血管性血友病因子Ala737→Glu突变体的构建和表达  

Construction and transient expression of an Ala 737→Glu mutant of vWF

作　　者：王迎春[1] 张敬宇[1] 万海英[1] 张威[1] 阮长耿[1] 

机构地区：[1]苏州医学院附属第一医院,江苏省血液研究所215006

出　　处：《中华血液学杂志》2000年第5期256-259,共4页Chinese Journal of Hematology

基　　金：国家自然科学基金!( 3 9670 3 3 1)

摘　　要：目的　研究血管性血友病因子 (vWF)Ala737→Glu突变对vWF功能的影响 ,分析vWFAla737→Glu突变致 2A型血管性血友病 (vWD)的分子病理机制。方法　用聚合酶链反应 (PCR)的方法对野生型的vWF全长表达质粒 (pSVvWF)进行体外定点诱变 ,在COS 7细胞中进行表达。分析其表达上清和细胞裂解液中的vWF∶Ag和上清的vWF的多聚物 ,观察患者血小板中的vWF多聚物及患者对DDAVP(糜凝 )的治疗反应。结果　vWFAla737→Glu的突变体上清vWF∶Ag是野生型的 76 .4% ,裂解液中vWF∶Ag是野生型的 98.8%。vWF突变体的多聚物形态和野生型相似 ,患者血小板中的vWF多聚物存在全部分子量的形式 ,糜凝治疗后vWF∶Ag明显升高。结论　vWFAla737→Glu的突变没有影响vWF的分泌和转运 ,vWFAla737→Glu突变致第Ⅱ类 2A型vWD。Objective To study the functional change of an Ala737→Glu substitution mutant of vWF and the molecular pathological mechanism of this mutant in the type 2A vWD. Methods The expression plasmid pSVvWF containing full length cDNA of vWF was used to site direct mutagenesis by polymerase chain reaction(PCR) and transitorily expressed in the COS 7 cells, vWF:Ag in the supernatant and the cell lysate between wild type and pSVAla737Glu vWF were measured. vWF polymers of the platelet of the patient with the mutation and response to DDAVP treatment of the patient were observed. Results vWF:Ag level of pSVAla 737Glu vWF was 76.4% and 98.8% of the wild types in the supernatant and cell lysate, respectively. The polymer pattern of extracellular pSVAla737Glu vWF was indistinguishable from that of the wild type, containing all kinds of molecular weight. Increased vWF antigen levels was observed in the patients after DDAVP treatment. Conclusion This mutant did not change the assembly and secretion of vWF. The mutant of vWF Ala737→Glu resulted in Group Ⅱ type 2A vWD.

关 键 词：血管性血友病 VWF 突变体 构建 基因表达 

分 类 号：R554.1[医药卫生—血液循环系统疾病]