COL2A1基因新发突变导致先天性脊柱骨骺发育不良的遗传分析及产前诊断

摘要:目的?对1例先天性脊柱骨骺发育不良(SEDC)患儿进行基因检测和蛋白质功能预测，确定其致病原因，为家系遗传咨询及产前诊断提供依据。方法?应用高通量测序法对先证者454个骨病相关基因进行检测，对检出的可疑基因突变进行Sanger测序检测，对父母进行验证并对胎儿进行产前诊断。结果?该患儿检出COL2A1基因c.3589G>A(p.Gly1197Ser)杂合错义突变，其父母均未携带该突变，先证者为新发变异，家系中胎儿产前诊断结果提示胎儿未携带与先证者相同的变异。Polyphen2、Mutation Taster软件对其蛋白质功能预测的结果为有害。结论?COL2A1基因c.3589G>A变异是该SEDC家系先证者的致病原因。     

脊柱骨骺发育不良的影像学表现

目的：分析脊柱骨骺发育不良的影像学特征。方法：12例脊柱骨骺发育不良患儿，其主要临床症状为躯干与四肢不成比例的矮小身材、背部和肢体疼痛、脊柱侧弯及畸形足等。所有患儿进行一系列的X线检查。结果：检查发现头颅直径增大3例，齿状突发育不良2例，扁平椎12例，椎间隙狭窄12例，髋内翻11例，股骨近端骨骺骨化延迟12例．青少年髋关节退行性改变6例。结论：脊柱骨骺发育不良的影像学表现有其特征性，结合其临床表现能够做出明确诊断。     

晚发性脊柱骨骺发育不良并进行性关节病1例

病例 女,14岁,双髋关节疼痛不适、跛行2年,加重半月.查身体矮小,相当于6-8岁儿童的身高,鸭形跛行,智力正常.实验室检查:血常规正常,HLA-B27阴性,类风湿因子(RF)阴性.     

胎儿左心发育不良综合征的产前超声诊断

先天性心脏畸形是一种常见的先天畸形，其发病率在活产新生儿中约占0.5%-1.0%，在死胎中可高达3%。左心发育不良综合征指左侧心腔，主动脉发育不良的一组复杂先天性心脏病，病变包括左心房和左心室发育不良，主动脉瓣和（或）二尖瓣口狭窄或闭锁，升主动脉发育不良等。本病的病因不明，约占先天性心脏病患者的1.3%-7.5%,，且预后极差，如能早期发现对患者和临床都具有很大意义。     

一例成骨不全Ⅱ型高危胎儿的产前基因诊断

目的对广东一疑似致死性侏儒症或成骨不全Ⅱ型的高危胎儿实施产前基因诊断,以阐明胎儿骨发育异常的真实病因,及时预防患胎出生。方法对经超声检查初诊为致死性侏儒症或成骨不全、已孕25周的高危胎儿,在抽取脐血制备DNA模板后,采用PCR-DNA直接测序法,分别对胎儿的FGFR3基因和COL1A1基因进行突变检测,然后对所发现的突变进行分析和鉴定。结果 FGFR3基因未发现病理性突变,而COL1A1基因发现一典型的杂合错义突变(c.3065G>T,p.G1022V),经查HGMD数据库证实为成骨不全Ⅱ型的致病性突变。结论 (1)此高危胎儿为成骨不全Ⅱ型患胎,应及时终止妊娠(胎儿已经引产,经复查证实与产前基因诊断结果完全一致)。(2)在超声初诊基础上,采用产前基因诊断可快速、有效对高危胎儿做出确诊,为出生缺陷的预防提供技术保障。     

COL3A1在胃癌中的表达及意义

目的 检测胃癌及其正常组织中COL3A1的mRNA及蛋白表达水平,初步探究COL3A1与胃癌临床及病理特征的关系.方法 从胃癌手术患者癌组织及其正常组织中收集新鲜标本各10例,石蜡标本各50例,分别使用实时定量PCR法(Quantitative Real-time PCR)和免疫组化法检测COL3 A1 mRNA及蛋白...     

1个X-连锁迟发性脊椎骨骺发育不良家系的TRAPPC2基因突变分析

目的确定1个迟发性脊椎骨骺发育不良(spondyloepiphyseal dysplasia tarda,SEDT)家系的致病基因。方法收集先证者及家系的临床资料,提取先证者及亲属外周血DNA,用高通量测序技术对先证者的COL2A1、COL1A1、MATN3、TRAPPC2、FGFR3等189个骨骼相关基因的外显子编码区测序,对发现的致病突变进行Sanger测序验证,并对家系其他成员进行该突变的检测。结果在先证者TRAPPC2基因第5外显子上发现了1个移码突变c.271_275del CAAGA半合子缺失,为X-SEDT的致病性突变,同时发现患者母亲为该突变的携带者,而患者父亲和妹妹未检测到该突变。结论用靶向二代测序和Sanger测序结合的方法确定了1个X-SEDT家系的移码突变c.271_275del CAAGA半合子缺失,为临床遗传咨询提供了分子依据。     

COL3A1对胃癌细胞增殖及侵袭影响的实验研究

目的 研究COL3 A1在胃癌细胞增殖、迁移及侵袭过程中的作用.方法 以靶向COL3 A1的干扰慢病毒(3个靶点)及阴性对照慢病毒转染胃癌细胞株MGC803,嘌呤霉素筛选稳转细胞株.qRT-PCR验证转染后MGC803细胞中COL3A1的mRNA表达,筛选出2个转染效率最高的细胞株,再以Western blot法进一步...     

LMX1A基因多态性与中国汉族人群先天性脊柱侧凸遗传易感性的关联

背景：先天性脊柱侧凸是由于胚胎期脊柱椎体发育异常引起的脊柱侧凸,其遗传病因学假说开始引起许多学者的重视。目的：通过候选基因LMX1A上关键单核苷酸多态性位点的筛查,探索LMX1A与中国汉族人群先天性脊柱侧凸及其不同临床表型之间的关联。方法：入选127例中国汉族先天性脊柱侧凸患者,另选取外伤、感染、炎症性疾病等患者127例为对照组。根据国际人类基因组单体型图计划提供的基因型数据,应用Haploview 4.1软件选取LMX1A的标签和功能单核苷酸多态性位点。根据椎体畸形特点、畸形部位、畸形受累程度、有无合并肋骨畸形和椎管内畸形将病例组进一步分为不同临床表型。所有样本应用SNPstream UHT Genotyping系统对所选单核苷酸多态性位点进行基因型鉴定;进一步进行基于基因型/等位基因频率的关联分析,并用Haploview 4.1软件分析对照组单核苷酸多态性位点间是否存在连锁不平衡。结果与结论：共筛选6个位点：SNP1（rs 1819768）、SNP2（rs 12023709）、SNP3（rs 16841013）、SNP4（rs 4656435）和SNP5（rs 4657412）和SNP6（rs 4657411）,其基因型分布在病例组和对照组中均符合Hardy-Weinberg平衡;在基于基因型的关联分析中发现阳性位点SNP1和SNP2,单位点分析显示两位点基因型在病例组和对照组中的分布频率差异有显著性意义（P=0.026和P=0.026）,在进一步非条件Logistic回归分析中发现这两个位点的基因型分布最符合Ressessive（OR=0.38;95%CI=0.15～0.94,P=0.029,AIC=354.9）遗传模型;SNP1、SNP2、SNP3和SNP6处于连锁不平衡状态,SNP4和SNP5也处于完全连锁不平衡状态,但单倍体型与先天性脊柱侧凸的发生风险之间不存在相关性;在进一步与先天性脊柱侧凸临床表型的关联分析中发现SNP1基因型AC型、SNP2基因型AG型、SNP3基因型CT型与有椎体形成障碍先天性脊柱侧凸的易感性升高有关。结果提示在中国汉族人群中LMX1A基因可能和先天性脊柱侧凸的发生、发展相关,是一个重要的易感基因。     

A novel COL2A1 mutation causing spondyloepiphyseal dysplasia congenita in a Chinese family

BackgroundSpondyloepiphyseal dysplasia congenita is an autosomal dominant cartilaginous dysplasia characterized by short trunk, abnormal epiphysis, and flattened vertebral body. Skeletal features of SEDC are present at birth and evolve over time. Other features of SEDC include myopia and/or retinal degeneration with retinal detachment and cleft palate. A mutation in the COL2A1 gene located in 12q13.11 is considered as one of the important causes of SEDC. In 2016, Barat‐Houari et al. reported a large number of COL2A1 mutations. Among them, a non‐synonymous mutation in COL2A1 exon 37, c.2437G>A (p. Gly813Arg), has been reported to cause SEDC in only one patient from France so far.MethodsWe followed up a patient with SEDC phenotype and his family members. The clinical manifestations, physical examination and imaging examination, including X‐ray, CT and MRI, were recorded. The whole‐exome sequencing was used to detect the patients'' genes, and the pathogenic genes were screened out by comparing with many databases.ResultsWe report a Chinese patient with SEDC phenotype characterized by short trunk, abnormal epiphysis, flattened vertebral body, narrow intervertebral space, dysplasia of the odontoid process, chicken chest, scoliosis, hip and knee dysplasia, and joint hypertrophy. Gene sequencing analysis showed that the patient had a heterozygous mutation (c.2437G>A; p. Gly813Arg) in the COL2A1 gene. No COL2A1 mutation or SEDC phenotype was observed in his family members. This is the first report of SEDC caused by this mutation in an East Asian family.ConclusionThis report provides typical clinical, imaging, and genetic evidence for SEDC, confirming that a de novo mutation in the COL2A1 gene, c.2437G>A (p. Gly813Arg), causes SEDC in Chinese population.     

多种分子遗传技术应用于2例小额外标记染色体胎儿的产前筛查与诊断

目的运用多种分子遗传技术对小额外标记染色体(small supernumerary marker chromosome,sSMC)胎儿进行产前筛查和诊断,探讨各分子遗传技术的优缺点。方法对2例无创产前基因检测(non-invasive prenatal tests, NIPT)高风险胎儿的孕妇进行羊膜腔穿刺术,采集羊水标本,运用多重连接依赖探针扩增技术(multiplex ligation-dependent probe amplification,MLPA)对2例胎儿进行染色体非整倍体快速诊断,同时进行细胞培养和G显带核型分析;采用染色体微阵列技术(chromosome microarray, CMA)对胎儿1(18三体高风险)进行验证,采用MLPA技术对胎儿2(性染色体异常高风险)进行Y染色体微缺失检测。2例胎儿双亲均进行外周血G显带核型分析。结果 MLPA技术快速诊断结果为:胎儿1的18号染色体短臂部分三倍体,胎儿2携带2条正常X染色体和1条短臂缺失的Y染色体;胎儿1羊水G显带核型结果为47,XY,+Mar(胎儿1),胎儿2为47,XX,+Mar;胎儿1的CMA检测结果为arr[hg19]18p11.32p11.21 (136,227-15,099)×4;胎儿2的Y染色体微缺失结果为SRY基因和生精区AZFc区缺失。2例胎儿父母双亲的外周血染色体核型均正常。结论 NIPT检测适用于sSMC胎儿的产前筛查,不同的分子遗传产前诊断技术检测sSMC胎儿各有优缺点,相互补充验证的结果可为sSMC胎儿临床遗传咨询提供更详细的信息。     

A novel ATP1A2 gene mutation in an Irish familial hemiplegic migraine kindred

OBJECTIVE: We studied a large Irish Caucasian pedigree with familial hemiplegic migraine (FHM) with the aim of finding the causative gene mutation. BACKGROUND: FHM is a rare autosomal-dominant subtype of migraine with aura, which is linked to 4 loci on chromosomes 19p13, 1q23, 2q24, and 1q31. The mutations responsible for hemiplegic migraine have been described in the CACNA1A gene (chromosome 19p13), ATP1A2 gene (chromosome 1q23), and SCN1A gene (chromosome 2q24). METHODS: We performed linkage analyses in this family for chromosome 1q23 and performed mutation analysis of the ATP1A2 gene. RESULTS: Linkage to the FHM2 locus on chromosome 1 was demonstrated. Mutation screening of the ATP1A2 gene revealed a G to C substitution in exon 22 resulting in a novel protein variant, D999H, which co-segregates with FHM within this pedigree and is absent in 50 unaffected individuals. This residue is also highly conserved across species. CONCLUSIONS: We propose that D999H is a novel FHM ATP1A2 mutation.     

Three novel polymorphic sequence variants in the type I collagen gene COL1A1, the main disease locus for Osteogenesis Imperfecta

Three novel polymorphic variants were found within COL1A1 genomic sequence (accession number AF017178) while screening several patients in the search of OI causal mutations. The three polymorphisms, located in intron 12, exon 26, and intron 29, respectively, can be detected by PCR amplification and digestion with appropriate restriction enzymes (Mbo II, Bst NI, Pvu II, respectively). Allelic frequencies within the Italian population were calculated.     

COL6A1基因嵌合突变的Bethlem肌病一例并文献复习

目的 报道1例由COL6A1基因嵌合突变所致的Bethlem肌病,并分析该突变的致病性。 方法 应用全外显子基因组测序法（trio-WES）对1例Bethlem肌病患儿及其父母进行基因测序,对检出突变进行生物信息学预测,并以“Bethlem肌病”“COL6A1”（包括中英文）为检索词在PubMed、CNKI、中华医学期刊全文数据库（MedBook）检索相关病例,在千人基因组数据库、ExAC数据库及ClinVar数据库检索患儿的基因突变。 结果 经trio-WES检测发现,患儿COL6A1基因第8外显子存在1个c.868G > A （p.G290R）错义突变（突变频率为48.13%）,该突变同时在其父亲外周血中检出,但突变频率仅7.89%,考虑为嵌合突变（突变频率<10%）,属于新发突变（PS2）;该突变导致的蛋白水平改变发生在甘氨酸位置,属于COL6A1基因热点区域突变（PM1）;同时该突变在正常人群突变频率数据库中均不存在（PM2）。经多种有害突变预测软件预测结果均提示c.868G > A （p.G290R）为有害突变（PP2+PP3）,根据美国医学遗传学与基因组学学会指南判定该新发错义突变为致病性突变（PS2+PM1+PM2+PP2+PP3）。在数据库检索到6例COL6A1基因突变所致Bethlem肌病先证者,无存在嵌合突变者。 结论 COL6A1基因c.868G >A （p.G290R）为该患儿罹患Bethlem肌病的致病原因,该突变未曾被报道,这在一定程度上扩充了COL6A1基因的变异谱。     

羊水细胞低密度脂蛋白受体基因突变分析在家族性高胆固醇血症产前诊断中应用

目的探讨羊水细胞低密度脂蛋白受体（low density lipoprotein receptor,LDLR）基因突变分析在家族性高胆固醇血症（familial hypercholesterolemia,FH）产前诊断中的应用价值。方法 3例曾生育FH重症患儿并再次妊娠的妇女及其核心家系成员,提取其外周血基因组DNA,筛查LDLR基因突变;于妊娠16～20周在超声引导下行羊膜腔穿刺术抽取羊水,提取胎儿脱落细胞DNA,分别对家系存在的LDLR基因突变进行检测,判断胎儿是否为重症FH。结果 3个家系均符合FH诊断,并分别在LDLR基因检测到2个互不相同的杂合突变位点;胎儿LDLR基因核苷酸序列分析证实,1号家系胎儿仅携带该家系1个突变位点判断为杂合（轻症）,2号家系胎儿携带该家系2个突变位点判断为复合杂合（重症）,3号家系胎儿未检到该家系的突变位点推测为正常个体。结论 FH孕妇羊水脱落细胞LDLR基因分析安全有效,可尽早发现FH纯合子患儿。     

Severe attacks of familial hemiplegic migraine, childhood epilepsy and ATP1A2 mutation

We studied four members of a family suffering from typical attacks of familial hemiplegic migraine (FHM) caused by a new mutation, R548C, of ATP1A2 gene in exon 12. One individual had also childhood absence epilepsy and generalized tonic-clonic seizures (GTCS). GTCS were followed by a severe attack of hemiplegic migraine at four times. Sodium valproate enabled control of both the epileptic seizures and the most severe FHM attacks. This association of FHM and epileptic seizures and their control with the same treatment suggest similar pathophysiological mechanisms.