甲基丙二酸血症伴同型半胱氨酸血症患儿基因突变分析  

Analysis of gene mutations in Chinese patients with methylmalonic acidemia and homocysteinemia

作　　者：王斐[1] 韩连书[1] 胡宇慧[1] 杨艳玲[2] 叶军[1] 邱文娟[1] 张雅芬[1] 高晓岚[1] 王瑜[1] 顾学范[1] 

机构地区：[1]上海交通大学医学院附属新华医院,上海市儿科医学研究所内分泌遗传代谢病研究室,200092 [2]北京大学第一医院儿科

出　　处：《中华儿科杂志》2009年第3期189-193,共5页Chinese Journal of Pediatrics

基　　金：十一五国家科技支撑计划课题（2006AB105A05,2006BA105A07）;国家高技术发展计划（2007AA022447）;上海市卫生局重点课题（2008ZD001）;上海市卫生局科研课题（2006043）

摘　　要：目的检测甲基丙二酸血症伴同型半胱氨酸血症患儿的MMACHC基因突变类型及突变频率。方法依据串联质谱检测血中的酰基肉碱、气相色谱-质谱检测尿甲基丙二酸、血清同型半胱氨酸测定及维生素B12负荷试验等，对28例甲基丙二酸血症伴同型半胱氨酸血症患儿进行诊断；应用聚合酶链反应（PCR）对这些患儿及其部分直系亲属、健康对照组的MMACHC基因外显子进行扩增，通过DNA直接测序进行基因突变分析。结果在28例患儿中27例检测到突变，共10种，其中2例仅检测到1个杂合突变，3例仅检测到多态性。突变集中在外显子3和4上（91．3％），以609G〉A（W203X）最常见，突变频率为53．6％，其中10例为纯合突变，10例为杂合突变，其次为658—660delAAG（K220del），突变频率为8．9％，均为杂合突变。另外检测到6种未报道突变，分别为1A〉G、365A〉T、658_del660AAG、301-3_327del30、567_568insT和625_626insT。3种基因多态性分别为-302T〉G（rs3748643）、-234A〉G（rs3728644）和321G〉A（rs2275276）。结论揭示了中国甲基丙二酸血症伴同型半胱氨酸血症患儿的部分基因突变谱，其中609G〉A（W203X）可能为热点突变。Objective Methylmalonic acidemia complicated with homocysteinemia, cblC type, is the most common inborn error of cobalamin metabolism. The gene MMACHC （ OMIM 277400） is located on chromosome lp34. 1 with four coding exons and a 5th non-coding exon. It encodes for a protein with 282 amino acid residues. So far, more than 40 mutations have been detected, in which 271dupA（R91KfsX14） is the hot spot of MMACHC gene. However, there have not been relevant reports in China. The present study aimed to identify the mutation types of MMACHC gene and analyze the genotype-phenotype correlations in Chinese patients. Method The diagnosis of this disease mainly depends on the measurement of C3 propionylcarnitine, C3/C0 （free carnitine） and C3/C2 （acetylcarnitine） in the blood by tandem mass spectrometry, the detection of methylmalonic acid in the urine by gas-chromatography mass spectrometry, the determination of total homocysteine in the serum, and the loading test of vitamin B12. The entire coding region of MMACHC gene was screened by polymerase chain reaction （PCR） combined with DNA direct sequencing in 28 Chinese patients. Genomic DNA was extracted using phenol-chloroform method from the peripheral blood leukocytes of each patient. PCR amplification products were checked by 1.8% agarose gel electrophoresis and were subsequently sequenced with both the forward and reverse primers. Mutational analyses were performed using normal human genomic MMACHC sequence as a reference （ GenBank ID： 25974）. Result In this study, ten mutations were identified in 27 of 28 Chinese patients. Most of them were located in exons 3 and 4 （91.3%）. We detected four mutations reported, which were 609G 〉 A （W203X）, 217C 〉T（R73X） , 271dupA（R91KfsX14）, and 394C 〉T（R132X） , and six novel mutations, which were 1A 〉 G, 365A 〉 T, 658_660delAAG, 301-3_327de1 30, 567_568insT, and 625_626insT. The 609G〉A （W203X） is the most common mutation, which was detected in 30 of 56 alleles （53.6%）, includi

关 键 词：甲基丙二酸 同型半胱氨酸 基因 突变 

分 类 号：R686[医药卫生—骨科学]