Okada Y, Naka K, Kawamura K, Matsumoto T, Nakanishi I, Fujimoto N, Sato H, Seiki M
Department of Pathology, School of Medicine, Kanazawa University, Japan.
Lab Invest. 1995 Mar;72(3):311-22.
Matrix metalloproteinase 9 (MMP-9, 92-kD gelatinase/type IV collagenase = gelatinase B) is a member of the MMP gene family and implicated in tissue destruction in the various pathophysiologic conditions. Our previous study showed that MMP-9 purified from human fibrosarcoma cells can cleave the cross-link-containing NH(2)-terminal telopeptides of the alpha 2 chain of type I collagen and collagen types III, IV, and V as well as gelatins.
To investigate the role of MMP-9 in bone resorption we have examined its localization in the human bone tissues by immunohistochemistry and in situ hybridization. The enzymic properties were also biochemically studied.
Immunohistochemistry using monoclonal antibodies against MMP-1 (interstitial collagenase), MMP-2 (72-kD gelatinase/type IV collagenase = gelatinase A), MMP-3 (stromelysin-1), MMP-9, and tissue inhibitor of metalloproteinases-1 demonstrated that MMP-9 is localized exclusively in osteoclasts of the bone tissues from normal subjects and patients with rheumatoid arthritis or metastatic carcinoma whereas some osteoclasts are also labeled by anti-(MMP-1) antibody. Northern blot and in situ hybridizations of rheumatoid bone tissues using a RNA probe for MMP-9 exhibited strong signals for the mRNA within osteoclasts. MMP-9 depolymerized acid-insoluble polymers of type I collagen and digested collagen fibrils in the demineralized bone. The gelatinolytic activity of the proteinase was optimal at pH 7.5, but 50 to 80% of the full activity was retained at pH 5.5 to 6.0. It was also 90% active in the presence of 100 mM Ca2+. Degradation of acid-soluble and -insoluble type I collagens by MMP-9 was enhanced at higher concentrations of Ca2+. The zymogen of MMP-9 was activated up to approximately 85% of full activity by incubation at pH 2.3.
These results demonstrate that MMP-9 is produced by osteoclasts in the human bone tissues and suggest that it can degrade bone collagens in concert with MMP-1 and cysteine proteinases in the subosteoclastic microenvironment. This proteinase may play a role in the normal bone remodeling and pathologic bone resorption in the human diseases.
基质金属蛋白酶9（MMP - 9，92-kD明胶酶/IV型胶原酶 = 明胶酶B）是MMP基因家族的成员，在多种病理生理条件下参与组织破坏。我们之前的研究表明，从人纤维肉瘤细胞中纯化的MMP - 9能够切割I型胶原α2链以及III型、IV型和V型胶原和明胶中含交联的NH₂-末端肽段。
为了研究MMP - 9在骨吸收中的作用，我们通过免疫组织化学和原位杂交检测了其在人骨组织中的定位。还对其酶学特性进行了生化研究。
使用针对MMP - 1（间质胶原酶）、MMP - 2（72-kD明胶酶/IV型胶原酶 = 明胶酶A）、MMP - 3（基质溶解素-1）、MMP - 9和金属蛋白酶组织抑制剂-1的单克隆抗体进行免疫组织化学分析表明，MMP - 9仅定位于正常受试者以及类风湿关节炎或转移性癌患者骨组织中的破骨细胞，而一些破骨细胞也被抗（MMP - 1）抗体标记。使用MMP - 9的RNA探针进行类风湿骨组织的Northern印迹和原位杂交显示破骨细胞内mRNA有强信号。MMP - 9使I型胶原的酸不溶性聚合物解聚，并消化脱矿骨中的胶原纤维。该蛋白酶的明胶水解活性在pH 7.5时最佳，但在pH 5.5至6.0时保留了50%至80%的全部活性。在100 mM Ca²⁺存在下其活性也为90%。在较高浓度的Ca²⁺下，MMP - 9对酸溶性和酸不溶性I型胶原的降解作用增强。通过在pH 2.3孵育，MMP - 9的酶原被激活至约85%的全部活性。
这些结果表明MMP - 9由人骨组织中的破骨细胞产生，并提示它可与破骨细胞微环境中的MMP - 1和半胱氨酸蛋白酶协同降解骨胶原。这种蛋白酶可能在人类疾病的正常骨重塑和病理性骨吸收中起作用。
