中国人群Leber遗传性视神经病变线粒体ND1基因突变频谱分析

摘要

背景 Leber遗传性视神经病变(LHON)是一种常见的遗传性眼病,常导致患者双侧中心视力下降,其基因突变频谱并不完全清楚. 目的 筛查中国人群LHON线粒体ND1基因的突变位点并明确其突变频谱.方法 经温州医学院伦理道德委员会批准,所有受试者知情同意.收集临床诊断为LHON患者894例和正常对照者134名,抽取受检者的外周血提取基因组DNA,采用聚合酶链反应(PCR)法对受检者线粒体ND1基因进行扩增和测序,并与修正的线粒体剑桥参考序列进行比对,筛查突变位点并分析其突变频率.结果 本研究在894例LHON患者的ND1基因中发现了7个与LHON相关的突变位点G3316A、T3394C、G3460A、C3497T、G3635A、G3733A、T4216C,携带这7个位点的患者共100例,占11.19％,其中与3个原发性突变位点共存者29例(3.24％).这几个突变位点在病例中的发生频率分别为2.57％、2.23％、1.45％、3.80％、0.67％、0.11％、0.34％,其中G3316A、T3394C、C3497T、T4216C在正常对照者中也可检测到,分布频率分别为4.48％、2.99％、4.48％和1.49％,经分析在LHON组和正常对照组之间差异均无统计学意义(x2=0.926,P=0.336;x2=0.052,P=0.820;x2=0.142,P=0.707;P=0.129),而其他一些在欧美人群中报道的位点,如G3376A、G3496T、G3700A、A4136G、T4160C、C4171A在中国LHON人群中并未发现. 结论 线粒体ND1基因是中国人群LHON的一个突变热点区域,约11.19％的中国LHON患者与ND1基因突变有关.G3635A、G3733A为中国人群中少见的致病突变位点,而G3316A、T3394C、C3497T、T4216C本身并不足以致病,但可能对LHON外显率和表型的表达起协同作用.%Background Leber hereditary optic neuropathy (LHON)is a common inherited eye disease,which generally affects young adults with bilateral loss of central vision.Mutation frequency of Leber hereditary has not been fully clarified. Objective This study was to investigate the mutation frequency of mitochondrial NDI gene associated with LHON in Chinese population. Methods The proposal of the study was approved by Ethic Committee of Wenzhou Medical College.Written informed consent was obtained from each subject initial of this trial.Eight hundred and ninety-four LHON patients and 134 normal subjects were collected.Genomic DNA was extracted from peripheral blood leukocytes of the all participants.Polymerase chain reaction (PCR) was used to amplify and sequence analysis of the mitochondrial ND1 gene was performed and aligned with revised Cambridge Reference Sequence(rCRS) of mitochondrial DNA.Then mutated gene frequency was screened and analyzed. Results Mutational analysis of mitochondrial ND1 gene in 894 LHON patients revealed the presence of G3316A,T3394C,G3460A,C3497T,G3635A,G3733A,and T4216C.11.19％ LHON patients (100/894 ) were found to be associated with the gene mutations mentioned above,and 3.24％ patients (29/894) showed the co-occurrence of three primary mutations.Mutation frequencies in LHON patients were 2.57％,2.23％,1.45％,3.80％,0.67％,0.11％,0.34％,respectively,and G3316A,T3394C,C3497T and T4216C also were detected in 134 normal controls with the mutation frequencies of 4.48％,2.99％,4.48％ and 1.49％,respectively.Mutation frequency analysis showed an insignificant difference in the mutations of G3316A,T3394C,C3497T and T4216C between LHON patients and normal controls (x2 =0.926,P=0.336;x2 =0.052,P=0.820; x2 =0.142,P=0.707;P=0.129).G3376A,G3496T,G3700A,A4136G,T4160C and C4171A were absent in Chinese LHON patients. Conclusions Mitoehondrial ND1 gene in LHON is a mutational hotspot in Chinese population,11.19％ (100/894)associated with LHON was caused by ND1 gene mutation.G3635A,G3733A may be rare pathological mutation in Chinese population.However,G3316A,T3394C,C3497T and T4216C are insufficient to produce the clinical phenotype,but they may play a synergic role for penetrance and phenotypic manifestation in LHON.