五例新生儿肝内胆汁淤积症患儿基因的突变分析CSCD

目的明确5例新生儿肝内胆汁淤积症患儿的分子机制。方法应用新一代测序技术对患儿的SLC25A13基因进行外显子捕获检测,对突变位点进行Sanger测序验证。用PolyPhen-2软件对新突变的致病性进行分析。结果 5例患儿均携带SLC25A13基因的复合突变,共发现8个突变位点,其中2个既往未见报道(c.1357A>G和c.1663dup23)。5例患儿的父母均为突变携带者。结论 SLC25A13基因的突变可能是5例患儿的发病原因,所携带的突变以851del4和1638-1660dup为主。新发现的c.1357A>G和c.1663dup23突变丰富了SLC25A13基因的突变谱。     

原发性肉碱缺乏症一家系的SLC22A5基因突变检测与产前诊断

目的对原发性肉碱缺乏症(PCD)病患者及一家系进行致病基因SLC22A5突变鉴定,为家系提供遗传咨询和产前诊断,同时研究c.394-1 GT突变所导致的SLC22A5基因剪接形式的改变。方法收集患者及父母的外周血标本,提取基因组DNA,采用Sanger法对家系中各成员进行SLC22A5基因的所有外显子进行测序,同时对发现的突变在100名正常对照中进行测序,以排除多态性位点;使用QIAGEN血液RNA提取试剂盒抽提患儿总RNA,通过RT-PCR和测序技术分析c DNA序列特征。对先证者母亲的羊水标本进行产前诊断。结果 Sanger法DNA测序检测出该家系中先证者为SLC22A5基因的两个杂合突变:c.394-1 GT,c.760CT(p.R254X),在100名正常对照中均未发现这两个位点的突变。先证者父亲、母亲分别携带SLC22A5基因c.394-1 GT、c.760 CT(p.R254X)杂合突变。先证者母亲的羊水标本的基因型与先证者一样。先证者的特定片段RT-PCR产物电泳时较非携带者的正常人多出一条小的亮带,经测序确认是由c.394-1 GT突变导致SLC22A5基因c DNA外显子2完整序列的缺失。结论 c.394-1 GT突变位点在国内外仅一例报道,本研究证实该突变的存在,同时证明该突变导致SLC22A5基因c DNA外显子2完整序列的缺失。这一发现丰富了SLC22A5基因突变谱,为原发性肉碱缺乏症(PCD)病的防治提供新的参考,Sanger测序等技术可为原发性肉碱缺乏症家系提供遗传咨询和产前诊断服务。     

遗传性出血性毛细血管扩张症基因突变分析

目的 分析遗传性出血性毛细血管扩张症(hereditary hemorrhagic telangiectasia,HHT)家系ENG、ACVEL1和SMAD4基因突变.方法 收集4个HHT家系临床资料并分析其临床特点,应用直接测序和多重连接探针扩增技术对11例临床确诊及可疑患者的ENG、ACVRL1和SMAD4基因进行突变分析,将结果与HHT基因突变数据库进行对比.结果 家系2先证者及2个妹妹的ENG基因发生了第2外显子c.207G＞A(p.L69L)同义突变、第8外显子c.1004A＞T(p.Q335L)错义突变、ACVRL1基因第7外显子c.817C＞T(L273L)同义突变;家系3先证者及其母亲和弟的ENG基因发生了第8外显子c.1004A＞T(p.Q335L)突变;也检测到家系4先证者及其兄的ENG基因第8外显子c.1004A＞T(p.Q335L)突变.家系1先证者及其他HHT患者,未检测到基因突变.其中ENG基因第8外显子c.1004A＞ T(p.335Q＞L)为新突变,在200名正常对照中也未检测到该突变.结论 HHT具有遗传异质性,ENG基因第8外显子c.1004A＞T(p.Q335L)为HHT新的致病突变.     

GJB2基因p.V37I突变致病性及家系临床表型分析

目的探讨GJB2基因突变耳聋家系p.V37I(c.109GA)突变致病性及分析家系患者临床表型。方法收集6个GJB2基因p.V37I(c.109GA)突变耳聋患者及家系的临床资料及外周血样本,运用PCR产物直接测序技术对家系耳聋患者进行GJB2、GJB3基因编码区、SLC26A4基因外显子7和8、以及线粒体m.1494CT、m.1555AG位点检测分析,另对家系5先证者行高通量全外显子序列检测。结果 6个家系均检出GJB2基因p.V37I突变,其中家系3、5为纯合突变,家系1、2、6先证者另复合GJB2基因c.235del C杂合突变,家系4患者另检出c.299-300del AT杂合突变;全部家系均未检出GJB3基因编码区、SLC26A4基因外显子7和8、以及线粒体m.1494CT、m.1555AG位点突变,家系5先证者高通量全外显子序列检测结果亦提示GJB2基因p.V37I纯合突变为其致聋原因。结论 GJB2基因p.V37I突变具有一定致病性,该位点纯合突变可导致轻度至中度听力损失,而p.V37I突变复合c.299-300del AT、c.235del C杂合突变可导致中度至重度感音神经性耳聋。     

糖原累积病Ⅰ b型1例的临床和分子遗传分析

目的初步探讨1例糖原累积病Ⅰb型(GSDⅠb)患者的临床特点和致病基因,分析该疾病发生的分子遗传机制。方法收集患者临床资料,抽提患者外周血白细胞基因组DNA,通过多聚酶链反应扩增葡萄糖-6-磷酸酶转位酶基因SLC37A4的9个外显子,用DNA直接测序法确定其突变位点。结果患者临床表现及实验室检查完全符合GSDⅠb。经PCR测序发现SLC37A4基因第3外显子572位碱基C→T纯合突变(c.572 C>T),造成第191位的脯氨酸被亮氨酸替代(p.P191L),导致葡萄糖-6-磷酸转位酶活性下降。结论 SLC37A4基因突变导致的葡萄糖-6-磷酸转位酶结构改变是该GSDⅠb患者临床表现的分子遗传基础。P191L纯合突变在中国大陆的报道尚属首次。相信不久DNA突变分析将会成为糖原累积病Ⅰb型的主要确诊方法。     

糖原累积病I b型1例的临床和分子遗传分析

目的 初步探讨1例糖原累积病I b型(GSD I b)患者的临床特点和致病基因,分析该疾病发生的分子遗传机制.方法 收集患者临床资料,抽提患者外周血白细胞基因组DNA,通过多聚酶链反应扩增葡萄糖-6-磷酸酶转位酶基因SLC37A4的9个外显子,用DNA直接测序法确定其突变位点.结果 患者临床表现及实验室检查完全符合GSD I b.经PCR测序发现SLC37A4基因第3外显子572位碱基C→T纯合突变(c.572 C>T),造成第191位的脯氨酸被亮氨酸替代(P.P191L),导致葡萄糖-6-磷酸转位酶活性下降.结论 SLC37A4基因突变导致的葡萄糖-6-磷酸转位酶结构改变是该GSD I b患者临床表现的分子遗传基础.P191L纯合突变在中国大陆的报道尚属首次.相信不久DNA突变分析将会成为糖原累积病I b型的主要确诊方法.     

变性高效液相色谱法分析非综合征型耳聋人群SLC26A4基因突变

目的 研究非综合征型耳聋(nonsyndromic hearing loss,NSHL)患者SLC26A4基因的突变情况,为临床上NSHL患者基因诊断提供指导.方法 PCR分别扩增SLC26A4基因的21个外显子及其侧翼序列,所得目的 片段用变性高效液相色谱(denaturing high-performance liquid chromatorgraphy,DHPLC)进行突变筛查,有异常峰形的样本进行DNA测序.结果 在所选30例无血缘关系且GJB2基因检测未发现突变的NSHL患者中,共检测出10种SLC26A4基因变异,其中包括7种已知突变,2种未见报道的新突变(F572L和D87Y),及一种已知多态(Ivs11+47T＞C),其中Ivs7-2A＞G是最常见的突变,约占总突变的40%.结论 SLC26A4基因为仅次于GJB2的导致NSHL的相关基因,在(GJB2基因检测未发现突变的NSHL人群中SLC26A4基因的检出率达到23.3%,其中Ivs7-2A＞G是其最常见的突变.     

感音神经性耳聋患者大前庭导水管综合征相关SLC26A4基因IVS7-2A＞G的全序列分析

目的 对携带SLC26A4基因IVS7-2A＞G单杂合突变感音神经性耳聋患者进行 SLC26A4基因的全序列检测,以期发现除IVS7-2A＞G以外的其他突变.方法 应用直接测序法对80例携带IVS7-2A＞G单杂合突变的感音神经性耳聋患者进行SLC26A4基因进行全序列测序.结果 80例患者中47例发现另1个突变位点,其余33例未发现复合杂合突变,IVS7-2A＞G单杂合突变找到另外1个突变的比例为58.8%(47/80).发现了 3个新的突变,分别是5+2T＞A、14-2A＞G和1825del G,最为常见的5种突变为H723R(20%)、T410M(5%)、15+5G＞A(5%)、L676Q(5%)、N392Y(3.75%).第17外显子是突变发生种类最多的外显子.结论 SLC26A4基因IVS7-2A＞G单杂合突变者应该进行其他突变的筛查,SLC26A4基因复合突变可以解释部分的耳聋原因.     

一例17α-羟化酶/17,20-碳链裂解酶缺陷症患儿的CYP17A1基因突变分析

目的 对1例17α-羟化酶/17,20-碳链裂解酶缺陷症(17α-hydroxylase/17,20-lyase deficiency,17OHD)的患儿进行CYP17A1基因突变分析,同时分析中国17OHD患者的CYP17A1基因突变特点.方法 收集患者的临床资料,应用PCR和DNA测序技术对患者进行CYP17A1基因的突变检测.结果 患者临床表现较典型:高血压、低血钾、性激素和皮质醇明显低于正常、促肾上腺皮质激素升高.患者的CYP17A1基因第6外显子上发现c.987C＞A(p.Y329X)以及c.985del(p.Y329TfsX89)复合杂合突变,分别生成包含328和417个氨基酸的截短蛋白.结论 在患者的CYP17A1基因上发现了c.985del和c.987C＞A复合杂合突变.我国患者中最常见类型为错义突变,且集中于第6和第8外显子.     

一种新复合杂合突变导致21-羟化酶缺陷症

目的探讨1例单纯男性化型21-羟化酶缺陷症(21-OHD)基因突变的类型和特点及临床表型与基因突变类型之间的关系。方法收集患者的临床资料,提取外周血白细胞DNA,用PCR方法扩增CYP21A2基因的10个外显子及内含子边界,测序鉴定CYP21A2基因突变位点,进一步分析突变位点与临床表型的关系。结果患者的临床表现主要为外阴发育异常。基因测序结果显示为复合杂合突变,其一个等位基因为c.515 T>A,p.I172N,另一个等位基因为c.593 T>G,p.L198X,此种复合杂合突变主要引起单纯男性化表现。p.L198X是至今尚未见报道的一种新突变。结论发现了CYP21A2基因一种新的突变p.L198X,丰富了CYP21A2基因突变数据库。同时从分子遗传学方面证实了对患者的诊断,患者基因型能很好地解释其临床表现。     

A Founder Effect of c.257 + 2T &gt; C Mutation in <Emphasis Type="Italic">NCF2</Emphasis> Gene Underlies Severe Chronic Granulomatous Disease in Eleven Patients

Chronic granulomatous disease (CGD) is the prototypic functional neutrophil disorder caused by genetic defects in one of the five genes encoding the superoxide-generating nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase subunits of phagocytes. Mutations causing the most prevalent form of CGD in western populations are located in the X-linked-CYBB gene. The four remaining autosomal recessive (AR) forms collectively account for one-third of CGD cases. We investigated the clinical and molecular features of eleven patients with CGD from 6 consanguineous families, originating from contiguous regions in the west of Tunisia. The patients’ clinical phenotype is characterized by a high incidence of mycobacterial infections. Five out of the eleven patients died despite treatment arguing in favor of a severe clinical form of CGD. These findings correlated with the absence of functional p67phox protein as well as the absence of residual reactive oxygen intermediates (ROI) production. Genetic analysis showed the presence, in all patients, of a unique mutation (c.257 + 2T > C) in NCF2 gene predicted to affect RNA splicing. Segregating analysis using nine polymorphic markers overlapping the NCF2 gene revealed a common haplotype spanning 4.1 Mb. The founder event responsible for this mutation was estimated to have arisen approximately 175 years ago. These findings will facilitate the implementation of preventive approaches through genetic counseling in affected consanguineous families.     

1例Wiskott-Aldrich综合征临床特征和致病基因突变分析

目的:分析1例Wiskott-Aldrich综合征患者的临床特征及分子遗传特征。方法:分析患者的临床特征,收集患者及其父母外周血,提取基因组DNA,针对WASP基因所有编码外显子及外显子和内含子交界处进行PCR扩增测序。结果:我们报道的患者具有典型的WAS表型,临床得分为5分;患儿Coomb’s试验,自身抗体中ANA及类风湿因子均阳性,伴有自身免疫性疾病。患儿为WASP基因第7外显子中第665位核苷酸C突变为T(c.665C>T),导致211位密码子发生无义突变,该位置提前出现终止密码(p.R211X);其母亲为此突变基因携带者。结论:这例男性中国Wiskott-Aldrich综合征患儿由于WASP基因突变致病,此基因型患儿(p.R211X)拥有典型的WAS表型且伴有自身免疫性疾病的临床型为国内首次报道。     

Familial tumoral calcinosis and testicular microlithiasis associated with a new mutation of GALNT3 in a white family

BACKGROUND: Familial tumoral calcinosis (FTC) is a rare autosomal recessive disease characterised by the development of multiple calcified masses in periarticular soft tissues; GALNT3 gene mutations have recently been described in an African American and in a Druse Arab family with FTC. OBJECTIVE: To report the clinical and histological features caused by a new GALNT3 mutation in a white family. RESULTS: Homozygosity for the nonsense mutation Lys463X was found in both affected siblings, who displayed a classic phenotype, the male also having testicular microlithiasis. He is the first subject described with testicular microlithiasis in FTC. CONCLUSIONS: The high testicular expression of GALNT3 suggests that the gene alteration could act locally by causing deposition of calcium, and the testis may be an underestimated site of calcification in FTC. Autoimmune diseases are present in several members of the family. Although immune disorders have been described in FTC, autoimmunity does not segregate with the GALNT3 mutation in this family.     

一例皮肤松弛症患儿的临床与遗传学分析CSCD

目的采用遗传病相关基因外显子测序的方法确定1例皮肤松弛症患儿的致病基因,并对其相关的临床表型及基因型进行总结。方法收集先证者及其父母的临床资料,首先对先证者进行遗传病相关基因的外显子测序,确定了可能的致病突变,对先证者及其父母进行Sanger测序验证,确定基因突变位点。结果先证者存在ATP6VOA2基因C．187C〉T（P．R63X）杂合突变和C．1189G〉C（P．A397P）杂合突变,先证者父母分别携带ATP6VOA2基因c．1189G〉c（P．A397P）杂合突变和C．187C〉T（P．R63X）杂合突变。结合患儿临床表型,诊断为常染色体隐性遗传皮肤松弛症2A型（autosomal recessive cutis laxa type2A,ARCL2A）。结论通过遗传学方法确诊了ARCL2A型患儿,总结了ARCL2A患儿的典型临床特征,新突变的发现扩大了ATP6VOA2基因的突变谱。外显子测序可作为诊断复杂遗传病致病基因的重要工具。     

Clinical,histopathological and genetic studies in a case of fatal familial insomnia with review of the literature

To explore clinical, histopathological and genetic features of a case with fatal familial insomnia (FFI) and review the related literatures. A middle-aged woman who complained of “insomnia for 9 months and psychosis for 3 months” was suspicious of FFI. The clinical features of the patient were analyzed, and the dead patient was examined by autopsy and the brain tissues were obtained for histopathological studies, and the blood samples from the patient and some of her familial members were collected for the sequencing of prion protein gene (PRNP). The main clinical features included intractable insomnia, psychiatric symptoms and abnormal night sleep behavior, unsteady gait, difficulty swallowing, sudden death, and positive family history. The pathological studies showed neuronal loss and gliosis of multiple brain tissues in the proband, predominated with thalamus; and analysis of PRNP revealed gene D178N mutation, and linkage with 129 methionine (Met) allele in the proband and a relative. FFI patients may manifest as sudden death, and may have prominent psychiatric symptoms; the corresponding gene mutation could occur in the asymptomatic carriers; the data of autopsy and brain tissue pathology is helpful for further understanding of this disease.     

一个遗传性牙本质发育不全家系的分子遗传研究

目的 研究一个常染色体显性牙本质发育不全家系发病的遗传基础.方法 通过对一个DSPP家系临床检查和家族史调查,连锁分析和DSPP基因的突变检测,以及限制性片段长度多态分析方法,分析该家系发病的分子基础.结果 连锁分析发现,该疾病致病基因与微卫星标记D4S1534完全连锁,对位于该区域的DSPP基因进行测序分析,发现一个新的致病突变(c.49C→T,p.Pro17Ser),该突变位于DSPP基因的第1外显子.该家系所有患者中都检测到了这一致病突变,但家系中的正常个体和100个无亲缘关系的正常人中未发现这个突变.结论 p.Pro17Ser是牙本质发育不全Ⅱ型致病基因DSPP的一个新的致病突变.我们的研究进一步拓展了对牙本质发育不全疾病分子遗传基础的认识.     

一个先天性小眼畸形家系的全外显子组测序分析及产前诊断

目的分析1个先天性小眼畸形家系的临床表型及遗传学病因。方法应用高通量测序技术对先证者及其父母进行全外显子组测序,筛选候选致病位点,对其家系进行Sanger测序验证,并通过羊水穿刺和Sanger测序为先证者母亲提供产前诊断。结果全外显子组测序和Sanger测序发现家系中的3例患者均携带OTX2基因c.289C>T(p.R97*)杂合变异,先证者母亲亦携带该变异,但无小眼畸形。先证者的父亲、舅母和胎儿未携带上述变异。结论OTX2基因c.289C>T(p.R97*)杂合变异很可能是该家系的发病原因。上述诊断将有助于该家系的遗传咨询和产前诊断。     

Analysis of MAT1A gene mutations in a child affected with simple hypermethioninemia北大核心CSCD

Objective To detect potential mutations of MATl A gene in a child suspected with simple hypermethioninemia by MS/MS neonatal screening. Methods Clinical data of the child was collected. Genomic DNA was extracted by astandard method and subjected to targeted sequencing using an Ion Ampliseq™ Inherited Disease Panel. Detectedmutations were verified by Sanger sequencing. Results The child showed no clinical features except evaluatedmethionine. A novel compound mutation of the MATIA gene, i. e. , c. 345delA and c. 529C>T, was identified in the child. His father and mother were found to be heterozygous for the c. 345delA mutation and c. 529C > Tmutation, respectively. Conclusion The compound mutation c. 345delA and c. 529C>T of the MATIA gene probably underlie the disease in the child. The semi-conductor sequencing has provided an important means for thediagnosis of hereditary diseases.     

临床病毒性脑炎标本的博尔纳病病毒核酸检测

目的 探讨临床病毒性脑炎(viral encephalitis,VE)患者与博尔纳病病毒(Borna disease virus,BDV)感染的关系,分析BDV感染的病毒性脑炎患者临床特征.方法 用荧光定量巢式逆转录聚合酶链反应(FQ-nRT-PCR)方法检测病毒性脑炎患者及非感染性疾病施行蛛网膜下腔阻滞麻醉的手术患者脑脊液单个核细胞(cerebrospinal fluid mononuclear cell,CSFMC)中BDV p24基因片段,同时用β-肌动蛋白(β-actin)作为内参照,脑脊液(CSF)阳性标本基因测序分析并总结出临床特征.结果 32例病毒性脑炎脑脊液标本BDV p24基因片段检出率为12.5%(4/32),拷贝数＞102/μl.对其中一份CSF阳性标本测序后,与BDV标准病毒株Ⅴ和马源的BDV病毒株H1766序列比较同源性分别为95.35%和98.84%.在4个位点出现基因突变(nt1649 T→C、nt1656 G→A、nt1670 C→T、nt1676 C→T).该目的基因片段与马源的BDV病毒株亲缘关系最近.阳性脑炎患者主要以精神行为异常为临床特征.结论 贵州省遵义市部分病毒性脑炎的发生与BDV感染有关,主要以精神行为异常为临床特征.     

A familial frontotemporal dementia associated with C9orf72 repeat expansion and dysplastic gangliocytoma

A hexanucleotide repeat expansion in the chromosome 9 open reading frame 72 gene (C9orf72) was recently identified as the most common genetic cause of frontotemporal dementia/amyotrophic lateral sclerosis. Here we describe the clinical, pathologic, and genetic features of a Finnish C9orf72 expansion carrier, who developed a dysplastic gangliocytoma (Lhermitte-Duclos disease), a rare hamartoma/overgrowth syndrome of cerebellar granule cells associated with mutations in the phosphatase and tensin homolog gene. In addition to the dysplastic gangliocytoma, the patient showed typical transactive response DNA-binding protein with M_r 43 kD (TDP-43) pathology mainly in the cortex and the substantia nigra and numerous p62-positive/TDP-43-negative inclusions in the cerebellar granule cells. His sister carried the same gene defect and showed a similar type of TDP-43/p62 pathology in her brain. Our findings confirm that the clinical and pathologic picture of C9orf72 mutation carriers is more heterogeneous than originally thought and warrants further studies on the possible involvement of phosphatase and tensin homolog gene pathway in the specific cerebellar granule cell pathology associated with C9orf72 expansion.