Investigating the molecular mechanism of P-gp/NHERF-1 network at feto maternal interface and role of paracrine signaling of EVs containing drug transporter proteins

摘要 妊娠期药物治疗是不可避免的，目前的干预策略并不成功 降低妊娠相关疾病的风险。更好地了解药物的转运机制 胎儿-母体接口(FMI)的参与是改善妊娠和新生儿结局所必需的。这个 药物转运蛋白的表达水平随着妊娠期的进展而降低。我们的数据 提示钠氢交换调节因子-1(NHERF-1)与外排转运蛋白相互作用 蛋白质，通透性糖蛋白(P-gp)，这种相互作用主要在胎膜中更多 而不是胎盘。与此数据相一致的是，外排药物机制在绒毛膜细胞中也较高。 胎盘滋养层细胞多于胎膜。另一方面，细胞外小泡来自于胎儿 膜参与旁分泌信号，引起母体蜕膜和 子宫肌层。此外，EV携带药物转运蛋白作为它们的货运蛋白，并参与 修饰母体细胞以进行转运蛋白功能活动。这项建议建立在这些观察的基础上； 核心假设是胎膜中的NHERF-1可能是调节药物的靶点 怀孕期间的交通工具。我们将确定P-gp/NHERF-1在胎儿体内的机制网络 母体接口使用创新的FM-解放军-片上器官(OOC)器件(AIM-1)。在目标1中，我们将使用 OOC模拟宫内环境以测试P-gp底物他克莫司在FMI中的动力学 正常的健康和疾病状况，以及NHERF-1的存在和不存在。的表达方式 P-gp和NHERF-1将通过FMI确定。来源于胎膜的EVS携带药物 转运蛋白，但它们在药物转运调节中的作用尚不清楚。在AIM-2中，我们将 探讨母体细胞摄取EVS的机制并确定其在药物调控中的作用 运输使用P-gp击倒小鼠。鉴于NHERF-1和来自胎儿的EVS的关键作用 膜调控药物转运具有重要的科学意义和临床意义。成功者 这项研究的完成将为了解胎膜TPS和EVS是如何在FMI中产生的提供新的见解 在妊娠期间超越胎盘界限的药物药代动力学中发挥作用，并将有助于 设计药物输送策略以治疗不良妊娠结局。

ABSTRACT Pharmacotherapy during pregnancy is inevitable and current interventional strategies are not successful in reducing the risks of pregnancy related disorders. A better understanding of the drug transport mechanisms involved at feto-maternal interface (FMi) is required to improve the pregnancy and neonatal outcomes. The expression levels of the drug transporter proteins decrease as the gestation period progresses. Our data suggested that sodium hydrogen exchange regulatory factor-1 (NHERF-1) interacts with efflux transporter protein, permeability glycoprotein (P-gp), and this interaction is predominantly more in the fetal membrane rather than the placenta. Consistent with this data, efflux drug mechanism is also higher in the chorion cells of fetal membrane than placenta trophoblast cells. On the other hand, extracellular vesicles derived from fetal membrane are involved in the paracrine signaling that induce inflammatory changes in maternal decidua and myometrium. Along with that, EVs carry drug transporter proteins as their cargo proteins and are involved in modifying maternal cells for transporter protein functional activities. This proposal builds on these observations; the central hypothesis is the NHERF-1 in the fetal membrane could be targeted to regulate the drug transportation during pregnancy. We will determine the mechanistic network of P-gp/NHERF-1 across fetal maternal interface using innovative FM-PLA - organ on chip (OOC) devices (Aim-1). In Aim 1, we will use the OOC that mimic an in-utero environment to test the kinetics of P-gp substrate, Tacrolimus across the FMi in normal healthy and disease conditions along with in presence and absence of NHERF-1. The expression of the P-gp and NHERF-1 will be determined across the FMi. EVs derived from the fetal membrane carry drug transporter proteins but their role in the regulation of drug transportation remains unknown. In Aim-2 we will explore the uptake mechanism of the EVs by maternal cells and determine their role in regulating drug transportation using P-gp knock down mice. Given the critical role of NHERF-1 and EVs from the fetal membrane regulates the drug transportation is of significant scientific and clinical importance. The successful completion of the study will provide novel insights into how the fetal membrane TPs and EVs derived at FMi have a role in drug pharmacokinetics during pregnancy beyond the boundaries of the placenta and will help to design drug delivery strategies to treat adverse pregnancy outcomes.