The Aging Pituitary/Gonadal Axis

摘要 绝经过渡期的开始缺乏合适的诊断标志物。过渡 伴随着显著的发病率，包括不规则的生殖周期，功能失调性子宫出血， 泌尿生殖系统变化、生育能力受损、骨量下降、血管扩张症状和心理 损伤该项目的长期目标是了解促卵泡激素的作用 (FSH)糖基化变体hFSH 21和hFSH 24在衰老中的作用。年轻女性拥有更多的低糖基 hFSH 21比完全糖基化的hFSH 24更少，而老年妇女拥有更少的hFSH 21。由于这两种糖型都存在于 在尿液中，两者都进入循环，在那里它们影响性腺和非性腺目标。在此前的融资中， 在此期间，我们确定hFSH 21在结合FSHR、激活性腺靶点方面比hFSH 24活性高得多 细胞，并在体内刺激卵巢。相比之下，hFSH 24比hFSH 21活性高得多， 激活破骨细胞分化。因此，我们的总体假设是，面对衰老的卵巢， 从hFSH 21到hFSH 24的额外转换进一步损害生育力并可能导致骨丢失。每个 该项目将通过以下具体目标测试这一假设的各个方面：1。确定 hFSH的部分糖基化导致FSHR结合显著增加的机制。我们 工作假设是hFSH 21和小分子结合改变FSHR构象，使更多的FSH 结合位点可用于两种糖型，从而增加生物活性。一个令人兴奋的元素是协作 超分辨率显微镜研究，将直接监测FSHR寡聚化。2.定义不同的 在传统的性腺靶点以及骨中，FSH糖型激活的信号传导途径， 非传统靶细胞。在这个目标下，项目2将比较小鼠或人的FSH糖型激活， 颗粒细胞或细胞系和破骨细胞前体。我们的工作假设是hFSH 21更活跃， 在性腺靶细胞中，hFSH 24的表达高于hFSH 24，而在非性腺靶细胞中，hFSH 24的表达高于hFSH 24。与年龄相关的转变 因此，糖型比例具有重要的生理学后果。3.使用遗传模型来研究 衰老卵巢中的FSH糖型与年龄相关的骨质流失在此目标下，FSH低糖基化 将由项目3评估糖型拯救Fshb缺失雌性小鼠免于不育的能力， 对骨质流失的影响。我们的工作假设是与年龄相关的从FSH 21到FSH 24的转变具有有害的 影响卵巢功能，但促进破骨细胞活化。这些项目将得到两个 科学核心核心B将为所有项目提供关键的FSH糖型制剂。核心C将协助 分析质谱数据，提供下一代测序（NGS）和分析NGS数据。 总之，这些项目将提供关于FSH糖型在生殖衰老中的作用的新知识， 可用于诊断绝经期过渡的开始，以及支持或反驳 FSH在老年妇女骨质流失中的作用存在争议。

Abstract The beginning of the menopausal transition lacks suitable diagnostic markers. The transition is attended by significant morbidity including irregular reproductive cycles, dysfunctional uterine bleeding, urogenital changes, impaired fertility, declining bone mass, vasomotor symptoms, and psychological impairment. The long-term objective of this project is to understand the role of follicle-stimulating hormone (FSH) glycosylation variants hFSH21 and hFSH24 in aging. Young women possess more hypo-glycosylated hFSH21 than fully-glycosylated hFSH24, while older women possess less hFSH21. As both glycoforms are found in urine, both enter the circulation where they affect gonadal and non-gonadal targets. In the previous funding period we determined that hFSH21 is much more active than hFSH24 in binding FSHR, activating gonadal target cells in vitro, and stimulating the ovary in vivo. In contrast, hFSH24 is much more active than hFSH21 in activating osteoclast differentiation. Thus, our overall hypothesis is, in the face of a senescing ovary, the additional switch from hFSH21 to hFSH24 further compromises fertility and may contribute to bone loss. Each project will test various aspects of this hypothesis via the following specific aims: 1. Determine the mechanism(s) by which partial glycosylation of hFSH leads to significantly increased FSHR binding. Our working hypothesis is that hFSH21 and small molecule binding alter FSHR conformation, making more FSH binding sites available to both glycoforms, thus increasing bioactivity. An exciting element is collaborative super resolution microscopy studies that will monitor FSHR oligomerization directly. 2. Define the different signaling pathways activated by FSH glycoforms in traditional gonadal targets as well as in bone, a nontraditional target cell. Under this aim, Project 2 will compare FSH glycoform activation of murine or human granulosa cells or cell lines, and osteoclast precursors. Our working hypothesis is that hFSH21 is more active than hFSH24 in gonadal target cells, while the reverse is true for non-gonadal target cells. The age-related shift in glycoform ratio thus has important physiological consequences. 3. Use genetic models to study the role of FSH glycoforms in the aging ovary and age-related bone loss. Under this aim, FSH hypo-glycosylated glycoforms will be evaluated by Project 3 for their ability to rescue Fshb null female mice from infertility and their effects on bone loss. Our working hypothesis is the age-related shift from FSH21 to FSH24 has deleterious effects on ovarian function, yet promotes osteoclast activation. These projects will be supported by two scientific cores. Core B will provide critical FSH glycoform preparations to all projects. Core C will assist in analyzing mass spectrometry data, provide next generation sequencing (NGS) and analyze NGS data. Together, the projects will provide new knowledge about the role of FSH glycoforms in reproductive aging that can be used to diagnose the onset of the menopausal transition, and evidence to support or refute the controversial role of FSH in bone loss in aging women.