Tezuka K, Nemoto K, Tezuka Y, Sato T, Ikeda Y, Kobori M, Kawashima H, Eguchi H, Hakeda Y, Kumegawa M
Department of Oral Anatomy, Meikai University School of Dentistry, Saitama, Japan.
J Biol Chem. 1994 May 27;269(21):15006-9.
Osteoclasts are multinucleate giant cells that play key roles in bone resorption. To identify genes predominantly expressed in osteoclasts, we screened a cDNA library of osteoclasts with cDNA probes of osteoclasts and alveolar macrophages. Clones specifically hybridizing to the osteoclast probe were isolated and sequenced. The nucleotide sequence of one such clone, F17, was found to share significant similarity with the sequences of human and mouse matrix metalloproteinase 9 (MMP-9) cDNA. By isolation and sequencing of the full-length cDNA, F17 was revealed to encode the rabbit counterpart of MMP-9. By Northern blotting, messenger RNA for MMP-9 was found to be highly and predominantly expressed in isolated osteoclasts when compared with its level in alveolar macrophages and other tissues. By gelatin zymography, gelatinase activity was detected in conditioned medium of isolated osteoclasts, suggesting that MMP-9 is secreted by isolated osteoclasts. Expression of MMP-9 was also observed in in vivo osteoclasts in metacarpal bones of newborn rabbits by in situ hybridization. These facts suggest that MMP-9 is one of the major proteases produced by osteoclasts and possibly plays a role in osteoclastic bone resorption.
破骨细胞是在骨吸收中起关键作用的多核巨细胞。为了鉴定主要在破骨细胞中表达的基因，我们用破骨细胞和肺泡巨噬细胞的cDNA探针筛选了破骨细胞的cDNA文库。分离出与破骨细胞探针特异性杂交的克隆并进行测序。发现其中一个这样的克隆F17的核苷酸序列与人和小鼠基质金属蛋白酶9（MMP-9）cDNA的序列有显著相似性。通过全长cDNA的分离和测序，发现F17编码MMP-9的兔对应物。通过Northern印迹法，与肺泡巨噬细胞和其他组织中的水平相比，发现MMP-9的信使核糖核酸在分离的破骨细胞中高度且主要表达。通过明胶酶谱法，在分离的破骨细胞的条件培养基中检测到明胶酶活性，表明MMP-9由分离的破骨细胞分泌。通过原位杂交也在新生兔掌骨的体内破骨细胞中观察到MMP-9的表达。这些事实表明MMP-9是破骨细胞产生的主要蛋白酶之一，可能在破骨细胞性骨吸收中起作用。
