Posttranslational Regulation of eNOS by Estrogen in the Vagina and Clitoris

描述（由申请人提供）：女性性唤起障碍（FSAD）是女性性功能障碍的主要组成部分，影响25-70%的女性。FSAD的机制尚不清楚。衰老和激素的影响与FSAD广泛相关。雌激素主要被认为在性反应过程中有助于控制生殖器的血液流动。然而，性类固醇激素的分子机制，特别是雌激素在血管生理过程中的作用，包括生殖器脉管系统的作用，尚不清楚。雌激素对血管的作用归因于其对内皮型一氧化氮合酶（eNOS）的调节，但一氧化氮（NO）在女性生殖道中的生理作用和雌激素调节eNOS的分子机制在很大程度上是未知的。体外研究表明，内皮细胞NO生成的增加并不一定需要eNOS丰度的增加；这可能是eNOS翻译后特异性修饰的结果。本研究的总体目标是探讨雌激素对女性生殖道eNOS的调控机制。我们将首先评估雌激素是否通过PI3K/ akt依赖机制刺激eNOS磷酸化，影响eNOS与调节蛋白的相互作用（增加其与热休克蛋白-90的相互作用，减少其与小窝蛋白-1的相互作用），并影响eNOS与女性生殖道血管中RhoA/Rho-激酶信号通路之间的反调节平衡（抑制RhoA/Rho-激酶收缩通路）。这一假设将在急性雌激素治疗后短期去卵巢的成年大鼠中进行验证，以确定雌激素对eNOS独立于eNOS基因调控的eNOS调控功能的影响。接下来，我们将评估eNOS翻译后修饰的这些机制是否会因衰老背景下的慢性雌激素缺乏而受损，以及早期和延迟长期雌激素给药是否可以保护eNOS和内皮功能。这一假设将在完整、去卵巢和去卵巢雌激素治疗的成年、中年和老年大鼠中进行验证。本研究通过阐明雌激素对血管作用的新机制，有助于理解和治疗女性性唤起障碍。同时，它可能推进与内皮功能障碍和激素环境改变相关的多种疾病状态的治疗。

DESCRIPTION (provided by applicant): Female sexual arousal disorder (FSAD) is a major component of female sexual dysfunctions in general, affecting 25-70% of women. The mechanisms for FSAD are poorly understood. Aging and hormonal influences are widely associated with FSAD. Estrogen primarily is believed to contribute to the control of genital blood flow during the sexual response. However, the molecular mechanisms of sex steroid hormone, and specifically estrogen action in vascular physiological processes, including that of the genital vasculature, are poorly understood. Vascular effects of estrogen are attributed to its regulation of endothelial nitric oxide synthase (eNOS), yet the physiological role of nitric oxide (NO) and the molecular mechanisms of eNOS regulation by estrogen in the female genital tract are largely unknown. In vitro studies have shown that the increase in endothelial NO production does not necessarily require an increase in eNOS abundance; it may result from specific posttranslational modification of eNOS. The overall goal of the proposal is to investigate the mechanisms of estrogen regulation of eNOS in the female genital tract. We will first evaluate whether estrogen stimulates eNOS phosphorylation through PI3K/Akt-dependent mechanisms, influences eNOS interactions with regulatory protens (increases its interaction with the heat shock protein-90 and decreases its interaction with caveolin-1), and affects the counterregulatory balance between eNOS and the RhoA/Rho- kinase signaling pathway (inhibits the RhoA/Rho-kinase contraction pathway) in female genital tract vasculature. This hypothesis will be tested in adult short-term ovariectomized rats after acute estrogen treatment to determine the effect of estrogen on eNOS regulatory function independent of eNOS gene regulation. Next we will evaluate whether these mechanisms of eNOS posttranslational modification are impaired by chronic estrogen deficiency in the context of aging, and whether early versus delayed long-term estrogen administration may preserve eNOS and endothelial function. This hypothesis will be tested in intact, ovariectomized, and ovariectomized estrogen-treated adult, middle-aged, and aged rats. The proposed study applies to the understanding and treatment of female sexual arousal disorder by elucidating novel mechanisms of vascular effects of estrogen. At the same time it may advance treatments for multiple disease states associated with endothelial dysfunction and alterations in hormonal milieu.