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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.

本研究的目的是研究质子（H+）的运输机制，在活跃的破骨细胞。我们用荧光pH指示剂测量了盖玻片或骨切片上分离的破骨细胞的细胞内pH（pHi）变化。破骨细胞通过酸负荷产生H^+，细胞的pH值迅速下降到静息水平以下。在无Na^+培养基中，降低的pHi不能恢复，但在富Na^+培养基中恢复。用含有阿米洛利（一种有效的钠质子交换剂（NHE）抑制剂）的富Na+培养基灌流时，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，而骨上的破骨细胞在加阿米洛利和不加Na+的情况下，其pHi恢复，表明活性破骨细胞具有除NHE以外的H ^+转运系统。提示破骨细胞在骨表面的粘附激活了细胞内信号转导系统。另一方面，我们研究了双膦酸盐对低钙饮食大鼠骨细胞的影响。骨形态计量学分析显示，双膦酸盐抑制破骨细胞数、骨细胞核数和成骨细胞数的增加。双膦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的增加。这些结果表明，双膦酸盐抑制营业额在骨代谢激活低钙饮食。