Age-Dependent N-Glycosylation of Follicle-Stimulation Hormone in Gonadotropes

促性腺激素中卵泡刺激激素的年龄依赖性 N-糖基化

• 批准号: 10679254
• 负责人: Rosemary McDonald
• 金额: $ 1.97万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2023-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10679254
• 关键词: Address; Adipose tissue; Affect; Age; Aging; Anterior Pituitary Gland; Binding; Biochemical; Bioinformatics; Biological; Biological Assay; Bone Tissue; Cardiovascular Diseases; Cells; Clinical Treatment; Complex; Cre driver; Data; ESR1 gene; Enzymes; Estrogen Receptors; Estrogen decline; Estrogens; Etiology; Event; Exhibits; Feedback; Female; Fluorescence-Activated Cell Sorting; Follicle Stimulating Hormone; Gene Expression; Genes; Genetic; Glycoproteins; Goals; Gonadotrope Cell; Hormones; Human; Immunofluorescence Immunologic; Inflammatory; Investigation; Knock-out; Knowledge; Life Style; Longevity; LoxP-flanked allele; Luteinizing Hormone; Menopausal Symptom; Menopause; Messenger RNA; Metabolic Diseases; Molecular; Mus; Obesity; Organism; Osteoporosis; Ovarian; Ovarian Steroid Hormone; Ovarian aging; Pathway interactions; Perimenopause; Phenotype; Physiological; Pituitary Gland; Postmenopause; Premenopause; Process; Progesterone; Progesterone Receptors; Proteins; Receptor Signaling; Regulation; Research; Role; Signal Transduction; Stains; Steroid Receptors; Steroids; Symptoms; Tissues; Up-Regulation; Urine; Visceral; Weight Gain; Woman; Work; age related; aged; body system; bone; bone loss; common symptom; enzyme activity; enzyme pathway; experience; glycosylation; human old age (65+); hypothalamic pituitary gonadal axis; in vivo; mRNA Expression; middle age; mouse model; novel; older women; promoter; protein expression; receptor-mediated signaling; reproductive; reproductive senescence; steroid hormone receptor; targeted treatment; transcriptome sequencing; young woman

PROJECT SUMMARY The menopausal transition represents a complex and dynamic physiological aging phenomenon and often leads to adverse life-style effects. Nearly 85% of post-menopausal women will experience multiple symptoms. Treatment of clinical symptoms in post-menopausal women is difficult without knowing the underlying biological mechanisms and molecular players and hence requires further investigation. The anterior pituitary-derived heterodimeric glycoprotein hormone, follicle-stimulating hormone (FSH) consists of an alpha and a beta subunit. The FSH subunit undergoes significant age-dependent changes in N-glycosylation. Higher ratios of fully glycosylated FSHβ subunit in older, post-menopausal women compared to younger, reproductively active women were identified in human pituitaries and urine. FSH levels rise significantly due to concomitantly declining ovarian estrogen and progesterone during menopause and high levels of old age specific FSH may be deleterious in tissues such as bone and adipose, where FSH acts noncanonically via inflammatory pathways. This observation is of great significance because osteoporosis and weight gain are two of the most common symptoms post-menopausal women experience. This project focuses on the N-glycosylation process within anterior pituitary gonadotrope cells and seeks to identify the molecular players involved in the “glycosylation shift” to a more fully glycosylated phenotype in older age females, as well as the factors that regulate this process. We have recently identified differentially regulated N-glycosylation enzymes expressed in both young (~4- months) and older age (8-month) female mouse gonadotrope cells via RNA-sequencing analysis. Several glycosylation pathway-encoding genes (for example, Man2a1, B4galt5) exhibited a significant upregulation in gonadotropes of older age mice. We hypothesize that these N-glycosylation enzyme- encoding genes may contribute to the age-related “glycosylation shift” in FSHβ seen in older, post-reproductively active female mice. In Specific Aim 1, we will define age-dependent changes in expression of N-glycosylation enzymes MAN2A1 and B4GALT4 in gonadotrope cells of young and old female mice (using a GFP-tagged gonadotrope mouse model) and immunostaining whole pituitary and gonadotrope cells, as well as performing enzyme activity assays. In Specific Aim 2, we will determine the role of estrogen and progesterone receptor signaling in regulation of the N- glycosylation pathway-encoding enzymes in gonadotrope cells of young and old female mice using gonadotrope-specific knockout of either Esr1 or Pgr and performing fluorescence-activated cell sorting (FACS), RNA-sequencing, qPCR, immunostaining, and enzyme activity assays in pituitary gonadotrope cells. Ultimately, the work proposed here represents a novel genetic and biochemical approach and provide new knowledge on the regulation of FSHβ N-glycosylation during reproductive aging and how this may indirectly contribute to the etiology of the well-known menopausal symptoms such as bone loss and adiposity.

项目摘要 绝经过渡期是一种复杂的动态生理老化现象， 不良生活方式的影响。近85%的绝经后妇女会出现多种症状。 绝经后妇女的临床症状的治疗是困难的，不知道潜在的生物学 机制和分子球员，因此需要进一步调查。垂体前叶 作为异二聚体糖蛋白激素，促卵泡激素（FSH）由α和β亚基组成。 FSH β亚基的N-糖基化发生显著的年龄依赖性变化。充分利用的比率较高 绝经后老年女性与生育活跃的年轻女性的糖基化FSHβ亚基比较 在人类垂体和尿液中发现了女性。FSH水平显著上升，由于伴随下降 绝经期卵巢雌激素和孕激素以及高水平的老年特异性FSH可能是 在骨和脂肪等组织中是有害的，其中FSH通过炎症途径非规范地起作用。 这一观察结果意义重大，因为骨质疏松和体重增加是最常见的两种 绝经后妇女经历的症状。本项目的重点是N-糖基化过程中， 垂体前叶促性腺激素细胞，并试图确定参与“糖基化”的分子球员 在老年女性中，糖基化表型向更完全的糖基化表型转变，以及调节这一过程的因素。 我们最近发现，在两个年轻的（~4- 3个月）和更大年龄（8个月）的雌性小鼠促性腺细胞。几 糖基化途径编码基因（例如，Man 2a 1，B4 galt 5）在细胞凋亡中表现出显著的上调。 老年小鼠的促性腺激素。我们假设这些N-糖基化酶编码基因可能 有助于在年老的、生殖后活跃的雌性小鼠中观察到的FSHβ中与年龄相关的“糖基化转变”。 在具体目标1中，我们将定义N-糖基化酶MAN 2A 1表达的年龄依赖性变化 和B4 GALT 4在年轻和年老雌性小鼠的促性腺细胞中的表达（使用GFP标记的促性腺细胞小鼠 模型）和免疫染色整个垂体和促性腺细胞，以及进行酶活性测定。 在具体目标2中，我们将确定雌激素和孕激素受体信号转导在调节雌激素受体和孕激素受体信号转导中的作用。 用免疫组织化学方法检测年轻和老年雌性小鼠促性腺细胞中N-糖基化途径编码酶 促性腺激素特异性敲除Esr 1或Pgr并进行荧光激活细胞分选（FACS）， 垂体促性腺激素细胞中的RNA测序、qPCR、免疫染色和酶活性测定。最后， 本文提出的工作代表了一种新的遗传学和生物化学方法，并提供了关于 生殖衰老过程中FSHβ N-糖基化的调节以及这可能如何间接促进 众所周知的绝经期症状如骨质流失和肥胖的病因。