凝血因子Ⅴ基因编码区单核苷酸多态性与静脉血栓栓塞症的相关性分析

The related analysis of venous thromboembolism and cSNPs of coagulation factor Ⅴ Gene

目的研究凝血因子Ⅴ（FⅤ）基因编码区单核苷酸多态性与静脉血栓栓塞症的关系。方法检测静脉血栓患者及正常对照者血浆中的FⅤ的活性（FⅤ：C）（一期法）及FⅤ抗原（FⅤ：Ag）（ELISA法），用Premier5软件设计5对与静脉血栓症相关的基因多态性（Asp79His、Arg306The、Arg306Gly、Arg506Gln、Ile359The和His1299Arg）引物，进行PCR扩增、产物纯化、限制性片段长度多态性聚合酶链反应（PCR-RFLP）检测。对所发现的多态性用直接测序法证实，并对含多态性的标本用直接测序法进行FⅤ基因全部外显子编码区的筛查。结果静脉血栓组（VTE组）FⅤ：C（106．9±28．0）％，FⅤ：Ag（110．4±33．3）％；对照组FⅤ：C（102．4±30．9）％，FⅤ：Ag（102．1±24．1）％。VTE组FⅤ：Ag明显高于对照组（P〈0．05），FⅤ：C两者之间差异无明显统计学意义。VTE组和对照组均无Asp79His、Arg306The、Arg306Gly、Arg506Gln和Ile359The多态性，5例静脉血栓栓塞症患者及3名正常人含有His1299Arg多态性。5例静脉血栓栓塞症患者同时伴有Met1736Val多态性，其中3例尚含Asp2194Gly多态性。结论FⅤ含量增高与静脉血栓栓塞症相关，静脉血栓栓塞症与Asp79His、Arg306The、Arg306Gly、Arg506Gln和Ile359The多态性基本无关；His1299Arg在VTE组中的分布频率高于对照组，但差异无统计学意义。Hisl299Arg与Metl736Ⅴal和As02194Gly多态性在人群中有连锁不平衡特性。

Objective To identify the relationship between coagulation factor Ⅴ （FⅤ） gene single nueleotide polymorphisms（SNPs） and venous thromboembolism（VTE）. Methods The FV clotting activity （ FⅤ ： C） and FⅤ antigen （ FⅤ ： Ag） in plasma of VTE group （ 111 patients ） and normol control （ 110 patients） were detected using one-stage clotting assay and ELISA, respectively. Five paires of primers of the FV polymorphisms including Asp79His, Arg306The, Arg306Gly, ArgS06Gln and Ile359The/His1299 Arg were synthesized and amplified by PCR. The PCR products were digested by restriction enzyme using PCR-RFLP. The detected polymorphisms were confirmed by direct sequencing. The samples containing the polymorphisms were screened for coding regions of all FⅤ exons with direct sequencing. Results The plasma levels of FⅤ ： C and FⅤ ： Ag of VTE group and normal control were （ 106.9 ± 28.0 ） % , （ 110.4 ± 33.3 ） % and （ 102.4 ± 30.9） %, （ 102.1 ± 24.1 ） %, respectively. The plasma level of FV： Ag was significantly different between VTE group and normal control. However, there was no difference in FⅤ： C levels. Polymorphisms for the forementioned 5 primer pairs were not found in either patients or normal controls. Polymorphism of His1299Arg was identified in 5 patients with VTE and 3 normal controls. And these 5 cases also combined Met1736Val polymorphism, 3 of them combined another Asp2194Gly polymorphism. Conclusion The higher plasma level of FV： Ag contribute to venous thromboembolism. There is no relationship between polymorphisms of Asp79His, Arg306The, Arg306Gly, Arg506Gln, Ile359The and venous thromboembolism in Chinese studied. Polymorphism His1299Arg is higher in VTE group than in normol control, but has no statistical difference. Polymorphisms of His1299Arg, Met1736Val and Asp2194Gly are linked disequilibrium in Chinese Han population.