Nordic Bioscience A/S, Herlev DK 2730, Denmark.
BMC Musculoskelet Disord. 2010 Jun 1;11:109. doi: 10.1186/1471-2474-11-109.
Normal osteoclasts resorb bone by secretion of acid and proteases. Recent studies of patients with loss of function mutations affecting either of these processes have indicated a divergence in osteoclastic phenotypes. These difference in osteoclast phenotypes may directly or indirectly have secondary effects on bone remodeling, a process which is of importance for the pathogenesis of both osteoporosis and osteoarthritis. We treated human osteoclasts with different inhibitors and characterized their resulting function.
Human CD14 + monocytes were differentiated into mature osteoclasts using RANKL and M-CSF. The osteoclasts were cultured on bone in the presence or absence of various inhibitors: Inhibitors of acidification (bafilomycin A1, diphyllin, ethoxyzolamide), inhibitors of proteolysis (E64, GM6001), or a bisphosphonate (ibandronate). Osteoclast numbers and bone resorption were monitored by measurements of TRACP activity, the release of calcium, CTX-I and ICTP, as well as by counting resorption pits.
All inhibitors of acidification were equally potent with respect to inhibition of both organic and inorganic resorption. In contrast, inhibition of proteolysis by E64 potently reduced organic resorption, but only modestly suppressed inorganic resorption. GM6001 alone did not greatly affect bone resorption. However, when GM6001 and E64 were combined, a complete abrogation of organic bone resorption was observed, without a great effect on inorganic resorption. Ibandronate abrogated both organic and inorganic resorption at all concentrations tested [0.3-100 microM], however, this treatment dramatically reduced TRACP activity.
We present evidence highlighting important differences with respect to osteoclast function, when comparing the different types of osteoclast inhibitors. Each class of osteoclast inhibitors will lead to different alterations in osteoclast quality, which secondarily may lead to different bone qualities.
正常的破骨细胞通过分泌酸和蛋白酶来吸收骨骼。最近对功能丧失突变影响这些过程的患者进行的研究表明，破骨细胞表型存在分歧。这些破骨细胞表型的差异可能直接或间接地对骨重塑产生次要影响，这一过程对骨质疏松症和骨关节炎的发病机制都很重要。我们用不同的抑制剂处理人破骨细胞，并对其功能进行了特征描述。
使用 RANKL 和 M-CSF 将人 CD14+单核细胞分化为成熟的破骨细胞。在存在或不存在各种抑制剂的情况下，将破骨细胞在骨上培养：酸化抑制剂（巴弗洛霉素 A1、二苯并庚酮、乙氧唑胺）、蛋白酶抑制剂（E64、GM6001）或双膦酸盐（伊班膦酸盐）。通过测量 TRACP 活性、钙、CTX-I 和 ICTP 的释放以及计数吸收陷窝来监测破骨细胞数量和骨吸收。
所有酸化抑制剂在抑制有机和无机吸收方面同样有效。相比之下，E64 抑制蛋白酶的活性强烈抑制有机吸收，但仅适度抑制无机吸收。GM6001 单独使用不会对骨吸收产生很大影响。然而，当 GM6001 和 E64 联合使用时，观察到有机骨吸收完全被阻断，而无机吸收的影响不大。伊班膦酸盐在所有测试浓度（0.3-100 μM）下都阻断了有机和无机吸收，但这种治疗会显著降低 TRACP 活性。
我们提供的证据强调了在比较不同类型的破骨细胞抑制剂时，破骨细胞功能方面的重要差异。每一类破骨细胞抑制剂都会导致破骨细胞质量的不同改变，这可能会导致不同的骨质量。
