家族性高胆固醇血症家系低密度脂蛋白受体活性及其基因突变分析

目的:对1例临床诊断为家族性高胆固醇血症(FH)纯合子患者家系进行低密度脂蛋白受体(LDL-R)活性研究及其基因突变分析,旨在探讨其分子病理机制。方法:对先证者及其父母进行血脂、颈动脉超声等检查;采用流式细胞仪检测先证者及其父母外周血淋巴细胞LDL-R表达量和功能;以先证者基因组DNA为模板,扩增载脂蛋白B100(ApoB100)基因3500区域片断,PCR产物进行核苷酸序列分析,排除由该突变导致家族性ApoB100缺陷症;用降落式PCR方法,在同一程序中分别扩增LDL-R基因的启动子和全部18个外显子片段,扩增产物进行核苷酸序列分析;发现突变后验证其父母是否存在相同基因突变。结果:先证者血清TC和LDL水平明显升高、颈动脉内中膜明显增厚;LDL-R的表达量和结合功能显著下降(分别为正常人的13.6%和21.1%);ApoB100基因3500区域未见突变,LDL-R基因第1864位碱基T取代了G发生纯合错义突变,导致第601位氨基酸天冬氨酸变成脯氨酸(D601Y)。其父母LDL-R基因发现了相同的杂合突变,经检索该突变在我国未见报道。结论:证实家系先证者存在LDL-R基因突变,分别来源于父系和母系的遗传,该突变可能是家系发病的分子基础;D601Y也可能是我国FH患者LDL-R基因一种新的突变类型。

Analysis of LDL receptor function and gene mutations in familial homozygous hypercholesterolemia

Objective:To screen the point mutation of LDL receptor gene in Han familial hypercholesterolemia(FH) patients,and characterize the relationship between the genotype and the phenotype for discussing the molecular pathologic mechanism.Method:We investigated a Han family with the clinical phenotype of homozygous FH,and detected the LDL receptor function in peripheral blood lymphocyte of the proband and his parents.Then we screened theirs gene mutations of promoter and all eighteen exons of LDL receptor(LDL-R) gene.Screening was carried out by touch-down PCR and agarose gel electrophoresis,combined with DNA sequence analysis.Then the result was compared with that in FH database.In addition,we screened the apolipoprotein B gene for known mutations(R3500Q) that caused familial defective Apo!B-100.Result:No mutation R3500Q of ApoB100 was observed.A homozygosis mutation in exon13 of the LDL-R was identified in the proband(D601Y) and his parents.The mutation has not been published in Han FH.Conclusion:The presence of the mutations at D601Y in LDL-R gene may be the cause of Han FH.Perhaps it is a particular pathogeneticgenotype for Chinese people.