Sex differences in in vitro and in vivo glaucoma models may predict gender specific dose adjustment needs

项目摘要 原发性开角型青光眼（POAG）最明确的流行病学特征之一是， 发病率显示出强烈的性别差异。一氧化氮（NO）直接参与调节 青光眼的眼内压（IOP）我们的实验室目前正在设计，合成和测试 在啮齿动物模型中具有神经保护活性的新型混合型NO供体降低IOP。几 流行病学研究表明，雌激素水平与视网膜血流量增加呈正相关 循环，增加NO的合成，并与眼内压（IOP）水平呈负相关。雌激素增加 内皮型一氧化氮合酶（NOS）的活性，从而调节平滑肌张力， 血管阻力由于雌激素受体位于小梁网、睫状体和 眼的流出系统，它可能通过调节房水的产生和 外流因此，抗青光眼药物的反应可能存在性别依赖性差异， 从而同时递送NO沿着抗氧化活性，其干扰男性中雌激素信号传导 和妇女该项目的目标是确定NO捐赠的性别特异性影响， 抗氧化剂杂合化合物对小梁网（TM）细胞的抗氧化酶状态的影响， 细胞外基质成分、雌激素调节酶水平和总体IOP降低 活动 我们合成了一种新型的多功能抗氧化剂和NO供体的杂化化合物SA-2， SA-2化合物的其他几种新型高抗氧化衍生物。我们发表的结果由此而来 一个项目证明，一个单一的眼滴的纳米封装悬浮液的SA-2降低IOP的50%， 小鼠以及大鼠青光眼模型（雄性和雌性）。另外，化合物SA-2是 在缺氧处理的视网膜外植体中的离体神经保护和在体内IOP非依赖性视神经挤压中的神经保护 (ONC)在雌性小鼠中进行的模型。在我们使用人类小梁网（hTM）细胞的初步结果中， 我们发现SA-2在1mM剂量下的细胞保护活性存在性别依赖性差异。我们 进一步发现，当受到ONC损伤时，雌性小鼠的反应不同于雄性小鼠。 具体地，对于该NIH补充申请，我们建议使用来源于以下两者的原代人hTM细胞： 雌性和雄性供体（具体目标1）以及来自三个动物组的昏迷TM组织： 女性、男性和卵巢切除（OVX）女性，其中OVX程序模拟绝经后状态 （具体目标2）。我们推测，化合物SA-2及其新型类似物SA-10将具有不同的细胞毒性， 以及取决于性别和雌激素相关信号的分子活性。成功完成 提出的研究将对开发青光眼的新疗法具有意义， 性别差异，以帮助设计更个性化的治疗沿着性别特异性剂量调整。

PROJECT SUMMARY One of the most definite epidemiological characteristics of primary open angle glaucoma (POAG) is that, its incidence shows a strong sex-related difference. Nitric oxide (NO) is directly implicated in the regulation of intraocular pressure (IOP) in glaucoma. Our laboratory is currently working on design, synthesis and testing of a novel hybrid class of NO donors to lower IOP with neuroprotective activity in rodent models. Several epidemiological studies have suggested that estrogen levels correlate positively to increased retinal blood circulation, increased NO synthesis and negatively with intra ocular pressure (IOP) levels. Estrogen increases the activity of endothelial-based nitric oxide synthase (NOS) and therefore regulates smooth muscle tone and vascular resistance. Since estrogen receptors are located in the trabecular meshwork, ciliary body and the outflow system of the eye, it might potentially influence IOP by regulating both aqueous humor production and outflow. Consequently, there may be gender-dependent differential response to anti-glaucoma drugs especially to that simultaneously deliver NO along with antioxidant activity which interfere with estrogen signaling in men and women. The goal of this project is to determine the gender specific effects of the NO donating- antioxidant hybrid compounds on the trabecular meshwork (TM) cell’s anti-oxidant enzyme status, extracellular matrix composition, estrogen regulating enzyme levels and on the overall IOP lowering activity. We have synthesized a novel multifunctional hybrid anti-oxidant and NO donating compound SA-2 as well as several other novel highly antioxidant derivatives of SA-2 compound. Our published results ensued from this project demonstrated that a single eye drop of a nano encapsulated suspension of SA-2 lowered IOP by 50% in mouse as well as in rat glaucoma model (in both males and females). Additionally, compound SA-2 is neuroprotective ex vivo in hypoxia treated retinal explants and in the in vivo IOP-independent optic nerve crush (ONC) model performed in female mice. In our preliminary result using human trabecular meshwork (hTM) cells, we found that there is a gender dependent difference in the cytoprotective activity of SA-2 at 1mM dose. We further found that, the female mice respond differently than male mice when subjected to ONC injury. Specifically, for this NIH supplement application, we propose to use primary human hTM cell derived both from female and male donors (specific aim 1) as well as in glaucomatous TM tissues from three animal groups: females, males and ovariectomized (OVX) females, in which OVX procedure models a postmenopausal state (specific aim 2). We hypothesize that, compound SA-2 and its novel analog SA-10 will have differential cellular and molecular activities depending on the gender and estrogen-related signaling. Successful completion of the proposed studies will have an implication for developing novel therapeutics for glaucoma with emphasis to a gender differences to help design more personalized therapies along with gender specific dose adjustment.