肝豆状核变性患者Wilson病蛋白及其基因的改变  

Changes of Wilson disease protein and its genein Wilsond isease patients

作　　者：侯国庆[1] 梁秀龄[1] 黄帆[1] 陈嵘[1] 欧翠华[1] 杨春水[1] 王莹[1] 

机构地区：[1]中山医科大学附属第一医院神经科,广东省广州市510080

出　　处：《世界华人消化杂志》2000年第9期988-992,共5页World Chinese Journal of Digestology

基　　金：国家自然科学基金项目;No.39670270;卫生部临床学科重大项目;No.37091;中山医科大学"211工程"重点建设项目;广东省自然科学基金项目;No.960128.~~

摘　　要：目的探讨肝豆状核变性(WD)发病的分子生物学机制.方法通过肝活检获得 WD 患者的肝标本,利用体外胶原酶温育分离并培养肝细胞,对 WD 患者肝细胞 WD 蛋白进行Western-blot 蛋白印迹检测;同时采用荧光 PCR 技术筛查其WD 基因突变,并进行直接 DNA 测序.结果肝细胞蛋白印迹可见特异 WD 蛋白(M_r155 000条带)的表达.3例 WD 患者中 WD1的 M_r155 000条带密度无明显变化,WD2,WD3的特异条带密度降低,仅有对照组的36%,提示其 WD 基因肝内表达异常;荧光 PCR 筛查发现一例患者存在exon8 Arg778Leu 突变,DNA 直接测序证实为2333 G→T杂合突变.结论 WD 基因在 WD 患者肝细胞的表达存在异常,这可能与WD 基因 Arg778Leu 位点突变有关,直接检测 WD 基因产物有助于研究 WD 的病理机制.AIM To study the molecular biological pathogenesis of Wilson disease (WD). METHODS Hepatocytes were isolated from three WD patients' bioptic hepatocytic samples and cultured in vitro.WD protein,the gene putative encoded products were detected by SDS-PAGE in conjunction with Western blot in liver samples of three patients and two controls. Their genomic DNAs were analyzed by means of fluorescent PCR to screen the mutations on Arg778Leu locus of WD gene and by direct DNA sequence. RESULTS The putative specific WD proteins (M_r155 000 lanes) were expressed in human hepatocytes,WD proteins from WD patients WD2 and WD3 were found to be poorly expressed,and WD1 showed normal expression. WD3 appeared as heterozygote of Arg778Leu mutation by fluorescent PCR screening,and DNA sequence proved the mutation of 2273G→T of WD gene in WD3. CONCLUSION The abnormally expressed putative WD proteins from WD patients might be related to the Arg778Leu mutation,and direct determination of WD gene products would help probe into the pathogenesis of WD.

关 键 词：肝豆状核变性 蛋白质类 基因 分子生物学 

分 类 号：R742.4[医药卫生—神经病学与精神病学]