DEVELOPMENT OF DRUG WHICH REGULATES BONE RESORPTION ACTIVITY IN OSTEOCLASTS VIA CYTOSKELTON AND SIGNAL TRANSDUCTION SYSTEM

通过细胞骨架和信号传导系统调节破骨细胞骨吸收活性的药物的开发

基本信息

批准号：
09470400
负责人：
OHYA Keiichi
金额：
$ 6.08万
依托单位：
Tokyo Medical and Dental University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

The aim of this study was to examine mechanisms of proton (H+ ) transport in active osteoclasts. We measured intracellular pH (pHi) changes in osteoclasts isolated on glasscoverslips or bone slices with a fluorescent pH indicator. H^+ was produced in osteoclasts by acid loading, and the pHi of the cells rapidly fell below the resting level. The decreased pHi did not recover in Na^+ free medium, but recovered in Na^+ rich medium. The pHi recovery was not observed on superfusion with Na+ rich medium containing amiloride, a potent sodium proton exchanger (NHE) inhibitor. The results reveal that osteoclasts on glass had H^+ transport system via NHE.In contrast, acid-loaded osteoclasts on bone showed their pHi recovery with amiloride and without Na+, presenting that active osteoclasts have H+ transport system other than NHE.From these results, osteoclasts have at least two different mechanisms of H+ transport compared to osteoclasts on glasscoverslips, suggesting that attachment to bone by osteoclasts activate intracellular signal transduction system.On the other hand, we evaluated the effects of bisphpsphonates on bone cells in rats fed with low calcium diet. Bone morphometrical analysis in femur and tibiae revealed that bisphpsphonates inhibited the increase of osteoclasts number, osteoclastic nucleus number, and osteoblasts number. Bisphpsphonates also inhibited the increment of mineralized nodule area and tartrate-resistant acid phosphatase positive-multi nuclear cells number. These results suggest that bisphpsphonates inhibit turnover in bone metabolism activated by low calcium diet.

本研究的目的是检查活性破骨细胞中质子 (H+) 运输的机制。我们使用荧光 pH 指示剂测量了玻璃盖玻片或骨切片上分离的破骨细胞的细胞内 pH (pHi) 变化。 H^+ 通过酸负荷在破骨细胞中产生，并且细胞的 pHi 迅速降至静息水平以下。降低的pHi在无Na^+培养基中没有恢复，但在富含Na^+培养基中恢复。在灌注含有阿米洛利（一种有效的钠质子交换器 (NHE) 抑制剂）的富含 Na+ 的培养基时，未观察到 pHi 恢复。结果表明，玻璃上的破骨细胞具有通过 NHE 的 H^+ 转运系统。相比之下，骨上的酸负载破骨细胞在阿米洛利和无 Na+ 的情况下显示其 pHi 恢复，表明活性破骨细胞具有除 NHE 之外的 H+ 转运系统。与玻璃盖玻片上的破骨细胞相比，破骨细胞至少有两种不同的 H+ 转运机制，表明破骨细胞附着在骨上激活了细胞内信号转导系统。另一方面，我们评估了双膦酸盐对低钙饮食喂养的大鼠骨细胞的影响。股骨和胫骨的骨形态测量分析表明，双磷酸盐抑制破骨细胞数量、破骨细胞核数量和成骨细胞数量的增加。双磷酸盐还抑制矿化结节面积和抗酒石酸酸性磷酸酶阳性多核细胞数量的增加。这些结果表明双磷酸盐抑制低钙饮食激活的骨代谢的转换。