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的机制知之甚少。衰老和激素的影响与FSAD广泛相关。雌激素主要被认为有助于在性反应期间控制生殖器血流。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，包括生殖器血管，知之甚少。雌激素的血管效应归因于其对内皮型一氧化氮合酶（eNOS）的调节，然而一氧化氮（NO）的生理作用和雌激素对女性生殖道中eNOS调节的分子机制在很大程度上是未知的。体外研究表明，内皮NO生成的增加并不一定需要eNOS丰度的增加，它可能是由eNOS的特异性翻译后修饰引起的。该提案的总体目标是研究雌激素调节女性生殖道eNOS的机制。我们将首先评估雌激素是否通过PI 3 K/Akt依赖性机制刺激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的相互作用并降低其与小窝蛋白-1的相互作用），并影响女性生殖道血管系统中eNOS与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的成年短期卵巢切除大鼠中进行测试，以确定雌激素对eNOS调控功能的影响，而不依赖于eNOS基因调控。接下来，我们将评估这些eNOS翻译后修饰的机制是否在衰老的背景下受到慢性雌激素缺乏的损害，以及早期与延迟长期雌激素给药是否可以保护eNOS和内皮功能。将在完整、卵巢切除和卵巢切除雌激素处理的成年、中年和老年大鼠中检验这一假设。这项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。与此同时，它可以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.