遗传性异常纤维蛋白原血症致病基因鉴定及其致病机制的研究

Congenital dysfibrinogenemia is a genetic disease caused by fibrinogen gene defects results in abnormal fibrinogen structure and function. Two methods were used to detect the activity and antigen of fibrinogen at the same time, which can improve the diagnostic efficiency of the CD. But misdiagnosis and missed diagnosis are still existed. 4 CD pedigrees were studied on our previously study in our region, analyzed fibrinogen defects and determines the gene mutations of CD. Different mutations have a close relationship with clinical manifestations of CD. So identification the disease-causing gene is the key of diagnosis, treatment and prevention. In our project, PCR and its derived techniques, high resolution melting analysis, DNA sequencing and others methods were used to establish genetic pedigree of CD patient’s pathogenic gene mutations. Western Blot was used to detect the content of fibrinogen and whether three peptides of fibrinogen migrate abnormally, confirming fibrinogen defect types. We analyze the aggregation function, coagulation of fibrinogen and fibrinolysis function of fibrin. Our project study the disease-causing gene and pathogenic mechanism of CD on gene, protein, and function, which will contribute to gene diagnosis method of defect fibrinogen, explore fibrinogen defect types and the gene mutations spectrum, build up the foundation for diagnosis, treatment and prenatal antenatal examination of CD.

遗传性异常纤维蛋白原血症（CD）是一种纤维蛋白原（Fg）基因缺陷导致Fg结构异常与功能缺陷的遗传性疾病。应用2种方法同时测定Fg活性和抗原浓度，可提高CD诊断效率，但仍有一些患者漏诊或误诊。我们在前期工作中，对本地区4个CD家系进行了Fg缺陷与基因突变的初步研究。不同基因突变位点与CD临床表现有着密切的关系。因此，致病基因鉴定才是CD诊断与治疗的关键。本课题拟应用PCR及其衍生技术、高分辨熔解曲线分析和DNA测序等方法，鉴定本地区人群CD的致病基因；应用Western Blot技术检测CD患者Fg三条肽链的相对表达量以及三条肽链有无异常迁移，以明确Fg缺陷类型；分析Fg聚集、凝固及纤维蛋白溶解功能。本课题从基因、蛋白及功能方面，对本地区CD的致病基因及其致病机制进行研究，将有助于改进我国CD患者的缺陷Fg基因诊断方法，探索本地区CD患者的Fg缺陷类型与基因突变谱，为CD诊断、治疗及产前优生检查奠定基础。

遗传性异常纤维蛋白原血症（CD）是一种纤维蛋白原（Fg）基因缺陷导致Fg分子结构异常与功能缺陷的遗传性疾病。临床上，CD患者往往容易被漏诊或误诊误治。本课题组通过收集CD患者，分别用两种方法（PT演算法与Clauss法）检测CD患者Fg浓度；应用PCR及其衍生技术、DNA测序等方法，鉴定CD患者的致病基因；应用SDS-PAGE和Western Blot技术检测CD患者Fg三条肽链的相对表达量以及三条肽链有无异常迁移，明确Fg缺陷类型；进行Fg聚集、凝固率及纤维蛋白溶解功能检测；应用血栓弹力图（TEG）完整地监测CD患者凝血功能。研究结果显示：（1）当Fg PT演算法/Clauss法比值＞1.43诊断CD的灵敏度与特异度为100%；（2）本地区CD患者以FGA基因Arg16和FGG基因Arg275突变位点为主，同时还发现了FGA基因Gly13Arg、FGG基因Ala327Val两个新CD突变位点以及一个新的FGG基因Cys165Arg杂合突变的遗传性低纤维蛋白原血症位点，均为国内或国际首次报道；（3）大部分CD患者均有Fg聚集功能受损和Fg凝固率降低，有血栓倾向的CD患者其Fg溶解率下降；（4）与正常对照组比较发现CD患者TEG的K值明显延长、Angle值明显减低，反映了CD患者的纤维蛋白原功能活性降低以及纤维蛋白块形成及相互聚集的速度减慢。并通过ROC曲线分析，发现当TEG的K＞2.5min、Angle≤59.3°时有助于辅助诊断CD。以上研究成果有利于改进CD实验诊断方案，同时丰富我国CD的遗传学数据，明确本地区CD患者 的Fg缺陷类型与基因突变谱，为CD诊断、治疗及产前优生检查奠定基础。

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