The pathophysiological significance of proteases in patients with chronic inflammatory joint disease

为了阐明蛋白水解酶在类风湿关节炎和骨性关节炎中的病理生理学意义，我们研究了类风湿关节炎和骨性关节炎患者滑液和滑膜组织中特异性激活的蛋白水解酶，以及丝氨酸蛋白酶对滑膜细胞释放炎性细胞因子的影响。(1)当酶底物与SF上清液孵育时，凝血酶是最优先的水解物，而胰酶底物和组织蛋白酶B底物有一定程度的水解性。在SF细胞提取液中，组织蛋白酶B底物最优先被水解，而其他底物几乎不被水解。在滑膜组织提取液中，组织蛋白酶B底物最优先被水解，而胰酶底物有一定程度的水解。总体而言，类风湿关节炎患者的蛋白水解酶活性显著高于骨性关节炎患者。(2)凝血酶(0.1~100单位/ml)可显著促进滑膜成纤维细胞样细胞(FC)的生长。当SFC与凝血酶(100单位/ml)或MCT(10 ng/ml)共同培养时，培养上清液中IL-8和IL-6的浓度显著高于对照组，而IL-1-、IL-1-和TNF-1的浓度在所有培养上清液中均未检测到。(3)从SF和滑膜组织中纯化的类组织蛋白酶B主要由组织蛋白酶B组成，从SF组织中纯化的组织蛋白酶B在体外能够激活尿激酶原为尿激酶。

To clarify the pathophysiological significance of proteases in RA and OA, we investigated what kinds of proteases were specifically activated in both synovial fluid and synovial tissue of RA ＆ OA patients , and effects of serine proteases on release of inflammatory cytokines from synovial cells.We obtained the following fmdings. (I) When the protease substrates were incubated with SF supernatants, thrombin was the most preferentially hydrolyzed while trypsin substrate and cathepsin B substrate were hydrolyzed to some degree. In the case of SF cell extracts, cathepsin B substrate was the most preferentially hydrolyzed while the other substrates were hardly hydrolyzed. In the case of synovial tissue extracts, cathepsin B substrate was the most preferentially hydrolyzed while trypsin substrate was hydrolyzed to some degree. Protease activities were in general significantly higher in RA than in OA. (II) Thrombin (0.1 to 100 units /ml) significantly enhanced growth of the synovial fibroblast-like cells (FC). When SFC were cultured with thrombin (100 units/ml) or MCT (10 ng/ml), concentrations of IL-8 and IL-6 in the culture medium were significantly increased compared with the control, while the concentrations of IL-1□, IL-1□ and TNF-□ were not detectable in all the culture mediums. (III) Cathepsin B-like protease purified from both the SF and synovial tissues ofRA patients was mainly composed of cathepsin B, and the cathepsin B purified from SF samples ofRA patients was capable of activating prourokinase to urokinase in vitro.