OPA1 an Estrogen-Mediated Modulator of Platelet Hyperactivation

强有力的流行病学证据表明绝经前雌激素暴露与静脉血栓风险增加有关 女性血栓形成，特别是在怀孕期间、产后初期以及服用药物的女性 含雌激素的口服避孕药（OCP）。该提案解决了链接的重大知识差距 雌激素具有血小板活化作用。过去的研究主要集中在雌激素如何改变止血和 纤溶系统。这项以血小板为重点的提案代表了双方之间富有成效的合作 爱荷华大学和犹他大学。我们对弗雷明汉队列和孕妇的初步研究， 揭示女性血小板 OPA1 水平升高，可预测冠状动脉疾病的风险，或与 妊娠晚期血小板活化增加。在血小板缺陷 OPA1 敲除 (KO) 小鼠中 我们发现，与雄性相比，雌性 OPA1KO 小鼠可以免受血栓形成。的 卵巢切除术后血栓形成表型得以逆转。根据这些初步发现，我们假设 OPA1 和 OPA1 活性调节因子 OMA1 和 YMEL1 在 雌激素依赖性方式和血小板中的 OPA1 水平预测雌激素调节血小板 激活。为了在 R61 阶段解决这一假设，目标 1 将确定以下分子机制： 使用具有血小板特异性的小鼠进行雌激素介导和 OPA-1 依赖性血小板活化调节 OPA1 缺失将产生 OMA1 和 YMEL1 血小板特异性 KO 小鼠，OPA1 活性增加。 目标 2 将确定血小板 OPA1、OMA1 和 YMEL1 表达，并将表达水平与血小板相关联 在怀孕和使用 OCP 的情况下激活。我们还将开发人类 iPSC 模型系统 生成 MK 以确定雌激素是否直接调节这些基因的表达。 R33阶段将 剖析女性 OPA1 增加与血小板活化之间的分子机制。目标1将 对从 OMA1 和 YMEL1 中分离的鼠血小板进行详细的生理和分子表型分析 KO 和 OPA 过表达转基因小鼠。此外，我们将研究增加的联系机制 OPA1 活性、线粒体能量学和血小板活化。目标 2 将通过以下方式补充动物研究 检查孕妇和孕妇的线粒体功能和血小板信号传导的变化 OCP。我们将从 iPSC 衍生的 MK 中基因去除 OMA1 和 YMEL1，以直接测试雌激素是否 以 OPA1 依赖性方式调节 MK 和血小板功能。通过整合体内和体外研究 在人类和小鼠模型中，我们的方法是转化性的、创新的并且对 RFA 高度敏感。 这些研究将首次确定怀孕期间雌激素暴露增加和 OCP 使用是否增加 重新编程 MK 和发育血小板以增加血栓并发症并确定新机制 对于这种增加的风险。这些研究的影响在于，它们可能会识别出可以预测的新生物标志物 易感女性在怀孕期间或使用 OCP 之前，血栓形成的风险增加。

Strong epidemiological evidence links pre-menopausal estrogen exposure to increased risk of venous thrombosis in women, particularly during pregnancy, in the immediate postpartum period, and in women taking estrogen-containing oral contraceptives (OCP). This proposal addresses a significant knowledge gap linking estrogens with platelet activation. Studies in the past have focused on how estrogens alter the hemostatic and fibrinolytic system. This proposal, which focuses on platelets, represents a productive collaboration between the Universities of Iowa and Utah. Our preliminary studies in the Framingham cohort and in pregnant humans, reveal increased platelet OPA1 levels in females, that predict risk of coronary artery disease, or correlate with increased platelet activation in the third trimester of pregnancy. In platelet-deficient OPA1 knockout (KO) mice we discovered that female OPA1KO mice were protected from thrombosis compared to males. The thrombosis phenotype was reversed after oophorectomy. Based on these preliminary findings, we hypothesize that OPA1 and regulators of OPA1 activity, OMA1 and YMEL1, modulate platelet activation in an estrogen-dependent manner and OPA1 levels in platelets predict estrogen modulated platelet activation. To address this hypothesis in the R61 phase, Aim 1 will determine the molecular mechanisms for estrogen-mediated and OPA-1-dependent modulation of platelet activation using mice with platelet specific OPA1 deletion and will generate OMA1 and YMEL1 platelet specific KO mice, with increased OPA1 activity. Aim 2 will determine platelet OPA1, OMA1, and YMEL1 expression and correlate expression levels to platelet activation in the setting of pregnancy and OCP use. We will also develop a human iPSC model system to generate MKs to determine if estrogen directly regulates the expression of these genes. The R33 phase will dissect the molecular mechanisms linking increased OPA1 with platelet activation in females. Aim 1 will perform detailed physiological and molecular phenotyping of murine platelets isolated from OMA1 and YMEL1 KO and OPA overexpressing transgenic mice. In addition, we will examine the mechanisms linking increased OPA1 activity, mitochondrial energetics and platelet activation. Aim 2 will complement animal studies by examining changes in mitochondrial function and signaling in platelets from pregnant women and women on OCPs. We will genetically ablate OMA1 and YMEL1 from iPSC-derived MKs to directly test if estrogen modulates MK and platelet function in an OPA1 dependent manner. By integrating in vivo and in vitro studies in humans and mouse models, our approach is translational, innovative and highly responsive to the RFA. These studies will determine for the first time whether increased estrogen exposure in pregnancy and OCP use reprogram MKs and developing platelets to increase thrombotic complications and identify novel mechanisms for this increased risk. The impact of these studies, is that they may identify novel biomarkers that may predict increased risk of thrombosis in susceptible females during pregnancy or prior to OCP use.