本文選題：耳聾 + 聽力損失　； 參考：《蘭州大學(xué)》2014年博士論文

【摘要】：聽力損失影響人的言語(yǔ)功能進(jìn)而導(dǎo)致社會(huì)交流障礙,因其發(fā)病率高而成為醫(yī)學(xué)界研究的熱點(diǎn)。從1995年克隆第一個(gè)耳聾基因起,超過80個(gè)非綜合征耳聾相關(guān)基因被發(fā)現(xiàn),越來越多的聾病分子病因得以揭示。中國(guó)人口的多民族結(jié)構(gòu)蘊(yùn)含著豐富的遺傳資源,特別是不同族源的少數(shù)民族人群更為珍貴。本研究旨在進(jìn)行中國(guó)西北地區(qū)少數(shù)民族耳聾資源的收集,建立一個(gè)覆蓋西北地區(qū)的包含主要少數(shù)民族的臨床資源和遺傳資源庫(kù)。在此基礎(chǔ)上,通過開展三種常見的耳聾相關(guān)基因的分子流行病學(xué)研究,并對(duì)一個(gè)耳聾家系進(jìn)行候選基因突變研究,試圖揭示這些基因在少數(shù)民族耳聾人群的流行狀況,繪制各民族的聾病基因突變頻譜,分析開展耳聾家系致病基因定位克隆的有效方法,探索針對(duì)少數(shù)民族的基因診斷策略。我們研究的主要內(nèi)容由以下四部分組成：第一章：中國(guó)西北地區(qū)少數(shù)民族聾病資源庫(kù)建設(shè) 少數(shù)民族耳聾群體較為稀缺,遺傳性耳聾資源有減少的趨勢(shì)。為了保護(hù)少數(shù)民族的遺傳性耳聾資源,我們研究建立西北地區(qū)少數(shù)民族的聾病資源庫(kù)。研究中制定了資源庫(kù)建設(shè)的規(guī)范流程,通過本地區(qū)各特教學(xué)校、各級(jí)殘聯(lián)及醫(yī)院門診等渠道收集耳聾散發(fā)病例及耳聾家系資源,進(jìn)行問卷調(diào)查、物理檢查、聽力學(xué)檢查并繪制家系圖譜,簽訂知情同意書并抽取外周靜脈血,提取基因組DNA,建立全套資料的數(shù)據(jù)化管理模式。我們共收集到少數(shù)民族1673例耳聾散發(fā)病例和146個(gè)遺傳性耳聾家系,完成了臨床和遺傳資源的收集,為進(jìn)一步開展遺傳學(xué)研究積累了資源。我們認(rèn)為需要進(jìn)一步加強(qiáng)少數(shù)民族耳聾資源的保護(hù),建立科學(xué)完善的資源庫(kù)管理體系。第二章：西北少數(shù)民族非綜合征感音神經(jīng)性聾的GJB2基因突變篩查研究 GJB2基因是目前發(fā)現(xiàn)的先天性耳聾最常見的分子病因,GJB2基因編碼縫隙連接蛋白Cx26,在內(nèi)耳電解質(zhì)交換和細(xì)胞間信息傳遞發(fā)揮重要作用,與人類聽覺功能關(guān)系密切。我們?cè)谖鞅鄙贁?shù)民族散發(fā)感音神經(jīng)性聾患者群體中開展GJB2基因的突變研究,目的是分析該基因的流行狀況,揭示少數(shù)民族的熱點(diǎn)突變,探討適合這些耳聾人群的基因診斷策略。研究中完成了共1330例非綜合征感音神經(jīng)性聽力損失患者的GJB2基因的突變篩查,并且完成了同地區(qū)457例漢族耳聾患者的比較研究。結(jié)果在少數(shù)民族發(fā)現(xiàn)了11種突變形式,包括一種新的突變：c.257CG。少數(shù)民族共有202例患者發(fā)現(xiàn)突變或序列改變,純合突變75例,雜合突變92例,復(fù)合雜合突變35例。這些基因型的突變頻率為15.19%(202/1330),同地區(qū)漢族人群的突變頻率為19.04%(87/457)。少數(shù)民族群體與漢族的突變頻率之間沒有顯著性差異,比較各民族與漢族的GJB2基因突變頻率差異,其中維吾爾族與漢族存在統(tǒng)計(jì)學(xué)差異,其他各民族之間無統(tǒng)計(jì)學(xué)差異。各民族GJB2基因突變基因型以框移突變?yōu)橹?這類基因型決定了耳聾表型是以重度及極重度聾為多見。在少數(shù)民族人群中等位基因頻率最高的四種突變是c.235delC、c.35delG、 c.109GA和c.299-300delAT,藏族的熱點(diǎn)突變是c.235delC和c.109GA,東鄉(xiāng)族的熱點(diǎn)突變分別是c.299-300delAT和c.235delC,回族是c.235delC和c.109GA,哈薩克族是c.35delG,維吾爾族是c.235delC、c.35delG,漢族人群的熱點(diǎn)突變是c.235delC、c.109GA和c.299-300delAT,各民族的GJB2基因突變頻率與其種族背景相一致,其中哈薩克族和維吾爾族具有明顯的高加索背景表現(xiàn)。研究中我們繪制了各民族完整的GJB2基因突變頻譜,可以為針對(duì)各民族的個(gè)性化的基因診斷和大面積篩查提供方法學(xué)依據(jù)。另外考慮到基因的結(jié)構(gòu)特點(diǎn)和突變位點(diǎn)的分散性,GJB2基因診斷模式以全編碼序列的篩查為宜。第三章：SLC26A4基因及IntDNA1555AG突變?cè)谖鞅鄙贁?shù)民族非綜合征感音神經(jīng)性聽力損失患者的篩查研究 前庭水管擴(kuò)大是先天性耳聾最常見的內(nèi)耳畸形,與這一疾病相關(guān)的SLC26A4基因與耳聾關(guān)系密切。線粒體DNAA1555G突變與氨基糖苷類抗生素導(dǎo)致耳聾有相關(guān)性,該突變檢測(cè)有利于揭示藥物性聾和環(huán)境因素的關(guān)系。我們對(duì)西北地區(qū)少數(shù)民族共1330例非綜合征感音神經(jīng)性聾患者,以PCR及直接測(cè)序檢測(cè)SLC26A4基因第8、第19外顯子的突變,運(yùn)用PCR-RFLP篩查1ntDNA1555AG點(diǎn)突變。在共1330例少數(shù)民族患者中,篩查出SLC26A4基因c.919-2AG、c.2168AG、c.2162CT三種突變形式,檢測(cè)到七種基因型,包括純合26例,雜合40例,復(fù)合雜合突變5例,其中雙等位基因的突變頻率為2.33%。同期進(jìn)行的本地區(qū)漢族457例聾人檢測(cè)到6種基因型,包括純合15例,雜合31例及復(fù)合雜合突變8例,雙等位基因突變頻率為5.03%。分析顯示漢族同藏族、東鄉(xiāng)族及維族的該基因突變頻率有統(tǒng)計(jì)學(xué)差異,回族與東鄉(xiāng)族和維族均有差異。研究證實(shí)c.919-2AG突變是漢族聾人的熱點(diǎn)突變,這一突變?cè)跐h族與藏族、東鄉(xiāng)族及維吾爾族均存在統(tǒng)計(jì)學(xué)差異,維族與回族及哈薩克族有差異,回族與東鄉(xiāng)族存在差異。研究中檢測(cè)出mtDNA1555AG突變的患者有28例,突變頻率為2.11%,同地區(qū)457例漢族患者中突變者有32例,突變頻率為7.00%,兩者之間有顯著性差異。在106例AAID患者中有9例檢測(cè)到mtDNA1555AG突變,AAID患者中突變頻率為8.49%。各少數(shù)民族耳聾患者頻率比較顯示漢族與回族、東鄉(xiāng)族及維吾爾族之間的突變頻率存在統(tǒng)計(jì)學(xué)差異。研究證實(shí)SLC26A4基因在各少數(shù)民族與漢族的突變頻譜不同,熱點(diǎn)突變各異,因此需要針對(duì)各少數(shù)民族研究其突變頻譜,制定適合各民族的基因診斷策略和產(chǎn)前診斷方法。研究表明有必要在本地區(qū)少數(shù)民族耳聾人群中開展mtDNA1555AG突變的基因診斷,在散發(fā)耳聾患者、遺傳性耳聾家系開展該突變檢測(cè),進(jìn)行科學(xué)干預(yù)有助于延緩和減少藥物性耳聾的發(fā)生。 第四章常染色體顯性遺傳非綜合征耳聾家系的候選基因分析 遺傳性疾病的基因定位克隆方法多樣,需要根據(jù)遺傳資源的特征選擇合適的研究方法。本文旨在探索適宜于遺傳性耳聾病因?qū)W研究的有效方法。針對(duì)一個(gè)遺傳性耳聾大家系,我們系統(tǒng)進(jìn)行臨床資料和表型特點(diǎn)研究,并通過候選基因突變篩查方法,明確了該家系的致病基因。首先通過對(duì)家系中耳聾患者發(fā)病特點(diǎn)的分析,我們明確其遺傳方式為常染色體顯性遺傳�？偨Y(jié)本家系表型特點(diǎn)有：耳聾首先以高頻聽力損失為主,后期影響到全頻聽力,部分患者表現(xiàn)有前庭功能障礙的癥狀。通過對(duì)表型的詳盡分析,研究中將候選基因局限在少數(shù)幾個(gè)基因,進(jìn)而對(duì)最符合表型特點(diǎn)的COCH基因全部外顯子進(jìn)行突變篩查,發(fā)現(xiàn)了該基因第8外顯子c.485GA突變?yōu)橹虏⊥蛔?該突變與家系中遺傳性狀共分離。研究中發(fā)現(xiàn)該家系中聽力損失外顯率為100%,聽力損失隨年齡增長(zhǎng)而逐漸加重,前庭功能障礙的外顯率為22.22%,前庭障礙癥狀與聽力損失的程度沒有相關(guān)性。從聽力學(xué)特點(diǎn)和前庭功能障礙特征判斷,該家系與梅尼埃病的臨床表現(xiàn)不符。我們認(rèn)為遺傳性耳聾家系的基因定位克隆,首選位置克隆和候選基因克隆法,通過連鎖分析能夠定位耳聾相關(guān)基因的位置,候選基因克隆法具有經(jīng)濟(jì)快捷的優(yōu)勢(shì)。收集詳實(shí)的臨床資料和聽力學(xué)資料、充分分析家系臨床特征對(duì)于選擇適宜的研究方法是十分必要的。
[Abstract]:Hearing loss affects human speech function and leads to social communication disorders. Because of its high incidence, the first deafness gene has been cloned in 1995, and more than 80 non syndromic deafness related genes have been found. More and more deafness molecular causes have been revealed. The multi ethnic structure of Chinese population is rich in abundance. The rich genetic resources, especially the ethnic minority groups of different ethnic groups, are more precious. This study aims to collect the deafness resources of ethnic minorities in Northwest China and to establish a clinical resource and genetic resource bank covering the main ethnic minorities in Northwest China. On the basis of this, three common deafness related genes are carried out. The study of molecular epidemiology and the study of the candidate gene mutation of a deafness family, try to reveal the epidemic situation of these genes in the deaf people of ethnic minorities, draw the frequency spectrum of gene mutation of deaf disease in all ethnic groups, analyze the effective methods to carry out the gene location cloning of the deafness family, and explore the gene diagnosis strategy for the minority nationalities. The main contents of our research are composed of the following four parts: Chapter 1: the construction of the national minority deaf resource pool in Northwest China.
In order to protect the genetic deafness resources of ethnic minorities, we have studied and established the deafness Resource Bank of minority nationalities in Northwest China. In the study, we set up a standard process for the construction of the resource bank, through the local special education schools, all levels of the disabled and hospital outpatients, and so on. Channel collection of deafness sporadic cases and deafness family resources, questionnaire survey, physical examination, audiology examination and drawing of family map, sign informed consent and extraction of peripheral venous blood, extraction of genomic DNA, and establish a data management model of complete set of data. We collected 1673 cases of deafness and 146 heredity in Ethnic Minorities. The family of sexual deafness has completed the collection of clinical and genetic resources and accumulated resources for further genetic research. We think it is necessary to further strengthen the protection of the minority deafness resources and establish a scientific and perfect resource bank management system. The second chapter: the GJB2 gene mutation screening of the NNW non syndromic sensorineural hearing loss. Investigation and research
The GJB2 gene is the most common molecular cause of congenital deafness. The GJB2 gene encodes the gap junction protein Cx26, which plays an important role in the internal ear electrolyte exchange and intercellular information transmission, which is closely related to the human auditory function. We carry out the mutation of the GJB2 gene in the population of the sporadic sensorineural deafness of the minority nationalities in Northwest China. The purpose of this study was to analyze the epidemic situation of the gene, reveal the hot spot mutation of ethnic minorities and explore the genetic diagnosis strategy suitable for these deaf people. In this study, a total of 1330 cases of GJB2 gene mutation screening in a total of patients with sensorineural hearing loss were screened, and 457 cases of Han deafness in the same area were compared. Results 11 forms of mutation were found in ethnic minorities, including a new mutation: 202 cases of c.257CG. minorities found mutations or sequence changes, 75 homozygous mutations, 92 heterozygous mutations and 35 complex heterozygous mutations. The mutation frequency of these genotypes was 15.19% (202/ 1330), and the mutation frequency of the Han population in the same area was 19.04% (87). /457). There is no significant difference between the mutation frequency of the ethnic minority group and the Han nationality, compared with the GJB2 gene mutation frequency difference between the ethnic group and the Han nationality. There is a statistical difference between the Uygur and the Han nationality, and there is no statistical difference between the other ethnic groups. The mutation genotype of the GJB2 gene in each ethnic group is mainly based on the frame shift mutation, and this type of genotyping is determined. The deafness phenotype is more common with severe and extremely severe deafness. The four mutation of the highest allele frequency in minority groups is c.235delC, c.35delG, c.109GA and c.299-300delAT. The hot mutation of the Tibetan people is c.235delC and c.109GA. The hot mutations in the Dongxiang nationality are c.299-300delAT and c.235delC, and the Hui nationality is c.235delC and c.109GA, ha. The sak nationality is c.35delG, the Uygur nationality is c.235delC, c.35delG, the hot mutation of the Han population is c.235delC, c.109GA and c.299-300delAT. The mutation frequency of the GJB2 gene of each ethnic group is in accordance with its racial background. The Kazakh and Uygur ethnic groups have obvious Caucasus back scenery. In the study, we draw the complete GJB2 of the ethnic groups. The gene mutation spectrum can provide a methodological basis for individual genetic diagnosis and large area screening for various nationalities. In addition, considering the structural characteristics of the gene and the dispersity of the mutation sites, the GJB2 gene diagnosis model is suitable for the screening of the full coding sequence. The third chapter: the SLC26A4 gene and the IntDNA1555AG mutation in the northwest minority nationality. Screening study of non syndromic sensorineural hearing loss
The enlargement of the vestibular water pipe is the most common deafness of the congenital deafness. The SLC26A4 gene related to this disease is closely related to the deafness. The mitochondrial DNAA1555G mutation is associated with the deafness caused by aminoglycoside antibiotics. The mutation detection is beneficial to reveal the relationship between drug deafness and environmental factors. In 1330 patients with sensorineural hearing loss, SLC26A4 gene eighth, nineteenth exon mutations were detected by PCR and direct sequencing, and 1ntDNA1555AG point mutation was screened by PCR-RFLP. Among the 1330 ethnic minority patients, the SLC26A4 gene c.919-2AG, c.2168AG, c.2162CT were screened and seven genotypes were detected, including the homozygous 26. There were 40 heterozygosity and 5 complex heterozygosity mutations. The mutation frequency of the double allele was 6 genotypes in 457 cases of the Han nationality in the local Han nationality of 2.33%., including 15 homozygous, 31 heterozygosity and 8 heterozygosity mutations. The mutation frequency of the double allele was 5.03%. analysis of the Han nationality, the Dongxiang and the Uygur. There is a statistical difference between the frequency of the mutation and the difference between the Hui and the Dongxiang and the Uygur people. The study confirms that the c.919-2AG mutation is a hot mutation of the Han and the deaf people. This mutation is statistically different in the Han and Tibetan, Dongxiang and Uygur ethnic groups. There are differences between the Uygur and the Hui and Kazakh people, and there are differences in the Hui and Dongxiang ethnic groups. The study detected M There were 28 cases of tDNA1555AG mutation, the frequency of mutation was 2.11%, and there were 32 cases of mutation in 457 Han patients in the same area, and the mutation frequency was 7%. There was a significant difference between the two. 9 cases in 106 cases of AAID were detected mtDNA1555AG mutation, and the frequency of mutation in AAID patients was 8.49%. of the minority deafness patients compared with the Han and the Han people. The mutation frequency between the Hui nationality and the Uygur ethnic group is statistically different. The study confirms that the mutation frequency of the SLC26A4 gene is different in the ethnic minority and the Han nationality, and the mutation frequency is different. Therefore, it is necessary to study the mutation spectrum of the ethnic minorities and formulate the genetic diagnosis strategies and prenatal diagnosis methods suitable for each ethnic group. The study shows that the SLC26A4 gene is suitable for all ethnic minorities. It is necessary to carry out the genetic diagnosis of mtDNA1555AG mutation among the deafness people in the local minority, and to carry out the mutation detection in the patients with deafness and hereditary deafness, and to intervene scientifically can help delay and reduce the occurrence of drug-induced deafness.
Fourth chapter analysis of candidate genes in families with autosomal dominant hereditary nonsyndromic hearing loss
The gene location and cloning methods of genetic diseases are diverse and need to be selected according to the characteristics of genetic resources. This paper aims to explore an effective method suitable for the etiological study of hereditary deafness. Through the analysis of the characteristics of the deafness in the family, it is clear that the hereditary mode of the family is autosomal dominant. The phenotypic characteristics of the family are as follows: the hearing loss is mainly high frequency hearing loss, the latter affects the full frequency hearing, and some patients show the vestibular function barrier. After a detailed analysis of the phenotype, the candidate genes were limited to a few genes, and then all the exons of the COCH gene, the most conforming to the phenotypic characteristics, were screened, and the mutation of the eighth exon c.485GA was found to be a pathogenic mutation, and the mutation was separated from the genetic traits in the family. The hearing loss was 100%, the hearing loss increased with age, and the vestibular dysfunction was 22.22%. The vestibular disorder was not related to the degree of hearing loss. Judging from the characteristics of the hearing and the characteristics of the vestibule dysfunction, the family was not in accordance with the clinical manifestation of the disease. We thought hereditary ear was inherited. The gene location cloning of the deafness family, the preferred location clone and the candidate gene cloning method, can locate the position of the deafness related genes through linkage analysis. The candidate gene cloning method has an economical and quick advantage. The detailed clinical data and audiological data are collected. Necessary.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R764.43

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本文編號(hào)：1970205