脊髓小脑性共济失调2型的分子遗传学诊断及临床分析

目的:研究分析脊髓小脑性共济失调2型(SCA2)的分子遗传学诊断、应用以及临床表现特征。方法:对来自广西地区临床诊断为SCA的1个家系2例患者和8名"健康"家系成员,以及35名正常对照人员,通过聚合酶链式反应、琼脂糖电泳等技术检测SCA2基因位点内CAG三核苷酸重复扩增次数,并对异常等位基因片段进行DNA测序。结果:我国广西正常人群SCA2等位基因CAG重复数为20～29次,1家系中2例患者与1例症状前患者存在SCA2(CAG)n扩展突变,拷贝数分别为42、45、55次。结论:首次发现广西SCA2,利用分子遗传学分析可进行SCA2基因诊断,为症状前诊断及遗传咨询提供依据。     

脊髓小脑性共济失调3型真核表达载体的构建及在PC12中的表达

目的构建以增强型绿色荧光蛋白(EGFP)作为报告基因、带有脊髓小脑性共济失调3型(SCA3)基因的真核细胞表达载体,并转染PC12细胞,观察外源基因的表达情况。方法将含有正常和突变SCA3基因的pcD-NA3．0-ataxin3Q28(基因内CAG重复28次)、pcDNA3．0-ataxin3Q84(基因内CAG重复84次)双酶切,水解片段插入pEGFP-C1,测序鉴定后脂质体转染法转染PC12细胞并优化转染条件,荧光显微镜观察EGFP的表达,Western印迹检测目的蛋白ataxin3的表达。结果真核表达载体pEGFPC1-ataxin3Q28、pEGFPC1-ataxin3Q84得以成功构建,EGFP及目的基因产物ataxin3Q28、ataxin3Q84顺利地在PC12细胞中表达,由此建立了SCA3的真核细胞表达模型。结论构建SCA3的真核表达载体并建立其细胞模型,对研究SCA3的发病机制有重要意义。     

遗传性脊髓小脑型共济失调7型遗传学诊断及临床特征

目的研究中国人遗传性脊髓小脑型共济失调(SCA)7型(SCA7)的基冈突变和临床特征。方法应用聚合酶链反应(PCR)、聚丙烯酰胺凝胶电泳(PAGE)等技术对临床表现为SCA的92个家系112例患者和16例散发SCA患者的SCA7基因内CAG三核苷酸重复序列进行检测,对异常等位基因片段进行DNA测序,分析基因型和表型之间的关系,并与表型正常的家系成员和健康人对照。结果在1个SCA7家系的6位成员中检测出2例患者的SCA7等化基因内CAG重复数目为71;临床表现主要为共济失调、视力下降、黄蓝色盲及视网膜色素变性。该家系内表型正常的4位成员SCA7等位基因CAG重复数目为7～9,另126例临床表现为SCA的患者、71名表型正常的家系成员及60名健康对照者SCA7等位基因内CAG三核甘三酸重复数为6—21。结论CAG过度扩增为SCA7的致病原因,分子遗传学分析有助于SCA7的诊断;视网膜色素变性为SCA7的重要特征。     

脊髓小脑性共济失调12型一家系的临床特征和基因突变分析

目的研究1个遗传性共济失调12型(spinocerebellar ataxia type 12 SCA12)家系的临床特征与基因突变特点。方法应用聚合酶链反应、毛细管电泳等方法对1个临床诊断为遗传性共济失调的家系进行SCA基因检测。结果确定该家系为遗传性共济失调SCA12型家系。共确诊7例现证患者,患者异常CAG的重复次数为51⁵5次。结论上肢震颤,逐渐出现共济失调、延髓麻痹,病理征阳性为SCA12型相对独特的临床表现。先证者异常片段CAG重复为55次,第3代患者Ⅲ2 CAG重复次数54次,发病年龄提前,可能存在遗传早现现象。SCAs核苷酸突变扩展的数目与年龄呈负相关,与症状严重程度呈正相关的特点可能也存在于SCA12型中。     

脊髓小脑共济失调2型、3型患者10例基因突变与临床表型分析

目的探讨脊髓小脑共济失调（SCA）2型、3型患者的基因突变特点及临床表型。方法总结4个SCAs家系的9例患者和1例散发患者的临床表现，对全部患者、43名家系成员及60名健康对照，采用PCR、荧光PCR、毛细管电泳等技术检测分析SCA2、3基因内CAG三核苷酸重复序列的长度及拷贝数。结果3个家系中的6例患者、1例散发患者存在SCA3／MJD（CAG）n扩展突变，CAG重复数为68—75次；1个家系的3例患者存在SCA2（CAG）n扩展突变，CAG重复数为39—41次。两型患者的临床表现有重叠之处，但在发病年龄、病程进展、神经系统受累部位等方面有明显差异。结论SCA3／MJD型与SCA2型在临床表现上存在一定差异性，有助于鉴别和分型，但基因检测是明确诊断的惟一方法。     

脊髓小脑性共济失调3型一家系报道

目的 探讨脊髓小脑性共济失调(spinocerebellar ataxia, SCA)3型(SCA3)的临床特点。方法 收集1例2016年6月在济宁医学院附属医院神经内科住院的SCA3患者及其家系的临床资料,基因检测明确家系分型,结合文献分析家系中的临床差异。结果 患者23岁发病,临床表现为行走不稳伴进行性下肢无力,肌张力正常,水平性诱发眼震,无跌倒及吟诗性语言,闭目难立征阳性,呈宽基底步态,双侧巴氏征未引出,腹壁反射减低,膝反射及跟腱反射活跃,深浅感觉未见异常。基因检测提示患者ATXN3基因CAG重复次数79次(正常人范围在12～44次),超过正常范围,患者其它(ATXN1、ATXN2、CACNA1A、ATXN7、ATXN8、PPP2R2B、TBP)基因的CAG重复次数均在正常范围内。患者的55岁父亲、6岁女儿及4岁儿子的ATXN3基因的CAG重复数分别为65、73、71次,超过正常范围,但均未发病。患者母亲ATXN3基因的CAG重复数29次,属正常范围,未见临床症状。结论 本例家系是SCA3型,具有临床异质性,基因检测有助于诊断。     

广东汉族人遗传性脊髓小脑型共济失调基因突变的研究

目的 研究广东汉族人遗传性脊髓小脑型共济失调（SCA）的亚型（SCA1、SCA2、SCA3和SCA7）基因突变分布频率。方法 对临床诊断为SCA的18个家系24例SCA患者、22例散发SCA患者、45名家系“正常人”及30名非家系健康人,通过聚合酶链反应（PCR）及聚丙烯酰胺凝胶电泳（PAGE）等技术检测SCA1、2、3、7基因内CAG三核苷酸重复序列突变,并利用ABI310、377测序仪对异常等位基因片段进行DNA测序。结果 在18个SCA家系中,1个家系（5.55％）有SCA1基因突变,2个家系（11.11％）有SCA2基因突变,7个家系（38.88％）有SCA3基因突变,未发现SCA7突变,。在家系成员中检出1例症状前SCA3患者。22例散发性患者中检出SCA1、SCA2、SCA3和SCA7各1例,各占4.54％。SCA1患者CAG重复数为53～56次,正常人19～34次;SCA2患者CAG重复数为38～45次,正常人20～25次;SCA3患者CAG重复数为74～81次,正常人14～40次。SCA7患者CAG重复数为65次,正常人为9～19次。结论 提示中国人SCA主要为SCA3/MJD型,其次为SCA2和SCA1。基因分析对家族性以及散发性SCA患者的临床确诊和遗传咨询均有重要意义。     

遗传性脊髓小脑性共济失调7型的基因突变及临床特征分析

目的 研究遗传性脊髓小脑性共济失调7型（SCA7）的基因突变和临床特征。方法 对临床诊断为脊髓小脑性共济失调（SCA）的15个家系24例患者、20例散发SCA患者、41名家系“正常人”及30名非序列的突变,并利用ABI373例序仪对异常等位基因片段进行DNA测序。结果 24例SCA患者的SCA7等位基因CAG重复数目为9－18。正常人SCA7等位基因CAG重复数目为9－19。检出1例散发患者为SCA7,经基因测序证实,其异常等位基因的CAG重复数目为63。结论 CAG过度扩增是SCA7的致病原因,利用基因突变分析可进行基因诊断,提供症状前诊断及遗传咨询的依据,为基因分型奠定基础。     

中国汉族人群CACNA1A基因CAG重复数目分布特点及其在脊髓小脑性共济失调6型基因诊断中的应用

目的 探讨我国汉族人群CACNA1A基因CAG重复数目分布特点及其在脊髓小脑性共济失调6型(spinocerebellar ataxias type 6,SCA6)基因诊断中的应用.方法 应用"两步PCR法"、变性聚丙烯酰胺凝胶电泳(DPAGE)和测序等方法对300名健康对照及109例无血缘关系的SCA患者进行CACNA1A基因CAG三核苷酸重复数目分析.结果 300名健康对照的CAG重复次数范围为3～18次,以13次最常见.在109例SCA患者中,发现1例SCA6患者,其CAG异常重复次数为24次,该患者的母亲和哥哥亦为SCA6患者,临床上均表现为缓慢进展的小脑性共济失调、构音障碍、眼震、轻度的振动及本体觉减退,遗传早现现象较明显.结论 SCA6病例在我国较少见,进行CACNA1A基因突变分析有助于临床诊断."两步PCR法"可提高CACNA1A基因突变分析的效率.     

中国汉族人群CACNA1A基因CAG重复数目分布特点及其在脊髓小脑性共济失调6型基因诊断中的应用

目的 探讨我国汉族人群CACNA1A基因CAG重复数目分布特点及其在脊髓小脑性共济失调6型(spinocerebellar ataxias type 6,SCA6)基因诊断中的应用.方法 应用"两步PCR法"、变性聚丙烯酰胺凝胶电泳(DPAGE)和测序等方法对300名健康对照及109例无血缘关系的SCA患者进行CACNA1A基因CAG三核苷酸重复数目分析.结果 300名健康对照的CAG重复次数范围为3～18次,以13次最常见.在109例SCA患者中,发现1例SCA6患者,其CAG异常重复次数为24次,该患者的母亲和哥哥亦为SCA6患者,临床上均表现为缓慢进展的小脑性共济失调、构音障碍、眼震、轻度的振动及本体觉减退,遗传早现现象较明显.结论 SCA6病例在我国较少见,进行CACNA1A基因突变分析有助于临床诊断."两步PCR法"可提高CACNA1A基因突变分析的效率.     

脊髓小脑型共济失调6型患者的临床特征及基因突变分析

目的研究脊髓小脑型共济失调(SCA)6型的临床特征和基因突变频率。方法采用聚合酶链反应(PCR)、聚丙烯酰胺凝胶电泳(PAGE)等技术,对160个SCA家系330例患者和77例散发SCA患者进行SCA6(CAG)n的重复数分析,对异常等位基因进行测序;并对SCA6患者进行临床及MRI检查。结果检测确定4个SCA6家系6例患者SCA6(CAG)n的重复数为25和26,正常人群SCA6(CAG)n的重复数为5～17,SCA6突变频率为本组SCA家系的2.5%。SCA6患者临床仅表现为缓慢进展的小脑性共济失调,MRI显示单纯小脑萎缩。结论SCA6是中国SCA中的少见亚型,SCA6与其他SCA亚型有不同的临床及影像学特征。     

Clinical features and genetic analysis of a Spanish family with spinocerebellar ataxia 6

Objective - To present the clinical features and DNA analysis of a Spanish SCA6 family. Material and methods - Four symptomatic members of the family (mean age at onset: 53.75 ± 5.21) were examined. SCA6 CAG trinucleotide repeat was analysed in the proband by the polymerase chain reaction (PCR). Results - Early dysphagia, ophthalmoparesis and neck dystonia in the oldest patient, without the loss of vibratory and proprioceptive sensation supporting the theory of phenotypic variability within families with SCA6 . Our results are in accordance with the theory that the size of the repeat pattern correlates with the age at onset of the symptoms. Analysis of the SCA6 CAG trinucleotide repeat at the CACNA1A gene in the patient's DNA demonstrated an expanded allele of 22 CAG repeat units. Conclusions - This study identifies phenotypic differences in the surviving kindred. The diagnosis of SCA6 in family members or single affected patients can be made by direct molecular analysis. This makes predictive testing possible.     

The molecular diagnosis of spinocerebellar ataxia type 1 in patients with ataxia

Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disease caused by an expanded CAG trinucleotide repeat in the SCA1 gene on chromosome 6. We screened 40 probands with ataxia for the CAG repeat expansion and found five probands with SCA1 representing five different families. The SCA1 gene was analysed in 50 members of these families, and the CAG repeat expansion was found in all 17 affected persons and in 14 unaffected at-risk individuals. The range of expansion was 41–53 repeat units, while the range of normal alleles was 22–36 repeat units. We found pronounced inter- and intrafamilial phenotypical variation. One of the families had a comparatively mild phenotype which correlates with a CAG repeat length in the low end of the range of expansions and a late age at onset. With few exceptions, normal alleles of the SCA1 gene have one to three CAT interruptions in the middle of the CAG repeat, while all expanded alleles are uninterrupted. We report the hitherto longest normal uninterrupted allele of 22 repeat units and stress the importance of analysis for the presence of CAT interruptions in the diagnosis of SCA1.     

Analysis of a fully penetrant spinocerebellar ataxia type 8 Brazilian family

Spinocerebellar ataxia type 8 (SCA8) is a progressive neurological disorder caused by the expanded repeat CTA/CTG of two overlapping genes, ATXN8OS and ATXN8, expressed bidirectionally. Normal alleles have 15‐50 repeats, and pathogenic alleles range from 71 to 1300 repeats. The disorder is relatively rare, accounting for about 2%‐5% of the autosomal dominant forms of hereditary ataxia worldwide. However, the prevalence of disease‐causing ATXN8OS/ATXN8 expansions is higher than the disease because of the reduced penetrance of the expanded allele. The aim of this study was to describe the first fully penetrant SCA8 family showing mixed Brazilian African and Amerindian origin. Eight members of this family were evaluated—the mother and seven offspring—through a complete neurological examination conducted at the Neurogenetics Clinic, HCFMRP—USP in Brazil. The number of CTA/CTG repeats was obtained after polymerase chain reaction (PCR) and fragment analysis. The haplotype analysis was conducted using a microsatellite marker, D13S1296, and four single nucleotide polymorphisms (SNPs), rs1831189, rs8002227, rs11841483, and rs72284461, all spanning a 70.1 Mb region on chromosome 13q21.3. The molecular analysis showed that the expansions ranged from 104 to 109 CTA/CTG repeats in the six affected individuals and were absent in two asymptomatic daughters (aged 53 and 40 years). Three SNPs cosegregate with the expanded alleles, confirming the connection between expansion and disease in this family. As the SCA8 diagnosis demands careful interpretation, we suggest the use of linkage analysis to observe segregation of the mutation, making more accurate its genotyping.     

脊髓小脑性共济失调的症状前诊断研究

目的:探讨脊髓小脑性共济失调的症状前患者分子遗传学诊断及其面临的有关问题。方法:对临床诊断为脊髓小脑性共济失调(SCA)患者进行基因诊断的同时根据申请者要求检测其家系“健康”个体,采用聚合酶链反应(PCR)对三核苷酸重复(TNR)片段进行扩增,聚丙烯酰胺凝胶电泳并计算其长度,推算正常和异常扩增等位基因内TNR重复拷贝数。结果:多数家系无症状患者均要求进行致病基因突变检测。通过基因检测诊断出6例SCA3、2例SCA1、2例SCA12症状前患者。结论:分子遗传学检测可作出可靠的症状前诊断。进行遗传咨询应考虑医学、伦理、法律及社会诸多问题。     

Dystonia in spinocerebellar ataxia type 6.

Spinocerebellar ataxias are heterogeneous disorders with overlapping clinical features. Spinocerebellar ataxia-6 is a dominantly inherited condition characterized by relatively pure ataxia with a paucity of other manifestations including extrapyramidal findings. We report on two patients with genetically proven SCA-6 who had dystonia. One patient presented initially with dystonia, which remained the most disabling problem. Dystonia may occur in SCA-6 and can be disabling.