APPLICATION OF GENETIC DEAFNESS GENE CHIP FOR DETECTION OF GENE MUTATION OF DEAFNESS IN PREGNANT WOMEN

Objective The study is to identify the carrier rate of common deafness mutation in Chinese pregnant women via detecting deafness gene mutations with gene chip. Methods The pregnant women in obstetric clinic without hearing impairment and hearing disorders family history were selected. The informed consent was signed. Peripheral blood was taken to extract genom- ic DNA. Application of genetic deafness gene chip for detecting 9 mutational hot spot of the most common 4 Chinese deafness genes, namely GJB2 （35delG, 176del16bp, 235delC, 299delAT）, GJB3 （C538T） ,SLC26A4 （ IVS72A〉G, A2168G） and mito- chondrial DNA 12S rRNA （A1555G, C1494T） . Further genetic testing were provided to the spouses and newborns of the screened carriers. Results Peripheral blood of 430 pregnant women were detected, detection of deafness gene mutation carri- ers in 24 cases（4.2%）, including 13 cases of the GJB2 heterozygous mutation, 3 cases of SLC26A4 heterozygous mutation, 1 cases of GJB3 heterozygous mutation, and 1 case of mitochondrial 12S rRNA mutation. 18 spouses and 17 newborns took further genetic tests, and 6 newborns inherited the mutation from their mother. Conclusion The common deafness genes muta- tion has a high carrier rate in pregnant women group, 235delC and IVS7-2A〉G heterozygous mutations are common.     

prevalence of the gjb2 mutations in deafness patients of different ethnic origins in xinjiang

Objective To investigate GJB2 mutation prevalences in the Uigur and Han ethnic groups in Xinjiang, China, and determine the relationship between ethnicity and GJB2 gene mutations. Methods Information regarding ethnicity of patients' families was obtained through medical records review and/or patient interview. Blood samples were collected from 61 Uigurs and 66 Hans for direct sequencing of the coding region and intron/exon boundaries of the GBJ2 gene. Results Carrier frequency of GJB2 mutations was similar between the Uigur and Han subjects. The GJB2 35delG mutation was seen only in Uigur patients with hearing loss, whereas the 235delC mutation was identified in both Uigur and Han patients. The allelic Frequency of 35delG mutation was 7.4% (9/122) in Uigur deaf students, but none in Han deaf students (0/128) and Uigur controls (0/196). The allelic frequency of GJB2 235delC mutation in Uigur and Han deaf students was 5.7% and 9.8%, and that of 299-300delAT mutation was 0.8% and 5.5%, respectively. V27I and E114G were the most frequent types of polymorphism. Conclusion We found an Asian-specific GJB2 diversity among Uigurs, and comparable GJB2 contribution to deafness in Uigur and Han patients. The high carrier frequency of 35delG in Uigurs (11.5%) is probably defined by gene drift/founder effect in a particular group. Even though GJB2 mutations have been widely reported in the literature, this discussion represents the first report of GJB2 mutations in Chinese multi-ethnic populations.     

screening of gjb2 mutations in chinese population

The GJB2 gene(connexin 26) has been shown to be responsible for DFNB1 and DFNA3. We screened the GJB2 gene in 488 patients with prelingual deafness(Group 1), 124 with postlingual deafness(Group 2), and 117 normal hearing subjects(Group 3). We found that, in Group 1, 65 patients(13.32%) were homozygotes or compound heterozygotes and 51 patients (10.45%) carried a single pathogenic mutation. The 235delC mutation was the most frequent mutation, accounting for 73.22% of the known pathogenic alleles in Group 1. No homozygotes or compound heterozygotes were detected in Group 2 or Group 3. Some postlingual deaf patients (2.42%) and normal hearing subjects(4.27%) were 235delC carriers. Our preliminary data indicate that 235delC, the most frequent mutation identified in this study, is a major cause for prelingual deafness.     

应用耳聋基因芯片探讨聋哑夫妇的生育风险

目的 通过耳聋基因芯片诊断技术,探讨耳聋易感基因筛选在聋哑人家庭优生优育中的意义.方法 52对聋人夫妻来自长春市某聋哑社区,平均((x)±s)年龄(58.3±6.7)岁.在受检者知情同意情况下采集外周静脉血3 ml,分离基因组DNA,利用遗传性耳聋基因检测芯片对GJB2、SLC26A4、GJB3和线粒体DNA等常见耳聋基因中的9个突变位点进行检测.通过直接测序法验证基因芯片结果.以50名年龄相仿的健康人为对照.结果 所有聋人夫妻纯音测听检查均为双耳非综合征型感音神经性聋.104例患者中,有32例出现GJB2基因突变,占耳聋总人数的30.7%(32/104),包括35delG、176del16、235delC、299delAT;其中18例存在235delC突变,占所有GJB2等位基因突变的59.1%(18/32).SLC26A4基因纯合突变4例,杂合突变3例,均为IVS7-2 A>G突变.问卷调查和基因检测分析发现,52对聋人夫妻,有4个家庭后代出现耳聋成员,占聋人家庭总数的7.6%(4/52).夫妻双方均携带相同基因突变,其子女均发生耳聋,耳聋发生风险为100%.基因芯片的结果与测序方法的结果完全一致.结论 聋哑家庭再生育聋人的风险较高,通过基因芯片技术进行耳聋易感基因检测,可避免明确病因的耳聋家庭出现新的耳聋病例.

Abstract：

Objective To explored the significance of screening the gene mutations of deafness related in deaf-mute ( deaf & dumb) family using DNA microarray. Methods Total of 52 couples of deafmute were recruited from Changchun deaf-mute community. With an averageage of (58. 3 ±6. 7) years old ((x) ± s) . Blood samples were obtained with informed consent. Their genomic DNA was extracted from peripheral blood and PCR was performed. Nine of hot spot mutations in four most common deafness pathologic gene were examined with the DNA microarray, including GJB2, GJB3,PDS and mtDNA 12SrRNA genes. At the same time, the results were verified with the traditional methods of sequencing. Fifty of normal people served as a control group. Results All patients were diagnosed non-syndromic sensorineural hearing loss by subjective pure tone audiometry. Thirty-two of 104 cases appeared GJB2 gene mutation (30. 7% ) , the mutation sites included 35delG,176del16,235delC and 299delAT. Eighteen of 32 cases of GJB2 mutations were 235delC ( 59. 1% ) . Seven of 104 cases appeared SLC26A4 gene IVS7-2 A > G mutation. Questionnaire survey and gene diagnosis revealed that four of 52 families have deaf offspring (7. 6% ). When a couple carries the same gene mutation, the risk of their children deafness was 100%.The results were confirmed with the traditional methods of sequencing. Conclusions There is a high risk of deafness if a deaf-mute family is planning to have a new baby. It is very important and helpful to avoid deaf newborns again in deaf-mute family by DNA microarray.     

新疆喀什地区维吾尔族耳聋人群常见基因突变的研究

目的 采用中国人群遗传性聋常见突变基因类型对新疆喀什地区耳聋人群进行筛查,探究新疆维吾尔族耳聋人群基因突变的检出率及其意义.方法 选取维吾尔族非综合征性聋患者174例作为研究对象,通过病史采集,血样抽取和DNA提取后对所选样本的35delG、176-191del16、235delC、299-300delAT、2168A＞C(H723R)、IVS7-2A＞G、1555A＞G、1494C＞T进行检测,对于位点缺失样本进行序列测定.采用SPSS 17.0软件对数据进行统计学分析.结果 GJB2是主要的突变基因,187delG在维吾尔族耳聋人群中首次发现,也是新发现的GJB2基因的病理性突变;SLC26A4突变在维吾尔族耳聋人群中的检出率低,与对照组比较差异无统计学意义(x2=0.000,P=1.000);mtDNA 12S rRNA突变在维吾尔族耳聋人群中的检出率低,在对照组中未检出;20例有明确耳聋家族史的患者中17例未检出突变.结论 维吾尔族耳聋人群耳聋基因常见突变有其特点,基因全序列分析和家系研究对丰富维吾尔族耳聋基因的突变谱是必要的.

Abstract：

Objective To investigate the frequency of the mutations in Uyghur nonsyndromic deafness groups in Kashgar region of Xinjiang province by means of screening the common mutations of known deafness genes in China. Methods One hundred and seventy-four Uyghur patients with hearing loss were involved in this study. Questionnaire survey was conducted and peripheral blood samples were collected for polymerase chain reaction. Screening was performed for 35delG, 176-191del16, 235de1C, 299-300delAT, 1555A ＞ G, 1494C ＞ T, 2168A ＞ G and IVS7-2A ＞ G. DNA sequence analysis was performed for the samples with absent signals at some loci. SPSS 17.0 software was used to analyze the data. Results Mutation of GJB2 was the most common among the three known deafness genes. 187delG was found for the first time in Uyghur groups with hearing loss and was a new pathological mutation of GJB2. The mutation rate of SLC26A4 was low in the experimental group with no significant difference when compared with the control group. The mtDNA 12S rRNA mutation rate in the deaf group was low but not detected in the control group.In addition, mutations were not dectected in 17 cases among the 20 patients with positive family history.Conclusion The mutation rate and dominant mutation of Uyghur ethnic nonsyndromic deaf groups have their own characteristics, it is necessary to conduct a sequence analysis and a stemma studying for an aim of perfecting the mutation spectrum of Uyghur deafness gene.     

GJB2 mutation spectrum in deaf population in a typical southeastern area of China

Mutations in GJB2 gene are the most frequently found mutations in patients with nonsyndromic hearing impairment. However, the spectrum and prevalence of mutations in this gene vary among different ethnic groups. In China, 30,000 infants are born with congenital hearing impairment annually. In order to provide appropriate genetic testing and counseling to the families, we investigated the molecular etiology of nonsyndromic deafness in 103 unrelated school children attending Nantong School for the Deaf and Mute in Jiangsu Province, China. The coding exon of the GJB2 gene was PCR amplified and sequenced. Sixty two GJB2 mutant alleles were identified in 35.9% (37/103) of the patients. Twenty five patients carried two pathogenic mutations and 12 patients carried one mutant allele. The 235delC was the most common mutation accounting for 69.4% (43/62) of GJB2 mutant alleles. The GJB2 mutant alleles accounted for 30.1% (62/206) of all chromosomes responsible for nonsyndromic hearing impairment. Testing of the 3 most prevalent deleterious frame shift mutations in this cohort detected 100% of all GJB2 mutant alleles. These results demonstrate that an effective genetic testing of GJB2 gene for patients and families with nonsyndromic hearing impairment is possible.     

hearing in old age - epidemiological and etiological aspects

Introduction The number of elderly persons is increasing all over the world. This strong demographic trend will affect our societies profoundly. Since the largest relative increase is expected to occur in the group of very old persons, over 80 years of age, the demands on the society and on relatives in terms of health- and social care are considerable. The influence of ageing on the auditory function is pronounced. Age-related hearing loss (ARHL), or presbyacusis, belongs to one of the three most frequently reported chronic health problems in old age, and is also the most prevalent cause of hearing loss. Communication with other people, and the auditory system is the most important link in communication. ARHL has often a devastating effect on the social contacts and quality of life of many elderly people.     

散发性耳聋GJB2基因突变分析

目的 通过分析内蒙古鄂尔多斯和呼和浩特地区散发性耳聋患者GJB2 235delC点突变,以探讨散发性耳聋患者的分子病因学。方法 对131例(汉族92例,蒙古族39例) 散发性耳聋患者进行耳聋病因学问卷调查、纯音听阈及声导抗测试。聚合酶链反应(polymerase chain reaction PCR)扩增目的片段并用限制性内切酶对其进行GJB2 235delC基因突变检测,对酶切检测结果呈阳性的样本用直接测序法进行验证。对50例健康中国人和100例健康加拿大白种人行限制性内切酶GJB2 235delC点突变检测,作为阴性对照。结果 131例散发性耳聋患者全部为感音神经性聋。在该群体中4例(汉族3例,蒙古族1例)存在GJB2 235delC纯合性突变;3例(汉族2例,蒙古族1例)存在GJB2 235delC杂合性突变。50例健康中国人对照组中检测出1例GJB2 235delC点突变携带者,100例健康加拿大白种人中未检测到GJB2 235delC点突变。结论 GJB2 235delC点突变是中国人散发性感音神经性耳聋的分子病因学之一。内蒙古地区汉族、蒙古族GJB2 235delC突变频率无明显差异,对GJB2 235delC点突变的基因筛查可以明确一些散发性耳聋患者的病因,从而对基因突变引起的散发性耳聋的早期诊断、遗传咨询及防聋治聋起到重要作用。     

常染色体隐性遗传耳聋家系的诊断和产前诊断

目的对常染色体隐性遗传耳聋家系进行基因检测与产前分子诊断。方法收集先证者临床及家系资料,应用聚合酶链式反应(PCR)、限制性内切酶法检测,并用直接测序技术确认,对该家系成员的GJB2基因外显子进行序列分析。并运用STR位点分析方法排除产前诊断中母体基因组DNA的污染。结果家系1先证者GJB2基因分型为235delC/299-300delAT复合杂合突变,系重度感音神经性耳聋,父亲和母亲分别为299-300delAT和235delC杂合突变携带者,临床表型均正常。家系2先证者系235delC纯合突变,其父母均为235delC杂合突变携带者,具有正常听力表型。产前诊断结果显示,家系1和家系2胎儿GJB2基因分型分别为299-300delAT杂合突变和235delC杂合突变。结论 GJB2基因235delC纯合性突变和235delC/299-300delAT复合杂合突变均为耳聋致病突变,结合运用STR位点分析方法有助于在产前诊断中排除母体DNA对胎儿DNA的污染,产前诊断和早期干预能避免耳聋患儿的出生。     

Long-term results of type I tympanoplasty with perichondrium reinforced cartilage palisade vs temporalis fascia for large perforations: A retrospective study

Objective:To compare the rate of graft uptake and postoperative hearing outcomes of Type I tympanoplasty with perichondrium reinforced cartilage palisade to temporalis fascia for large tympanic membrane(TM)perforations over 5 years follow-up period.Materials and methods:This was a retrospective comparative study involving patients with chronic otitis media with large TM perforations.The patients underwent type I tympanoplasty using either perichondrium reinforced cartilage palisade(CP group)or temporalis fascia(TF group)as the graft via a transmeatal approach and under local anesthesia.Morphological and functional results were recorded at three-and five years follow-up.Demographic profiles including age and sex,surgery side,contralateral disease and graft uptake rate,as well as hearing outcomes,were compared between the two groups.Results:At three years follow-up,graft uptake was 94.87%for perichondrium reinforced cartilage palisade and 80.7%for fascia,respectively,(p?0.67).At five years follow-up,the uptake rate dropped to 87.17%in the CP group,but to 66.6%in the TF group(p?0.019).Hearing improved after surgery in both groups,and showed no significant difference between the two groups.Conclusion:Over long-term,perichondrium reinforced palisade showed a statistically significant better outcome regarding graft uptake than temporalis fascia in type Itympanoplasty for large TM perforations with comparable audiometric results.     

Audiometric evaluation of carriers of the connexin 26 mutation 35delG

Mutation in a gap junction protein gene (GJB2 also named connexin 26) is a major cause of autosomal recessive congenital deafness, which is responsible for about 80% of the cases in Mediterranean families, but actually little is known about the influence of GJB2 mutations on the hearing of obligate carriers. We examined GJB2 35delG mutation carrier individuals to test the possible presence and incidence of audiometric abnormalities among carriers of 35delG mutations. Tonal audiometric analysis was performed on a 35delG mutation carrier group (H) and on a non-carrier control group (N). Audiometric evaluations in the control group showed the presence of thresholds within normal limits at all frequencies, while carriers of 35delG mutations presented a decrease of hearing principally at 6,000 and 8,000 Hz. The difference at 6,000 and 8,000 Hz between groups H and N is statistically significant.     

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...     

GJB2 gene mutations in newborns with non-syndromic hearing impairment in Northern China

Mutations in GJB2 account for the majority of recessive forms of prelingual hearing loss. However, in most previous studies it was not possible to distinguish between congenital (present at birth) and non-congenital prelingual hearing loss. In the present study, the frequency of GJB2 alleles in 20 newborns with bilateral severe-to-profound non-syndromic hearing impairment (NSHI) who were found at birth through newborn hearing screening and clinical examination is reported. PCR was used to amplify the coding region of GJB2 gene followed by sequencing analyses. Fifty volunteers with normal hearing were included as controls. Results showed that three cases were 235delC/235delC homozygotes; one was 235delC/605ins46 compound heterozygotes, 605ins46 mutation was a novel mutation reported in the Chinese population; another was 235delC/299-300delAT compound heterozygotes. 25% (5/20) of the deafness in newborns studied was caused by GJB2 gene mutations. The frequency of 235delC allele carrier in patients and in control group was 22.5% and 1%, respectively. One case was identified as being a 235delC heterozygote without other mutations detected. Besides, multiple polymorphisms such as V27I, V37I, E114G, T123N were also detected. In conclusion, GJB2 analysis is an important test that identifies a major cause of newborns with bilateral severe-to-profound NSHI screened by universal newborn hearing screening in Northern China. The most common pathologic mutation of GJB2 in studied cases was 235delC. Molecular analysis and genetic counseling will be extremely important for congenital deafness present at birth.     

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。     

江苏南通地区非综合征性耳聋GJB2基因突变分析

目的 研究南通地区非综合征性耳聋GJB2基因突变情况。方法 收集南通地区海安县和如皋县聋哑学校学生100名和健康对照组50名，利用PCR扩增及限制性内切酶酶切分析初筛GJB2 235delC突变者，然后再行DNA直接测序。结果 耳聋组中共发现三种突变：235delC、176—191del16、299—300delAT。235delC是主要突变方式．约30％的患者携带此突变；299—300delAT和176-191del16突变检出率分别为9％和8％。对照组未发现这些突变。结论 南通地区非综合征性耳聋GJB2基因突变率较高，因此在南通地区进行广泛的生育前耳聋基因筛查工作有重要意义。     

河北涿州、高碑店市特教学校非综合征性聋分子病因学分析——GJB2 235delC突变、SLC26A4 IVS7-2A＞G突变和线粒体DNA12SrRNA A1555G突变筛查报告

目的 对河北涿州、高碑店地区重度耳聋患者进行分子流行病学调查，了解耳聋的常见分子病因。方法 对河北涿州、高碑店市特殊教育学校64名耳聋学生进行遗传性耳聋问卷调查、全面的体格检查、耳鼻咽喉专科检查以及听力学评估（包括纯音测听和声导抗）。对64名非综合征型感音神经性耳聋患者分别进行GJB2基因235delC突变、线粒体DNA 12SrRNA基因A1555G点突变的限制性内切酶分析。应用直接测序法检测SLC26A4基因IVS7—2A〉G突变。结果7例（10．93％）携带GJB2基因235delC纯合突变；9例（14．06％）携带GJB2基因235delC杂合突变；6例（9．37％）携带SLC26A4基因ⅣS7—2A〉G纯合突变，12例（18．75％）携带SLC26A4基因IVS7—2A〉G杂合突变：未发现携带线粒体DNA 12SrRNA基因A1555G点突变者。结论 河北涿州、高碑店地区非综合征型耳聋患者存在较高的GJB2基因235delC和SLC26A4基因ⅣS7—2A〉G突变发生率，而线粒体DNA 12SrRNA基因A1555G突变发生率低于全国平均水平。聋病分子流行病学调查提示河北涿州、高碑店地区20.3％的非综合征型耳聋患者在分子水平能够明确诊断．另有32．81％的患者有遗传倾向。进行准确的耳聋早期诊断、遗传咨询、及时干预和治疗在这一地区的聋哑人群中非常重要。     

武汉地区非综合征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 分析武汉地区非综合征性耳聋（nonsyndromic hearing impairment，NSHI）患儿GJB2 235delC突变率和线粒体DNA A1555G突变率。方法 收集武汉市艺萌听力康复中心的94例耳聋患儿血样，非综合征性耳聋患儿88例，提取DNA后经聚合酶链反应（PCR）分别扩增GJB2基因编码区及线粒体DNA，ApaI酶切分析GJB2 235位点的C缺失突变，Prev—DAF药物性耳聋基因诊断试剂盒分析线粒体1555位点的A—G突变，对GJB2 235ddC及线粒体DNA A1555G的突变率进行统计分析。结果 88例患儿中9例（10．23％）为GJB2 235delC纯合突变，7例（7．96％）为GJB2 235delC杂合突变；2例（2．27％）存在线粒体DNA A1555G点突变。在分子水平能够明确诊断者占20．46％。结论 武汉地区耳聋患者存在较高的遗传性耳聋发生率，应用基因诊断技术可以在耳聋患者病因调查中进行快速诊断筛查，达到防止再生育聋儿、指导聋儿康复等积极效果。     

大同市特教学校非综合征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 进行山西省大同地区重度耳聋的分子流行病学调查。方法 对山西省大同市特殊教育学校152名耳聋学生进行遗传性耳聋问卷调查、全面的体格检查、耳鼻咽喉专科检查以及包括纯音测听和声导抗在内的听力学评估。对148名非综合征型感音神经性耳聋患者分别进行线粒体DNA 12SrRNA基因A1555G点突变和GJB2基因235delC突变的限制性内切酶分析。结果 3例（2．03％）存在线粒体DNA 12SrRNA基因A1555G点突变，16例（10．8l％）存在GJB2基因235delC纯合突变，2l例（14．19％）存在GJB2基因235delC杂合突变，能够明确进行基因诊断者占27．03％。结论 山西省大同地区非综合征型耳聋患者存在较高的GJB2基因235delC突变发生率．而线粒体DNA 12SrRNA基因A1555G突变发生率低于全国平均水平。通过聋病分子流行病学调查．提示27．03％的非综合征型耳聋患者具有明确或强烈的遗传倾向，对于大同地区耳聋的预防、治疗及康复有着较好的意义。