Isolated osteoclasts resorb the organic and inorganic components of bone.
Isolated osteoclasts resorb the organic and inorganic components of bone.
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DOI：
10.1083/jcb.102.4.1164
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发表时间：
1986-04
期刊：
影响因子：
--
通讯作者：
Teitelbaum SL
中科院分区：
文献类型：
--
作者：
Blair HC;Kahn AJ;Crouch EC;Jeffrey JJ;Teitelbaum SL
破骨细胞是骨的主要吸收细胞，但其降解骨基质主要有机成分胶原蛋白的能力尚不清楚。因此，我们以体内用L-[5-~3H]Pro或45Ca标记的失活大鼠骨和成骨样大鼠骨肉瘤细胞体外产生和矿化的类骨基质为底物，研究了高度浓缩的分离鸡破骨细胞群体的骨吸收活性。当与放射性标记基质共培养时，破骨细胞介导的矿物质动员在2 h内达到最大速率，而有机基质的降解速度较慢，在12-24 h达到最大速率，此后，当有多余的基质时，有机基质和无机基质的吸收速率基本上是线性的，至少持续6 d。氨基酸分析证实破骨细胞介导的骨胶原降解。溶解的氚有39%是以反式-4-羟基脯氨酸的形式回收的，47%是以Pro的形式回收的。10,000个破骨细胞在5天内溶解了100微克25-50微米骨片中总放射性的70%和[~3H]-反式-4-羟基脯氨酸的65%。几乎所有释放的氚标记蛋白都是低分子量，99%的MR小于或等于10,000，65%的MR小于或等于1,000。此外，当14%的被吸收的[~3H]Pro标记的多肽的MR大于或等于2,000时，当检测到哺乳动物胶原酶活性的特征初始产物TCA和TCB的存在时，在SDS PAGE中没有检测到。此外，破骨细胞条件培养液不具有溶胶原活性，外源性TCA和TCB片段不被破骨细胞降解。破骨细胞裂解产物在酸性缓冲液中具有胶原酶活性，而在中性缓冲液中不具有胶原酶活性，在pH值为4.0时活性最大。这些数据表明，破骨细胞具有通过定位于破骨细胞及其附着部位的过程吸收骨的有机相的能力。这一过程似乎独立于中性胶原酶的分泌，可能反映了酸性蛋白酶的活性。
Osteoclasts are the principal resorptive cells of bone, yet their capacity to degrade collagen, the major organic component of bone matrix, remains unexplored. Accordingly, we have studied the bone resorptive activity of highly enriched populations of isolated chicken osteoclasts, using as substrate devitalized rat bone which had been labeled in vivo with L-[5-3H]proline or 45Ca, and bone-like matrix produced and mineralized in vitro by osteoblast-like rat osteosarcoma cells. When co-cultured with a radiolabeled substrate, osteoclast- mediated mineral mobilization reached a maximal rate within 2 h, whereas organic matrix degradation appeared more slowly, reaching maximal rate by 12-24 h. Thereafter, the rates of organic and inorganic matrix resorption were essentially linear and parallel for at least 6 d when excess substrate was available. Osteoclast-mediated degradation of bone collagen was confirmed by amino acid analysis. 39% of the solubilized tritium was recovered as trans-4-hydroxyproline, 47% as proline. 10,000 osteoclasts solubilized 70% of the total radioactivity and 65% of the [3H]-trans-4-hydroxyproline from 100 micrograms of 25-50 micron bone fragments within 5 d. Virtually all released tritium- labeled protein was of low molecular weight, 99% with Mr less than or equal to 10,000, and 65% with Mr less than or equal to 1,000. Moreover, when the 14% of resorbed [3H]proline-labeled peptides with Mr greater than or equal to 2,000 were examined for the presence of TCA and TCB, the characteristic initial products of mammalian collagenase activity, none was detected by SDS PAGE. In addition, osteoclast-conditioned medium had no collagenolytic activity, and exogenous TCA and TCB fragments were not degraded by osteoclasts. On the other hand, osteoclast lysates have collagenolytic enzyme activity in acidic but not in neutral buffer, with maximum activity at pH 4.0. These data indicate that osteoclasts have the capacity to resorb the organic phase of bone by a process localized to the osteoclast and its attachment site. This process appears to be independent of secretion of neutral collagenase and probably reflects acid protease activity.