一例复合杂合F11基因变异所致的遗传性凝血因子Ⅺ缺陷症家系的分析

Analysis of a pedigree affected with hereditary coagulation factorⅪdeficiency due to compound heterozygous variants of F11 gene

目的对1个遗传性凝血因子Ⅺ(coagulation factor FⅪ,FⅪ)缺陷症患者家系进行临床表型和基因变异分析,寻找致病变异并初步探讨其分子机制。方法在Stago全自动血凝仪上检测先证者及其家系成员(共3代6人)活化部分凝血活酶时间(activated partial thromboplastin time,APTT)等血凝相关检测项目,FⅪ活性(FⅪactivity,FⅪ∶C)及其他有关凝血因子的活性;用ELISA法检测FⅪ抗原(FⅪantigen,FⅪ∶Ag)。提取基因组DNA,用Sanger测序法测定F11基因所有外显子及侧翼序列;用ClustalX-2.1-win软件分析变异氨基酸的保守性;用MutationTaster在线生物信息学软件分析变异是否有害;用Swiss-Pdb Viewer模型分析软件分析变异氨基酸对蛋白结构的影响。结果先证者APTT明显延长,为94.2 s;FⅪ∶C和FⅪ∶Ag分别降低为1%和1.3%;先证者父亲、母亲、儿子和女儿APTT分别为42.1 s、43.0 s、42.5 s和41.0 s,FⅪ∶C和FⅪ∶Ag均降低至正常值的一半左右;先证者丈夫APTT、FⅪ∶C和FⅪ∶Ag均正常。测序结果显示先证者F11基因存在第10外显子c.1103G>A(p.Gly350Glu)杂合错义变异和第13外显子c.1556G>A(p.Trp501stop)杂合错义变异。先证者父亲和女儿携带p.Gly350Glu杂合错义变异;母亲和儿子携带p.Trp501stop杂合无义变异。保守性分析表明Gly350和Trp501在同源物种间高度保守。MutationTaster在线生物信息学软件对两个变异的预测结果均为"disease causing",表明变异有害。蛋白模型分析表明,p.Gly350Glu变异影响了蛋白质的结构及稳定性。结论p.Gly350Glu和p.Trp501stop复合杂合变异可能是该家系遗传性FⅪ缺陷症的分子发病机制。

Objective To analyze the clinical phenotype and genetic basis for a Chinese pedigree affected with coagulation factorⅪ(FⅪ)deficiency.Methods Activated partial thromboplastin time(APTT)and other blood coagulation factors,FⅪactivity(FⅪ∶C)and other relevant coagulation factor activities for a large Chinese pedigree including 6 patients from 3 generations were determined on a Stago automatic coagulometer.The FⅪantigen(FⅪ∶Ag)was determined with an ELISA method.All exons and flanking regions of the F11 gene were subjected to Sanger sequencing.ClustalX-2.1-win software was used to analyze the conservation of amino acids.Pathogenicity of the variants was predicted with Mutation Taster online bioinformatics software and Swiss-Pdb Viewer.Results The APTT of the proband was prolonged to 94.2 s.The FⅪ∶C and FⅪ∶Ag were decreased to 1%and 1.3%,respectively.The APTT of her father,mother,son and daughter was 42.1 s,43.0 s,42.5 s and 41.0 s,respectively.The FⅪ∶C and the FⅪ∶Ag of them were almost half of the normal values.The APTT,FⅪ∶C and FⅪ∶Ag of her husband were all normal.Genetic testing revealed that the proband has carried a heterozygous missense c.1103G>A(p.Gly350Glu)variant in exon 10 and a heterozygous missense c.1556G>A(p.Trp501stop)variant in exon 13 of the F11 gene.The father and daughter were heterozygous for the c.1103G>A variant,whilst the mother and son were heterozygous for the c.1556G>A variant.Both Gly350 and Trp501 are highly conserved among homologous species,and both variants were predicted to be"disease causing"by Mutation Taster.Protein modeling indicated there are two hydrogen bonds between Gly350 and Phe312 in the wild-type,but the p.Gly350Glu variant may add a hydrogen bond to Glu and Tyr351 and create steric resistance between the two,both may affect the structure and stability of protein.Conclusion The c.1103G>A and c.1556G>A compound heterozygous variants probably underlay the pathogenesis of congenital FⅪdeficiency in this pedigree.