Study on direct action of estrogen on mature osteoclasts

雌激素缺乏，由更年期卵巢切除术引起，导致病理性骨质流失，这可以通过雌激素替代疗法来预防。尽管人们认为雌激素预防骨质流失的主要作用是通过抑制破骨细胞骨吸收，但这种作用的确切机制尚不清楚，这主要是由于纯化的功能性破骨细胞在技术上存在困难。在整个研究过程中，我们采用了我们最近开发的两种独特的高纯度哺乳动物破骨细胞的分离方法：一种是在塑料培养皿上纯化获得大量破骨细胞用于分子生物学分析，另一种是在细胞悬液中分离破骨细胞用于估计破骨细胞的骨吸收活性。在雌激素的生理浓度范围内（10^<-12>-10^<-10>M），雌激素抑制破骨细胞骨吸收活性。在相同浓度范围内，雌激素还降低了破骨细胞中大量特异性表达的组织蛋白酶K和破骨细胞中参与质子产生的碳酸酐酶II的mRNA水平。此外，雌激素诱导破骨细胞凋亡呈剂量和时间依赖性。ICI1164、384和他莫昔芬分别作为纯拮抗剂和部分拮抗剂，完全和部分阻断了雌激素抑制破骨细胞骨吸收和诱导破骨细胞凋亡的作用。最后，我们检测了雌激素受体（ERA） mRNA的表达，但没有检测到BRb mRNA的表达。这些数据表明，雌激素对绝经后骨质疏松症的保护作用部分是通过雌激素受体介导的机制直接减少破骨细胞骨吸收的关键酶和直接诱导破骨细胞凋亡。

Estrogen deficiency, cause by either menopause ovariectomy, results in pathological bone loss, which can be prevented by estrogen replacement therapy. Although it is believed that estrogen's main action in preventing bone loss is through inhibition of osteoclastic bone resorption, the precise mechanism of such effect is not clear, largely due to technical difficulties in obtaining purified functional osteoclasts. Throughout this study, we used two unique isolation methods of highly purified mammalian osteoclasts, which were recently developed by us : one is the purification on plastic dished to get a large number of osteoclasts for molecular biological analyzes, and the other is the isolation of osteoclasts in cell suspension for estimation of osteoclastic bone-resorbing activity. In a range of physiological concentrations of estrogen (10^<-12>-10^<-10>M), estrogen inhibited osteoclastic bone resorbing activity. In the same concentration range, estrogen also reduced mRNA levels of cathepsin K,which is abundantly and specifically expressed in osteoclasts, and of carbonic anhydrase II,that is involved in proton production in osteoclasts. Furthermore, estrogen induced osteoclast apoptosis in a dose-and time-dependent manner. ICI1164,384 and tamoxifen, as pure and partial antagonists, respectively, completely and partially blocked the effect of estrogen on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis. Finally, we detected the mRNA expression of estrogen reccptor (ERA), but not BRb. Thses data suggest that the protective effects of estrogen against postmenopausal osteoporosis are mediated in part by the direct reduction of key enzymes for osteoclastic bone resorption and the direct induction of osteoclast apoptosis via estrogen receptor-mediated mechanisms.