶切结果，分析线粒体DNA 12SrRNA A1555G在甘肃地区的突变频率。结果802例聋哑学生中有67例经酶切及直接测序证实为线粒体DNA A1555G突变，突变频率为8．4％（67／802）。其中有15例母系家庭成员中还有2例以上耳聋患者。结论线粒体DNA 12SrRNA A1555G点突变在甘肃地区非综合征型聋患者中占有很高的比例，高于国内外其他地区的相关报道。此研究结果不仅为绘制中国人群线粒体DNA A1555G突变频谱增添新的内容，也为因地制宜地开展耳聋基因诊断、实施遗传咨询及积极预防耳聋的发生有重要的指导意义。     

GJB2, SLC26A4, and mitochondrial DNA12S rRNA hot-spots in 156 subjects with non-syndromic hearing loss in Tengzhou,China

Conclusion: In this cohort of 156 non-syndromic hearing-impaired subjects of Tengzhou area, the most common deafness-associated genes GJB2, SLC26A4 and mtDNA 12S rRNA were investigated by SNPscan efficiently. GJB2 c.235delC and SLC26A4 c.IVS7-2A?>?G were the most common mutation sites. Objectives: Until now, there is no systematic gentic analysis in patients with non-syndromic hearing loss for Tengzhou area, so we evaluated the molecular etiology to investigate the hot-sports. Methods: Peripheral blood samples were obtained from 156 patients with severe-to-profound non-syndromic deafness in Tengzhou. The SNP scan assay technique was performed for a rapid multiplex genetic screening to detect the 115 mutations of the most common three genes. All results were statistically analyzed with SPSS software. Results: Among the 156 analyzed patients, 60 patients were demonstrated with deafness genes, accounting for 38.46% (60/156), including GJB2 (22.44%, 35/156), SLC26A4 (13.66%, 22/156), and mtDNA 12S rRNA (2.56%, 4/156). In this study, we confirmed 23 deafness-causing mutations and 27 different allelic combinations including GJB2 (eight variants, 11 allelic combinations), SLC26A4 (13 variants, 16 allelic combinations) and mtDNA 12S rRNA (two variants). The occurrence rates of these deafness-causing mutations GJB2 c.235delC and SLC26A4 c.IVS7-2A?>?G were significantly higher than other mutation sites (p?相似文献   

云南省10个非综合征性聋家系mtDNA 12S rRNA A1555G突变流行病学调查及分析

目的:探讨云南省10个非综合征性感音神经性聋家系的mtDNA 12SrRNA A1555G突变筛查、流行状况、遗传规律及对于特定药物干预预防的意义。方法:对10个家系以现场问卷方式调查母系家族耳聋的发病情况并绘制出详细的母系家系图,然后对自愿参与检测的母系成员采外周静脉血提取DNA,PCR扩增目的片段、限制性酶切检测A1555G突变阳性个体。结果:10个家系中参与采血者共96例,其中听力正常36例,感音神经性聋患者60例;经过筛查,4例无A1555G位点突变,92例(95.8%)有A1555G位点突变,其中7例为异质性表现,85例为均质性突变;73例有明确氨基苷类抗生素用药史,其余抗生素用药史不明确。结论:云南省药物性聋患者的比例较大,并且mtDNA 12SrRNA A1555G突变率高,对该地区进行mtDNA 12SrRNA A1555G突变筛查及药物干预预防宣教有重要意义。     

贵阳市盲聋哑学校非综合征性耳聋分子病因学分析——GJB2 235delC突变和线粒体DNA 12SrRNA A1555G突变筛查报告

目的 进行贵州省贵阳地区非综合征性耳聋分子病因学调查。方法 对贵阳市盲聋哑学校150名聋哑学生进行耳聋病因问卷调查、纯音听阈测试，对其中139名非综合征性耳聋患者进行线粒体DNA 12SrDNA A1555G点突变和GJB2基因235delC突变限制性内切酶的分析。结果 139名非综合征性耳聋患者中。6例（4132％）存在线粒体DNA 12SrDNA A1555G点突变；17例（12．23％）存在GJB2 235delC纯合突变；9例（6．47％）存在GJB2235delC杂合突变，在分子水平能够明确诊断者占23．02％。结论 贵阳地区耳聋患者存在较高的遗传性耳聋发生率，线粒体DNA A1555G突变发生率和GJB2 235delC突变发生率均高于全国平均水平。耳聋基因诊断技术可以应用在地区性耳聋病因调查中进行快速筛查、诊断，并可达到防止再出生聋儿，指导聋儿康复等积极效果。     

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.     

Absence of GJB6 mutations in Indian patients with non-syndromic hearing loss

Objective

Hearing loss is the most frequent sensory defect in human being. Genetic factors account for at least half of all cases of profound congenital deafness. The 13q11-q12 region contains the GJB2 and GJB6 genes, which code connexin 26 (CX26) and connexin 30 (CX30) proteins, respectively. Mutations in the gene GJB2, encoding the gap junction protein connexin 26, are considered to be responsible for up to 50% of familial cases of autosomal recessive non-syndromic hearing loss and for up to 15-30% of the sporadic cases. It has also been reported that mutations in the GJB6 gene contribute to autosomal recessive and autosomal dominant hearing defects in many populations. The 342-kb deletion [del(GJB6-D13S1830)] of the Cx30 gene is the second most common connexin mutation after the CX26 mutations in some NSHL populations. The aim of this study was to screen GJB6 gene mutations in Asian Indian patients with autosomal non-syndromic hearing loss.

Methods

We screened 203 non-syndromic hearing loss patients, who were negative for homozygous mutations in GJB2 gene, for GJB6-D13S1830 deletion and mutations in coding regions of GJB6 using polymerase chain reaction, denaturing high performance liquid chromatography and direct sequencing.

Results

No deleterious mutation in GJB6 gene was detected in our study cohort.

Conclusion

The present data demonstrated that mutations in the GJB6 gene are unlikely to be a major cause of non-syndromic deafness in Asian Indians.     

A novel variant of SLC26A4 and first report of the c.716T>A variant in Iranian pedigrees with non-syndromic sensorineural hearing loss

The autosomal recessive non-syndromic hearing loss (ARNSHL) can be associated with variants in solute carrier family 26, member 4 (SLC26A4) gene and is the second most common cause of ARNSHL worldwide. Therefore, this study aims to determine the contribution of the SLC26A4 genotype in the hearing loss (HL) of 40 ARNSHL pedigrees in Iran. A cohort of the 40 Iranian pedigrees with ARNSHL, having no mutation in the GJB2 gene, was selected. The linkage analysis with five short tandem repeat (STR) markers linked to SLC26A4 was performed for the 40 ARNSHL pedigrees. Then, two out of the 40 pedigrees with ARNSHL that linked to DFNB4 locus were further screened to determine the variants in all exons of SLC26A4 gene by direct DNA sequencing. The 21 exons of SCL26A4 were analyzed for the two pedigrees. A known variant (c.716T>A homozygote), it is the first reported incidence in Iran, a novel variant (c.493A>C homozygote) were detected in the two pedigrees and pathogenesis of c.493A>C confirmed in this study with review 100 hearing ethnically matched controls by PCR-RFLP analysis. The present study suggests that the SLC26A4 gene plays a crucial role in the HL occurring in Iranian pedigrees. Also, the results probably support the specificity and unique spectrum of SLC26A4 variants among Iranian HL patients. Molecular study of SLC26A4 gene may lead to elucidation of the profile of the population-specific variants which has importance in diagnostics of HL.     

非综合征型感音神经性聋儿GJB2基因突变分析

目的 对散发聋病患儿进行GJB2基因突变检测,探究其在遗传性聋临床工作中的意义.方法 收集门诊139例散发非综合征型感音神经性聋患儿及150例听力正常个体的外周血DNA样本共289例,采用聚合酶链反应分析方法扩增GJB2基因片断进行序列分析.结果 139例病患组中发现GJB2基因突变31例,占22.30%.其中235d...     

线粒体DNA1555位点和GJB2基因及SLC26A4基因的诊断方法及临床应用

目的建立常见耳聋基因如线粒体DNA（mtDNA）1555位点、GJB2基因、SLC26A4（Pendren’s syndrome gene，PDS gene）基因突变的临床检测方法。方法来自门诊的散发耳聋患者367例，有母系家族遗传史耳聋患者60例（27个家系），来自聋哑学校的先天性聋患者20例，来自门诊经高分辩CT证实双侧前庭水管扩大患者3例，无感音神经性聋病史的对照个体50例。应用线粒体基因A1555G突变检测试剂盒检测线粒体基因1555位点的突变情况；针对20例语前聋患者进行GJB2全序列分析；针对3例大前庭水管综合征的患者，应用变性高效液相色谱技术进行SLC26A4基因的全部外显子筛查，出现异常波形之外显子行序列分析。结果在26个家系的59例患者和5例散发患者中发现mtDNA A1555G突变；20例先天性聋中发现2例GJB2 235delC纯合突变，酶切加测序发现1例235delC＋299-300delAT复合突变，均为先天性聋的肯定原因，另外2例具有109G-A杂合突变；3例大前庭水管综合征患者的变性高效液相色谱技术筛查均发现包含第7、8外显子的扩增子具有异常波形，测序证实1例为杂合的SLC26A4 G316X突变；另2例为919-2 A-G纯合突变。结论耳聋基因诊断具有显著的临床意义，可操作性强，在不远的将来耳聋的基因诊断可能会正式列为耳科临床检测项目。