INHIBITORS FOR VACUOLAR THPE HィイD1+ィエD1-ATPaseINDUCES APOTOSIS IN OSTEOCLASTS

空泡 THPE D1+D1-ATP 酶抑制剂诱导破骨细胞凋亡

• 批准号: 10671729
• 负责人: OKAHASHI Nobuo
• 金额: $ 1.66万
• 依托单位: NATIONAL INSTITUTE OF INFECTIOUS DISEASES
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671729/
• 关键词: OSTEOCLAST; APOPTOSIS; PERIODONTITIS; CASPASE; LIPOPOLYSACCHARIDE; CYTOKINE; プロトンATPase

Osteoclasts are multinucleated bone resorbing giant cells. Osteoclasts resorb bone minerals by acid. Production of acid by osteoclast is controlled by carbonic anhydrase and vacuolar type ATPase. Recently, we found that the inhibitors of vacuolar type ATPase trigger apoptotic cell death of osteoclasts. In this research project, we investigate the cellular mechanism of apoptotic events induced by the ATPase inhibitors.Purified osteoclasts were prepared from mouse bone marrow cultures. Vacuolar type ATPase inhibitors, concanamycin A and bafilomycin induced cell death of purified osteoclast within 24 h. Hoechst stain and FITC-TUNEL revealed apoptotic features of this cell death of osteoclasts. Measurement of caspase activities revealed that ATPase inhibitors activated caspase-1 and caspase-3 during apoptotic events of osteoclasts. Furthermore, activation of caspase-8 and caspase-9 also detected. Fluorescenece labeling of osteoclastic mitochondorial membrene by MitAlert revealed that the mitochondoria membrane was destroyed by the ATPase inhibitors. These results suggested that vacuolar type ATPase plays an important role in bone resorption as well as survival of osteoclasts.

破骨细胞是多核骨吸收巨细胞。破骨细胞通过酸吸收骨矿物质。破骨细胞产酸受碳酸酐酶和空泡型ATP酶控制。近年来，我们发现空泡型ATP酶抑制剂可诱导破骨细胞凋亡。本研究旨在探讨ATP酶抑制剂诱导破骨细胞凋亡的细胞机制。极性ATP酶抑制剂康卡霉素A和巴弗洛霉素在24 h内诱导破骨细胞死亡。Hoechst染色和FITC-TUNEL显示破骨细胞的凋亡特征。caspase活性的测量显示，ATP酶抑制剂激活caspase-1和caspase-3在破骨细胞凋亡事件。此外，还检测到caspase-8和caspase-9的活化。MitAlert荧光标记显示ATP酶抑制剂可破坏线粒体膜。提示空泡型ATP酶在破骨细胞的生存和骨吸收过程中起重要作用。

项目成果

Ohguchi,M.,et.al.: "Actinobacillus actinomycetemcomitans toxin induces both cell cycle arrest in the G2/M phase and apoptosis" Infect.Immun.66. 5980-5987 (1998)

Ohguchi,M.,et.al.：“Actinobacillus actinomycetemcomitans 毒素诱导细胞周期停滞在 G2/M 期并诱导细胞凋亡”Infect.Immun.66。

Koide, M., et al.: "Inhibition of experimental bone resorption and osteolast formation and survival by 2-aminoethansulfonic acid"Arch. Oral Biol.. 44. 711-719 (1999)

Koide，M.，等人：“2-氨基乙磺酸对实验性骨吸收和成骨细胞形成和存活的抑制”Arch。

Okahashi,N.,et.al.: "Specific inhibitors of vacuolar H-ATPase trigger apoptotic cell death of osteoclasts" J.Bone Miner.Res.12. 1116-1123 (1997)

Okahashi,N.,et.al.：“液泡 H-ATP 酶的特异性抑制剂触发破骨细胞凋亡”J.Bone Miner.Res.12。

Okahashi,N.,et.al.: "Caspases(interleukin-1b converting enzyme family proteases)are involved in the regulation of the survival of osteoclasts" Bone. 23. 33-41 (1998)

Okahashi,N.,et.al.：“半胱天冬酶（白细胞介素 1b 转换酶家族蛋白酶）参与破骨细胞存活的调节”骨。

Koide, M., et al.: "Bone morphogenetic protein-2 enhances osteoclast formation mediated by interleukin-1a through upregulation of osteoclast differentiation factor and cyclooxygenase-2"Biochem. Biophys. Res. Commun.. 250. 97-102 (1999)

Koide, M. 等人：“骨形态发生蛋白 2 通过上调破骨细胞分化因子和环氧合酶 2 来增强白细胞介素 1a 介导的破骨细胞形成”Biochem。