GNAS新错义突变导致D/E型短指畸形

目的对我国一个短指家系进行致病基因进行鉴定。方法应用全外显子测序对该短指家系进行基因突变检测,并用Sanger测序对突变位点进行验证。结果该家系共有3名患者,均为短指表型,符合常染色体显性遗传。全外显子检测提示患者均存在GNAS基因(NM_001077488:exon13:c.A1145T:p.D382V)杂合错义突变,Sanger测序验证与全外显子结果一致。结论 GNAS基因第13号外显子上c.A1145T杂合错义突变是该短指家系的致病原因。     

近亲婚配后代卵巢早衰姐妹的致病基因研究

先证者为两位非孪生姐妹,均因卵巢早衰、婚后不孕就诊。其父母为姑表近亲结婚。进行现存家族成员外显子测序,Sanger测序方法对家系内成员进行共分离。筛选出两个可能致病位点,分别位于基因AHDC1和PTPN14上;共分离验证显示,基因AHDC1c.C874T和PTPN14c.G3344A与表型共分离。基因AHDC1和PTPN14可能造成该家系成员罹患特发性卵巢早衰,可能为特发性卵巢早衰的候选基因。     

一个多发性骨骺发育不良大家系的临床特点及致病基因分析

目的通过对一个5代疑似多发性骨骺发育不良(multiple epiphyseal dysplasia,MED)的大家系(患者17例)进行临床特征分析和致病基因的筛查,为遗传咨询和产前分子诊断提供实验依据。方法采集家系成员病史,一般体检、关节、髋部X线片资料;收集该家系外周血样,提取样本DNA,靶向基因高通量测序方法对先证者DNA临床全外显子进行测序,使用Next Gene软件对测序序列进行比对分析,并进一步利用Ingenuity软件对存在的突变进行功能注释,寻找先证者致病突变。针对可疑突变,PCR和Sanger测序对家系其他成员DNA样本进行验证。结果该家系共5代,现存家系成员38人,系谱分析符合常染色体显性遗传特征。家系共有患者17例,其临床表现为:幼时出现走路姿势异常,后出现髋关节及膝关节疼痛,X线有典型骨骺发育不良病理改变。高通量测序及数据分析后,筛选出先证者(Ⅳ-3)软骨低聚物基质蛋白(cartilage oligomeric matrix protein,COMP)基因c.1153G>A(p.Asp385Asn)错义杂合突变,该突变导致其编码蛋白的第385位天冬氨酸被天冬酰胺替代。先证者家系其他成员符合基因型与表型共分离。结论COMP基因c.1153G>A错义杂合突变是导致该MED家系患者发病的分子机制,该突变首次在大家系中被报道,进一步明确了COMP基因c.1153G>A突变的致病性,有利于家系患者的进一步的诊治,也为产前诊断提供了实验依据。     

两个遗传性并多指(趾)家系致病基因突变分析

目的探讨2个并多指(趾)畸形(SPD)家系的致病基因。方法收集2019年1月、2020年12月就诊于临沂市人民医院的2个SPD家系的临床资料, 采集先证者及家系成员静脉血样本, 提取基因组DNA, 对先证者行全外显子组测序筛选候选基因变异;采用Sanger测序对2个家系成员验证其突变位点;采用生物信息学软件PolyPhen-2和PROVEAN对突变位点的致病性进行预测分析, 结合美国医学遗传学与基因组学学会(ACMG)指南对突变位点进行致病性判断。结果家系1三代成员中共有5例患者(男2例、女3例), 先证者为8岁女性, 表现为右手第3、4指并指, 指蹼融合和远端指甲融合, 其余手指活动自如, 双脚未见异常;家系2三代成员中共有4例患者(均为女性), 先证者为4岁女性, 表现为双手第3、4指并指, 示指侧弯。全外显子组测序分别在2个SPD家系中检出同源盒D13(HOXD13)基因c.917G>A和c.917G>T突变, 且2个突变均呈现基因型-表型共分离, 其中HOXD13基因c.917G>T突变未见数据库收录, 为新发杂合错义突变。生物信息学软件预测这2个突变位点均为...     

1例隐睾症家系的外显子测序分析

目的:对收治的1个隐睾症家系进行全外显子测序(WES),筛选并分析该家系中与隐睾症发生相关的突变位点,为该疾病的遗传学病因提供证据。方法:收集本中心收治的一对同时患有单侧隐睾且存在家族史的同卵双生双胞胎兄弟及其父母的血液DNA样本,对该家系进行NextSeq500 PE150测序,以GATK、VarScan软件对SNP和InDel进行低频变异检测,并关联多个数据库(dbSNP、1000g、ESP6500、HGMD、OMIM等)对变异结果进行注释。结果:基因组分析显示,双胞胎患儿中,ROR2基因(c.2249G>A.p.G750D)存在杂合子非同义变异,变异来源为父亲。AR基因(c.1625G>A;p.R542Q)存在半合子非同义变异,变异来源于母亲。这两处变异在基因数据库中较为少见,预测与隐睾相关。结论:ROR2基因突变(c.2249G>A.p.G750D)与AR基因突变(c.1625G>A;p.R542Q)可能是该家系发生隐睾症的遗传学病因。     

常染色体显性遗传Alport综合征致病基因 COL4A4杂合剪接突变的致病性分析及基因型-表型关联分析

目的探讨常染色体显性遗传Alport综合征(autosomal dominant Alport syndrome, ADAS)致病基因COL4A4杂合剪接突变的致病机制及基因型与表型的关联, 以加深对COL4A4剪接突变的认识以及对ADAS表型异质性的理解。方法本研究为病例系列分析。从3家医院收集5个ADAS家系先证者及家系成员的临床资料。对从先证者中经全外显子组测序(whole exome sequencing, WES)发现的COL4A4杂合剪接变异, 通过RNA体内剪接或Minigene体外实验分析其对mRNA正常剪接的影响。结果在此5个ADAS家系患者中经WES发现了4个COL4A4杂合剪接变异位点。家系1、家系2、家系3和家系4中多数患者呈孤立性镜下血尿或合并微量蛋白尿, 个别患者合并显性蛋白尿、进入老年期后出现肾功能轻度减退。家系5中4例患者均呈快速肾功能进展, 于28～41岁进展至终末期肾病。家系1、家系2患者携带的c.735+3A>G和家系3患者携带的c.694-1G>C均可引起COL4A4的第12号外显子跳跃导致42 bp核苷酸框内缺失(c.694     

全外显子组测序发现中国Joubert综合征家系C5orf42基因的新突变

目的探究一个中国Joubert综合征家系的致病基因及位点。方法对该Joubert综合征家系的先证者进行全外显子组测序,按照ACMG解读规则筛选致病性变异位点,利用聚合酶链式反应(PCR)和Sanger测序进行验证,并在其他家庭成员及192例健康对照者中进行测序验证。结果在先证者的5号染色体的C5orf42基因上存在2个杂合突变位点,分别为c.3676CT,p.R1226X以及c.8310_8311insT,p.L2770fsX5,这两个位点在家系中符合遗传共分离规律。结论全外显子组测序结合Sanger测序发现C5orf42基因的c.3676CT,p.R1226X以及c.8310_8311insT,p.L2770fsX5位点为引起该Joubert综合征家系临床病变的突变位点。     

FSHR剪切变异c.299+2T>G致卵巢抵抗综合征的家系分析

目的 探讨卵巢抵抗综合征(ROS)不孕患者的遗传学病因,为临床遗传咨询及助孕方案提供诊疗依据。方法 收集先证者家系临床资料及遗传学检测指标,绘制家系图谱,进行染色体G显带核型分析,应用高通量基因测序技术(NGS)进行全外显子组测序;对先证者父母与兄长的变异位点进行Sanger测序验证,并进行生物信息学分析及诊断。结果 该女性先证者临床诊断为原发性闭经、ROS;先证者母亲的祖母与父亲的祖父为亲兄妹。全外显子组测序及Sanger测序检测到先证者存在卵泡刺激素受体(FSHR)基因经典剪切变异NM＿000145.4 c.299+2T>G(纯合型),且该变异为未报道的致病性变异;其兄长该位点突变与先证者一致,其父母均存在FSHR剪切变异c.299+2T>G(杂合型)。结论 FSHR新发现的剪切变异c.299+2T>G的检出,丰富了FSHR基因突变谱,为ROS的临床治疗和家系成员的遗传咨询提供了参考。     

局灶节段性肾小球硬化患者致病基因突变检测

目的对散发性或家族性局灶节段性肾小球硬化（focal segmental glomerulosclerosis,FSGS）患者进行FSGS致病基因热点突变进行筛查,了解这些热点突变在我国FSGS患者的发生情况。方法研究对象为我院经肾脏活检确诊的40例FSGS散发病例及一个22人的FSGS家系LF-01。收集散发病例的外周血,采用盐析法提取基因组DNA;对LF-01家系进行调查,收集实验室检测数据,并留存外周血提取基因组DNA。对40例散发病例及所有家系成员进行ACTN4第8外显子,TRPC6第2、5、12、13外显子和INF2基因2、3、4外显子筛查,通过PCR扩增外显子后,直接测序进行基因突变检测。结果LF-01家系共9人为患病或可能患病状态,该家系表现为不完全外显遗传模型。40例散发FSGS患者,平均发病年龄37岁,男女比例为26：14,其中22例患者临床表现为肾病综合征。我们对所有样本进行了ACTN4第8外显子,TRPC6第2、5、12、13外显子和INF2基因2、3、4外显子筛查,均未发现有已知的基因突变。结论国外报道的FSGS致病基因的热点突变ACTN4、TRPC6和INF2可能不是中国汉族人群FSGS的致病基因。     

迟发性家族性局灶节段肾小球硬化的足细胞分子基因突变研究

目的 了解迟发性家族性局灶节段肾小球硬化（FSGS）的足细胞分子基因致病突变特点。 方法 研究对象为上海瑞金医院肾脏科1997年9月至2007年10月收集的31个迟发性家族性FSGS家系。诊断标准：（1）成员年龄大于12岁；（2）1个家系中有2例或2例以上患者经肾活检证实为FSGS，或家系成员中有1例肾活检证实为FSGS，另有1例成员有蛋白尿或肾功能不全。100例健康人为对照组。外周血基因组DNA 经PCR扩增后直接对NPHS2、ACTN4、TRPC6基因行测序分析。 结果 发现ACTN4基因新错义突变L316P，该家系患病成员起病年龄平均（38.7±7.4）岁，肾功能损害进展相对缓慢，家系3例患病成员均为突变杂合子。发现TRPC6基因新杂合错义突变Q889K，该家系患者起病年龄平均（38.0±4.2）岁，肾功能损害进展也较缓慢，家系中临床表现存在个体差异，家系中3例患病成员均为突变杂合子。发现TRPC6静止突变G467G。所有家系中未发现NPHS2致病突变。健康对照组200条染色体亦未发现以上突变。 结论 在31例迟发性FSGS家系中发现2个家系携带致病相关突变：ACTN4新突变L316P和TRPC6新突变Q889K。在中国人群家族性迟发性FSGS中，ACTN4及TRPC6基因突变是致病原因之一，尚未发现NPHS2相关致病突变。     

汉族人群青少年特发性脊柱侧凸家系中存在复合型杂合DOCK9基因突变

目的探讨青少年特发性脊柱侧凸(AIS)家系中可能的致病基因突变。方法对一个汉族AIS家系的3名成员(先证者及其表型正常的父母)进行全基因组测序,筛选潜在的致病基因突变,并通过Sanger测序验证所有发现的突变。结果在AIS家系中发现DOCK9基因存在复合型杂合基因突变c.3259TC(p.F1087L)和c.2465AG(p.Y822C)。患者的父母是未出现AIS表型的突变基因携带者,父亲携带c2465AG突变,而母亲携带c.3259TC突变。结论复合型杂合DOCK9基因突变可能导致AIS的发生,其在AIS发生机制中的作用有待于进一步探索。     

A novel VCP mutation as the cause of atypical IBMPFD in a Chinese family

IntroductionInclusion-body myopathy (IBM) with Paget's disease of bone (PDB) and frontotemporal dementia (FTD), designated as IBMPFD, is a rare, autosomal dominant disorder (MIM 605382). IBMPFD is caused by mutations in the gene that encode valosin-containing protein (VCP). We investigated a Chinese family in which multiple members were diagnosed with PDB and suffered from weakness of the limbs. However, no members of this family were diagnosed with FTD. We made a preliminary diagnosis of PDB, but failed to identify an SQSTM1 mutation in any of the patients. We used whole-exome sequencing to identify the pathogenic gene mutation affecting the Chinese male proband.Materials and methodsAltogether, 254 subjects, including one 56-year-old male proband, four affected, related individuals and additional nine family members from a non-consanguineous Chinese family, and 240 healthy donors were recruited and genomic DNA was extracted. All eight exons and the exon–intron boundaries of the SQSTM1 gene were amplified by polymerase chain reaction (PCR) and directly sequenced in five patients (II13, II4, II5, II8, II9). Using whole-exome sequencing, we identified a novel mutation in VCP as the disease-causing mutation. We confirmed the result by sequencing a 500-bp region of the promoter and the coding region of VCP in all 254 of the participants using Sanger sequencing.ResultsNo mutation in the SQSTM1 gene was identified in the five patients examined using direct Sanger sequencing. However, through whole-exome sequencing we were able to identify a novel missense mutation in exon 3 of the VCP gene (p.Gly97Glu) in the Chinese male proband. This mutation was confirmed using Sanger sequencing. The proband, four affected individuals and three unaffected individuals carried this mutation. We were able to correctly diagnose the patients with atypical IBMPFD. Structural analysis of the p.Gly97Glu mutation in the VCP protein showed that the affected amino‐acid is located in the interface of the protein. This abnormality may therefore interfere with protein function.ConclusionsThis is the first report of a family from China with IBMPFD. A novel VCP mutation was found as the cause of atypical IBMPFD in a Chinese family. Our findings confirm that VCP gene mutations can be a pathogenic cause of IBMPFD.     

中国人恶性高热家系蓝尼定受体-1基因的筛查

目的 筛查中国人恶性高热(MH)家系的蓝尼定受体-1(RYR1)基因.方法 提取确诊为MH的先证者及家系其他成员外周血白细胞的基因组DNA,采用PCR扩增先证者RYR1基因的部分外显子,并进行直接测序,采用Fok Ⅰ限制酶分析验证先证者及家系其他成员基因突变情况.结果 先证者RYR1基因第6 724位碱基C突变为T(c.6724 C＞T),所编码第2 206位氨基酸由苏氨酸变为甲硫氨酸(p.T 2206 M),此突变在白种人已报道;先证者的4个子女中有2个为该错义突变携带者,为MH易感者.结论 中国人MH易感者携带与白种人MH易感者相同的RYR1基因突变.     

常染色体隐性遗传Alport 综合征一家系COL4A3及COL4A4基因突变分析

目的 通过对有近亲婚配史的Alport综合征一家系Ⅳ型胶原α3和α4链的 COL4A3/COL4A4 基因分析，明确常染色体隐性遗传Alport综合征的基因突变,为该病的基因诊断和家系遗传咨询提供更为全面的理论基础｡ 方法 PCR扩增先证者DNA COL4A3/COL4A4 基因的共98个外显子，经直接测序，寻找突变位点，对有意义的突变经限制性内切酶AvaⅡ酶切在家系中分析验证｡ 结果 在该患者中共发现1个错义突变和10个序列变异｡其中在COL4A3 基因上发现一个位于42号外显子上的错义突变 G3725A,导致蛋白质Gly1242Asp的突变｡错义突变在患者中是纯合子，携带者中是杂合子，其他正常家系成员及筛查100条正常人染色体，未发现该突变｡10个序列变异为单核苷酸多态性改变｡ 结论 报道了一个国内较少见的常染色体隐性遗传Alport 综合征家系，同时经基因突变筛查发现Ⅳ型胶原α3链的一个新的致病性的基因突变｡     

COQ2 mutation associated isolated nephropathy in two siblings from a Chinese pedigree

BackgroudCoenzyme Q10 (CoQ10) is involved in the biosynthesis of adenosine triphosphate (ATP), and is most abundant in the mitochondrial membrane. The primary CoQ10 deficiency caused by COQ2 defect is mostly manifested as encephalopathy, encephalopathy with nephropathy, and rarely as an isolated nephrotic syndrome.MethodsClinical and pathological data and peripheral blood samples of 2 siblings with steroid-resistant nephrotic syndrome (SRNS) and their family members of a Chinese pedigree were collected. DNA was extracted and subjected to next-generation sequencing of target genes of hereditary nephropathy.ResultsCompound heterozygous mutations of COQ2 (c.1058A > G, p.Y353C, paternal and c.973A > G, p.T325A, maternal)were identified in both siblings of the pedigree. Mutation of p.Y353C was novel. The proband was a girl, who presented with SRNS at the age of 7 months. CoQ10 was administered after the gene sequencing results came out. Proteinuria decreased gradually to 1+, occasionally negative. The child was normal in growth and intelligence. She is now 4 years old. The second patient was her elder brother. He was found to have SRNS at the age of 2 years old. Renal pathology indicated focal segmental glomerulosclerosis (FSGS). Electronic microcopy revealed that a large quantity of mitochondria with normal contour was accumulated within the podocytes. Both patients were in normal intelligence without convulsion.ConclusionThe 2 cases harboring COQ2compound heterozygous mutations presented with isolated SRNS, with a renal pathology of FSGS and a large quantity of mitochondria with normal contour accumulated within the podocytes. CoQ10 was efficacy in eliminating proteinuria.     

Multiple epiphyseal dysplasia: A clinical and genetic study of 12 cases in a Swedish 6-generation family

Background Multiple epiphyseal dysplasia (MED) is a common genetically and clinically heterogeneous skeletal dysplasia characterized by early-onset osteoarthritis, mainly in the hip and knee, and mild-to-moderate short stature. Here we report on a 6-generation MED family with 17 affected members.Method The clinical and radiographic data on the 12 affected members still living were scrutinized. A structured inquiry comprising state of health and MED-related symptoms since birth up to the present time and the osteoarthritis outcome (KOOS) questionnaire were sent to all living family members with MED. The 5 known gene loci for autosomal dominant MED were analyzed for linkage, using fluorescence-labeled microsatellite markers. Linkage was ascertained with markers close to the COL9A2 gene, which was analyzed for mutations by sequencing.Results We identified an exon 3 donor splice mutation in the COL9A2 gene in all affected family members. Clinical, radiographic, and questionnaire data from affected family members suggested that MED caused by COL9A2 mutations starts in early childhood with knee pain accompanied by delayed ossification of femoral epiphyses. The disease then either stabilizes during puberty or progresses with additional joints becoming affected; joint surgery might be necessary. The progression of the disease also affects muscles, with increasing atrophy, resulting in muscle fatigue and pain. Muscular atrophy has not been reported earlier in cases with COL9A2 mutations.Interpretation In a patient with clinically suspected or verified MED, it is important to perform DNA-based analysis to identify a possible disease-causing mutation. This information can be used to carry out genetic risk assessment of other family members and to achieve an early and correct diagnosis in the children.     

Clinical significance of MEFV mutations in ankylosing spondylitis

ObjectiveThe aim of the present study was to investigate the prevalence of MEFV gene mutations in patients with ankylosing spondylitis (AS) and to assess the clinical significance of the MEFV gene mutations in AS.MethodsEighty AS patients and 85 healthy controls were examined for 12 common MEFV mutations via strip-assay technique. Bath ankylosing spondylitis disease activity index (BASDAI), bath ankylosing spondylitis functional index (BASFI), visual analogue scale (VAS) for pain, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), Schober test, chest expansion measurements, hip involvement, ocular involvement, articular pain, and presence of syndesmophytes were used to assess the disease severity in patients.ResultsTwenty-four of the AS patients (30%) and 17 of the healthy controls (20%) were found to carry a single MEFV mutation. There was no significant difference between the AS patients and controls in terms of MEFV gene mutation frequency (p = 0.13, OR: 1.71, 95% CI: 0.83–3.50). When the patients were divided into two groups as MEFV mutation carriers and noncarriers, there was significant difference between the groups regarding BASFI and BASDAI whereas there was no significant difference in VAS score for pain. No association was found with the clinical findings and MEFV mutation except hip involvement. While there was no significant difference in CRP levels, individuals with MEFV mutation had a higher ESR than the noncarriers.ConclusionMEFV gene mutation carriage rate was not found to be significantly higher in AS patients when compared with healthy controls. However having an MEFV mutation seems to aggravate the disease course in AS.     

COL1A1新发突变致儿童成骨不全症 1 例报道并文献复习

目的分析一例I型成骨不全症(osteogenesis imperfecta,OI)患儿的临床特点,并研究患者及其家系的基因突变情况及致病性鉴定。方法详细询问病史,分析骨转换指标、骨密度、骨骼X线特点。采用高通量测序法,对患儿骨病检测包225个相关致病基因各外显子编码区域的序列变异情况进行检测分析,采用PCR结合Sanger测序的方法验证突变位点变异情况。结果骨标志物提示高转换水平,影像学提示骨量低下,四肢长骨纤细、骨皮质变薄,基因检测发现患儿COL1A1基因编码区杂合变异c.2911_2912insAG(p.G971Efs*138),经Mutation Taster预测显示为致病性突变。先证者母亲、父亲以及妹妹均未携带该突变基因。结论发现了OI患者COL1A1基因新的突变位点c.2911_2912insAG(p.G971Efs*138),丰富了中国人OI群体COL1A1基因致病突变谱。