TFPI, Protein S, and Plasma FIXa in Hormone-Induced Hypercoagulability

静脉血栓栓塞症（VTE）在妇女服用口服避孕药（OC）是一个广泛的临床问题， 至少部分由雌激素对凝血因子表达的影响介导的显著性 肝脏或脉管系统中的蛋白质。我们的总体目标是识别和表征潜在的凝血 导致OC诱导的高凝状态的途径，并确定在个体中普遍存在的遗传因素， 最高的风险是静脉血栓栓塞。OC的使用降低了两种内源性抗凝蛋白的血浆浓度， 组织因子途径抑制物（TFPI）和蛋白S（PS），其调节血浆FIXa产生和活性。 我们建立了一种高灵敏度、高特异性（<10 pmol/L）的血浆FIXa测定方法 使用增强的凝血酶生成测定法测定活性，该测定法通过以下途径放大来自受试者血浆的FIXa信号： FIX缺陷血浆中凝血酶生成。在献血者的血浆样本中使用这种检测方法，我们发现 OC诱导的TFPI和PS的变化与血浆因子IXa（FIXa）活性升高有关， 约30%的女性服用OC。我们的中心假设是OC的使用改变了促凝血剂的抑制作用 天然抗凝剂TFPI和PS的反应，产生血浆因子IXa（FIXa）的大幅增加 活动和全身高凝状态。这一假设将被探讨，以确定生物化学和 OC诱导的高凝状态的遗传相关性。使用重组FIX变体的生化研究 抗凝血酶或PS结合将被用来定义TFPI和PS如何调节FIXa的产生， 人类血浆系统生化检查结果的临床影响将通过 TFPI、PS和FIXa的血浆水平测量、GWAS分析和血栓形成史问卷 在两个绝经前妇女队列中：1）约1000名绝经前献血者的横断面队列; 和2）一个纵向队列的约45个青春期女孩进行了为期一年的跟踪后，开始OC治疗。 将对横断面队列中收集的数据进行建模，以确定非遗传和遗传因素， 与OC相关的血浆TFPI、PS和FIXa差异以及血栓形成史相关。数据 在纵向队列中收集的数据将用于检查OC诱导的时间过程和稳定性 血浆高凝状态个体内重复测量分析。这些发现将用于 验证血浆蛋白变化和GWAS在横断面队列中鉴定的SNP的影响。 拟议的实验将确定OC诱导的超增殖的生物化学和遗传基础， 凝血状态集中在我们发现血浆中活化凝血因子FIXa的含量升高 大约30%的女性使用OC。这些结果也将为进一步的临床研究确定依据， 血浆FIXa和相关遗传标记物对OC诱导血栓形成和促进血栓形成预测价值 为考虑使用OC和相关激素治疗的妇女提供基于证据的决策。

Venous thromboembolism (VTE) in women taking oral contraceptives (OC) is a problem of broad clinical significance that is mediated, at least in part, by estrogen effects on the expression of blood coagulation proteins by the liver or vasculature. Our overall goal is to identify and characterize the underlying coagulation pathways leading to OC-induced hypercoagulability and to identify genetic factors prevalent in individuals at highest risk for VTE. OC use decreases the plasma concentration of two endogenous anticoagulant proteins, tissue factor pathway inhibitor (TFPI) and protein S (PS) that modulate plasma FIXa production and activity. We have developed a highly sensitive and specific (<10 pmol/L) technique for measurement of plasma FIXa activity using an enhanced thrombin generation assay, which amplifies the FIXa signal from subject plasma via thrombin generation in FIX-deficient plasma. Using this assay in plasma samples from blood donors we found that OC-induced changes in TFPI and PS are associated with elevated plasma factor IXa (FIXa) activity in ~30% of women taking OC. Our central hypothesis is that OC use alters the inhibition of procoagulant responses by the natural anticoagulants TFPI and PS, producing large increases in plasma factor IXa (FIXa) activity and a systemic hypercoagulable state. This hypothesis will be probed to identify biochemical and genetic correlates of OC-induced hypercoagulability. Biochemical studies using recombinant FIX variants resistant to antithrombin or PS binding will be used to define how TFPI and PS modulate FIXa generation in human plasma-based systems. The clinical impact of the biochemical findings will be examined by measurement of plasma levels of TFPI, PS, and FIXa, GWAS analysis, and a thrombosis history questionnaire in two cohorts of pre-menopausal women: 1) a cross-sectional cohort of ~1000 pre-menopausal blood donors; and 2) a longitudinal cohort of ~45 adolescent girls to be followed for one year after initiation of OC-therapy. The data collected in the cross-sectional cohort will be modeled to identify non-genetic and genetic factors that correlate with OC-associated differences in plasma TFPI, PS and FIXa and thrombosis history. The data collected in the longitudinal cohort will be used to examine the time course and stability of the OC-induced plasma hypercoagulable state within individuals using repeated measures analysis. These findings will be used to validate plasma protein changes and the effect of SNPs identified by GWAS in the cross-sectional cohorts. The proposed experiments will define the biochemical and genetic underpinnings of the OC-induced hyper- coaguable state focused on our finding of elevated amounts of an activated clotting factor, FIXa, in the plasma of about 30% of women using OC. These results will also establish rationale for further clinical studies to define the predictive value of plasma FIXa and associated genetic markers for OC-induced thrombosis and facilitate evidence-based decision-making for women considering use of OC and related hormonal therapies.