MOLECULAR MECHANISMS OF ESTROGEN-REGULATED BONE RESORPTION

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。这个项目的主要目的是加强我们对骨吸收增加的机制的理解，这伴随着雌激素缺乏。骨吸收参数在6个月大的对照女性、卵巢切除女性（OVX）、接受激素替代疗法（HRT）、接受钙通道拮抗剂（地尔硫卓、硝苯地平、维拉帕米）或接受这两种药物联合治疗的女性之间进行了比较。正如预期的那样，OVX大鼠表现出骨骼间室骨吸收率增加，HRT降低了这一效果。各种拮抗剂对无定形腔室的作用与雌激素拮抗，对钙化腔室的作用具有协同作用，表明不同的作用机制。钙通道拮抗剂对平均尾动脉压的影响如假设的那样进行，表明即使与HRT联合使用，与对照组相比，所有三类钙通道拮抗剂也能显著降低血压。设计用于检测尿液中各种骨降解因子（脱氧吡啶啉（DPD），吡啶啉（PYD）和螺旋肽）的ELISA测定提供了监测骨骼代谢的替代途径。OVX雌性小鼠DPD、PYD和螺旋肽的排泄明显增加，表明骨转换增加，而接受HRT的雌性小鼠与对照组无显著差异。钙拮抗剂的使用减少了DPD、PYD和螺旋肽的排泄，表明骨转换减少，其作用模式与雌激素相似。为了确定雌激素在成骨细胞样细胞（7F2和UMR-106）中引发这种作用的机制，在缺乏雌激素化合物（FBS和酚红）的富集培养基（OPTI-MEM）中进行了实验。与用FBS培养的对照相比，该方案没有改变细胞的活力或形态。实验结果表明，雌激素浓度范围（0.1nM ~ 1.9mM）在24小时孵育后可刺激细胞增殖，表明雌激素具有成骨活性。我们最初的基因组研究表明，雌激素上调了转化生长因子β诱导的早期生长反应（TIEG）和骨保护素（OPG）的转录。这些实验结果支持了雌激素在骨骼转换中的抗吸收作用的假设。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The primary objective of this project is to enhance our understanding of the mechanisms of increased bone resorption, which accompanies estrogen deficiency. Bone resorption parameters have been compared between control females six months of age, ovariectomized females (OVX), females receiving hormone replacement therapy (HRT), and females receiving a calcium channel antagonists (Diltiazem, Nifedipine, Verapamil), or females receiving a combination of the two agents. As expected OVX rats displayed an increased rate of bone resorption from both skeletal compartments and HRT reduced that effect. The actions of the various antagonists were antagonistic to that of estrogen on the amorphous compartment and synergistic on the calcified compartment indicating differing mechanisms of action. The effects of the calcium channel antagonists on mean tail arterial pressure have proceeded as hypothesized demonstrating that all three classes of calcium channel antagonists significantly lowered blood pressure as compared to controls even when given in combination with HRT. ELISA assays designed to detect various bone degradation makers (deoxypyridinoline (DPD), pyridinoline (PYD), and helical peptide) in urine have provided an alternate avenue by which to monitor skeletal metabolism. OVX females display a significant increase in the excretion of DPD, PYD, and helical peptide, indicating an increase in bone turnover, while those receiving HRT were not significantly different from controls. The utilization of calcium antagonists decreases the excretion of DPD, PYD, and helical peptide, indicating a decrease in bone turnover and suggesting a similar mode of action as estrogen. In order to determine the mechanism by which estrogen elicits this action in osteoblast-like cells (7F2 & UMR-106), experiments were carried out in an enriched media (OPTI-MEM) in the absence of estrogenic compounds (FBS & phenol red). This protocol did not alter the viability or morphology of the cells as compared to controls grown with FBS. Experimental results indicate that estrogen over a range of concentrations (0.1nM to 1.9mM) stimulates cell proliferation following 24 hr incubation, demonstrating estrogen's osteoblastic activity. Our initial genomic studies suggest that estrogen up regulates the transcription of Transforming Growth Factor Beta Inducible Early Growth Response (TIEG) and osteoprotegerin (OPG). These experimental findings support the hypothesis of an antiresorptive role for estrogen in skeletal turnover.