MOLECULAR MECHANISMS OF THE SKELETAL EFFECTS OF ESTROGEN IN OLD AGE

雌激素对老年骨骼影响的分子机制

• 批准号: 7094994
• 负责人: STAVROS C. MANOLAGAS
• 金额: $ 21.49万
• 依托单位: UNIV OF ARKANSAS FOR MED SCIS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7094994
• 关键词: aging; animal old age; antioxidants; apoptosis; biological signal transduction; bone density; bone imaging /visualization /scanning; cellular pathology; computed axial tomography; cytokine; developmental genetics; estrogen receptors; estrogens; female; free radical oxygen; genetically modified animals; glutathione; laboratory mouse; mitogen activated protein kinase; molecular pathology; nuclear factor kappa beta; osteoblasts; osteoclasts; osteoporosis; oxidative stress; tissue /cell culture

Estrogen loss increases osteoclastogenesis, decreases osteoclast and increases osteoblast apoptosis, and causes loss of bone by decreasing anti-oxidant defense in these cell types. In addition, an age-dependent decrease in bone strength and mass in ovary-intact C57BL/6 mice is temporally associated with an agedependent increase in the prevalence of osteoblast apoptosis, decreased glutathione reductase (GSR) activity and corresponding increases in the production of reactive oxygen species (ROS) in the bone marrow and the phosphorylation of p66shc in vertebrae. Based on this evidence, it is hypothesized that increased ROS levels with advancing age is a fundamental mechanism of the age-dependent decline of bone strength and mass, and loss of estrogens exaggerates the adverse effects of aging on bone by decreasing defense against ROS, thereby, contributing perpetually to the loss of bone mass and strength that persists for decades after menopause, and is associated with old age. Estrogens enhance defense against ROS by modulating the activity of anti-oxidant enzymes, attenuating ROS-mediated induction of cytokines, and modulating signaling cascades that have been implicated in the defense against oxidative stress, apoptosis, and aging. All these effects are mediated by extranuclear (nongenotropic) actions of the ER on cytoplasmic kinases and result in changes in the birth and lifespan of osteoblasts, osteoclasts, and osteocytes. To test these interrelated hypotheses, in vitro studies are proposed to investigate whether the effects of estrogens on osteoblast and osteoclast apoptosis, as well as on osteoclastogenesis, are, at least in part, a direct consequence of the ability of these hormones to: a) antagonize the actions of ROS by increasing the levels of glutathione; and/or b) counteract ROS-activated signals on cytoplasmic kinases, and/or downstream transcription factors and the resulting upregulation of cytokines. Further, the role of oxidative stress in the molecular and cellular mechanism responsible for the loss of bone strength and mass associated with aging and the extent to which estrogen deficiency and the resulting attenuation of defense against oxidative stress in osteoblasts or osteoclasts contributes to these changes will be studied in mice over-expressing GSR in osteoblasts or osteoclasts. Finally, mice in which ERa will be deleted from osteoblasts or osteoclasts by means of CreXLoxP recombination; mice heterozygotes for an ERa knock-in mutant which cannot bind to ERE; and mice over-expressing an ERa mutant, which is incapable of membrane localization and prevents kinase-mediated signaling of the endogenous receptor, will be used to examine whether the antioxidant actions of estrogens on osteoblasts and osteoclasts in vivo are ER-dependent and nongenotropic. This work should help us understand how women become increasingly more susceptible to fractures as they grow old. Moreover, it mav lead to the discovery of more effective treatments or even a cure for osteoporosis.

雌激素丢失会增加破骨细胞的生成，减少破骨细胞，增加成骨细胞的凋亡， 通过减少这些细胞类型中的抗氧化防御而导致骨骼丢失。此外，与年龄相关的 卵巢完整的C57BL/6小鼠骨强度和骨量的降低与年龄依赖性的时间相关性 成骨细胞凋亡率增加，谷胱甘肽还原酶降低 骨髓中活性氧簇(ROS)的活性和相应的增加 椎骨中p66shc的磷酸化。根据这一证据，可以假设增加了 ROS水平随年龄增长是骨强度随年龄增长而下降的基本机制 和体重，雌激素的损失，通过减少防御，夸大了衰老对骨骼的不利影响 对抗ROS，从而永久地促进骨量和力量的损失 绝经后几十年，与老年有关。雌激素通过以下途径增强对ROS的防御 调节抗氧化酶的活性，减弱ROS介导的细胞因子的诱导，以及 调节信号级联反应，参与防御氧化应激，细胞凋亡， 和衰老。所有这些效应都是由ER对细胞质的核外(非遗传性)作用所介导的 并导致成骨细胞、破骨细胞和骨细胞的出生和寿命的改变。为了测试 这些相互关联的假说、体外研究都是为了调查雌激素的作用 关于成骨细胞和破骨细胞的凋亡，以及关于破骨细胞的形成，至少在一定程度上是直接的 这些荷尔蒙能力的结果是：a)通过增加ROS的水平来对抗ROS的作用 和/或b)中和细胞质上的ROS激活的信号，和/或下游 转录因子和由此产生的细胞因子上调。此外，氧化应激在 与衰老相关的骨强度和骨量丧失的分子和细胞机制 以及雌激素缺乏的程度和由此导致的对氧化应激防御的减弱 在成骨细胞或破骨细胞中，对这些变化的贡献将在GSR过度表达的小鼠身上进行研究 成骨细胞或破骨细胞。最后，将ERA从成骨细胞或破骨细胞中删除的小鼠 CreXLoxP重组的方法；不能结合的Era敲入突变体的小鼠杂合子 以及过度表达Era突变体的小鼠，该突变体不能进行膜定位并阻止 激酶介导的内源性受体信号传递，将被用来检测抗氧化剂是否 体内雌激素对成骨细胞和破骨细胞的作用是ER依赖性和非遗传性的。这部作品 应该有助于我们理解女性如何随着年龄的增长变得越来越容易骨折。 此外，它可能会导致发现更有效的治疗方法，甚至是治疗骨质疏松症的方法。