Ehrlich M G, Mankin H J, Jones H, Wright R, Crispen C, Vigliani G
J Clin Invest. 1977 Feb;59(2):226-33. doi: 10.1172/JCI108632.
In advanced osteoarthritis, all of the cartilaginous components are lost from the joint surface. Although mechanisms exist for proteoglycan degradation, there is not known to be any system for removal of the collagen. This study suggests that the loss of the collagen components may be a function of articular cartilage collagenase. The enzyme in normal human cartilage is bound to an inhibitor and appears to be present in very small amounts. Attempts to demonstrate collagenase activity in ground human articular cartilage or in its lysosomal fraction were unsuccessful. 7-Day cartilage tissue cultures also failed to demonstrate the presence of the enzyme; but the same culture fluid, incubated with trypsin, showed significant degradation of collagen, suggesting that trypsin destroyed the inhibitor. 7-Day culture fluids were then chromatographed on a heparin-charged Sepharose 4B affinity column that had been activated with cyanogen bromide. This removed the inhibitor, and the chromatographed fluid from osteoarthritic cartilage released 42% of the incorporated counts of the collagen substrate, whereas normal cartilage released 10.1% and a trypsin control, 6.4%. Electrophoresis of the degradation products of the enzyme-collagen complex incubated at 37 degrees C revealed breakdown was complete to small dialyzable fragments, while at 25 degrees C larger fragments were split off.
在晚期骨关节炎中，关节表面的所有软骨成分均已丧失。尽管存在蛋白聚糖降解机制，但目前尚不清楚是否存在清除胶原蛋白的系统。本研究表明，胶原蛋白成分的丧失可能是关节软骨胶原酶作用的结果。正常人体软骨中的这种酶与一种抑制剂结合，且含量似乎极少。在人关节软骨粉或其溶酶体组分中检测胶原酶活性的尝试均未成功。7天的软骨组织培养也未能证明该酶的存在；但将相同的培养液与胰蛋白酶一起孵育时，显示出胶原蛋白有明显降解，这表明胰蛋白酶破坏了抑制剂。然后将7天的培养液在经溴化氰活化的肝素化琼脂糖4B亲和柱上进行层析。这去除了抑制剂，骨关节炎软骨的层析液释放了胶原蛋白底物掺入量的42%，而正常软骨释放了10.1%，胰蛋白酶对照释放了6.4%。对在37℃孵育的酶 - 胶原蛋白复合物降解产物进行电泳显示，降解完全为可透析的小片段，而在25℃时则会裂解出较大的片段。
