家族性高胆固醇血症患儿及其家系基因分析

Gene Mutation Analysis of A Child with Familial Hypercholesterolemia and His Family

目的通过对1例伴血清胆固醇水平极度增高的疑诊为家族性高胆固醇血症(FH)的多发性皮肤黄瘤病患儿及其2代家系成员进行基因检测和系谱分析,探讨多发性皮肤黄瘤病的发病机制。方法收集1例多发性皮肤黄瘤病患儿的家系2代共5例血标本及临床资料,对其家系成员进行血脂测定;对患儿行心电图、血管超声、心脏彩超和冠脉CT造影检查;酚-氯仿抽提法提取患儿及家系成员基因组DNA,应用PCR-单链构象多态性(PCR-SSCP)分析结合DNA直接测序方法 ,检测其低密度脂蛋白受体(LDLR)基因的全部18个外显子和启动子,以及载脂蛋白B(apoB100)R3500Q位点;核苷酸序列分析结果与GenBank比对,寻找突变。结果 1.该患儿父母胆固醇水平均高于正常上限。2.该患儿双侧颈总动脉分叉处中段内-中膜轻度增厚,局部可见强回声斑块;左房轻度增大,二尖瓣轻度返流,冠状动脉血流储备减低;冠状动脉前降支中段轻-中度狭窄。3.该家系排除apoB100基因R3500Q突变。4.核苷酸序列分析证实先证者LDLR基因第13外显子发生D601Y纯合突变,为1864位G>T碱基置换,导致天冬氨酸改变为脯氨酸;患儿父母LDLR基因第13外显子均发生D601Y杂合突变;患儿2个姐姐未检测到突变。结论该例多发性黄瘤病患儿确诊为纯合子FH,其LDLR基因存在D601Y纯合突变分别来源于父系和母系的遗传,该突变可能为该家系中FH的致病突变。

Objective To screen the mutation of certain gene of a 10 - years - old boy with multiple xanthomas and very high level of cholesterol who could be diagnosed as homozygous familial hypercholesterolemia （FH） ,to explore the relationship between the genotype and phenotype,and to discuss the molecular pathologic mechanism. Methods The basic information of life styles were asked from the boy and his familial members. The blood was drown to examine the lipid and genes. The boy was examined with electrocardiogram examination, uhrasonog- raphy and coronary CT angiography （CTA） to evaluate the degree of atherosclerosis. Peripheral blood DNA of the boy and his parents were extracted by phenol - chloroform method and investigated for mutations of promoter and all 18 exons of low density lipoprotein receptor（LDLR） gene. Screening was carried out by using Touch -down polymerase chain reaction （PCR） and single strand conformation polymorphism（ PCR - SSCP） ,combined with DNA sequence analysis. In addition,the apolipoprotein B100 gene（apoB100） for known mutations （R3500Q） which caused familial defective apoB100 was screened by PCR - DNA sequence analysis. Results 1. Tile level of cholesterol of his parents were higher than the normal. 2. Several clinical manifestations of atherosclerosis were detected from that boy. Increased intima - media thickness and plaques were detected in the common carotid artery. Mitral valve regurgitation was found by eehocardiography. Coronary stenosis was confirmed by CTA. 3. No mutations R3500Q of apoBl00 was observed. 4. A homozygous mutation in exonl3 of the LDLR gene （I）601Y） were identified in the boy and his parents harbour D601Y heterozygous mutation due to a single base pair substitution of G for T in the eodon for residue 1864. Conclusions The final diagnosis of the boy with multiple xanthomas was homozygous FH. His disease was caused by D601Y homozygous mutation in exonl3 of the LDLR gene inherited from his heterozygous parents.