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目的探讨AT-Ⅲ基因突变及多态性与女性颅内静脉系统血栓形成(cerebral venous thrombosis,CVST)的相关性。方法采集南方医院生育期女性CVST住院患者50例及严格配伍的健康女性52例外周静脉血。采用ELISA法及发色底物法分别检测AT-Ⅲ抗原水平(AT-Ⅲ:Ag)、AT-Ⅲ血浆活性水平(AT-Ⅲ:A)。提取所有受试者全血DNA,PCR扩增后应用变性高效液相色谱(denaturing high performance liquid chromatography,DHPLC)技术分析抗凝血酶-Ⅲ(antithrombin,AT-Ⅲ)基因的启动子区,第1～6外显子及其侧翼序列的基因变异情况。结果病例组与对照组比较,AT-Ⅲ抗原含量水平相当(F=0.754,P=0.472),AT-Ⅲ活性明显降低(P<0.001)。DHPLC筛查发现突变位点1个已报道的致病突变Intron1 5+5 G>A突变,SNP位点5个,其中4个为SNP库已报道的SNP位点,1个为新发现的SNP位点。对照组亦发现4个为SNP库已报道的SNP位点。结论AT-Ⅲ基因突变可能是导致生育期女性CVST的遗传因素之一,功能性SNP位点也可能参与了该人群CVST的发生。
Objective To investigate the association between AT-Ⅲ gene mutation and polymorphism in female patients with cerebral venous thrombosis (CVST). Methods A total of 50 CVST female patients and 52 healthy volunteers with peripheral blood were collected from the Southern Hospital. The AT-Ⅲ antigen level (AT-Ⅲ: Ag) and AT-Ⅲ plasma level (AT-Ⅲ: A) were detected by ELISA and chromogenic substrate method respectively. All subjects’ whole blood DNA was extracted. After PCR amplification, the promoter region of antithrombin (AT-Ⅲ) gene was analyzed by denaturing high performance liquid chromatography (DHPLC) Gene variation of exons 1-6 and their flanking sequences. Results Compared with the control group, the level of AT-Ⅲ antigen was similar (F = 0.754, P = 0.472) and AT-Ⅲ activity was significantly lower (P <0.001). DHPLC screening found a mutation in the mutation site Intron1 5 +5 G> A mutations, SNP sites 5, of which 4 SNP sites have been reported SNP sites, a newly discovered SNP Site. The control group also found 4 SNP sites that have been reported for the SNP pool. Conclusions AT-Ⅲ gene mutation may be one of the genetic factors that lead to CVST in female during birth. Functional SNP may also be involved in the occurrence of CVST.