Quantitive Analysis of Matrix Metalloproteinases Activity in Synovial Fluids of the Temporomandiblar Joint

近年来对颞下颌关节（TMJ）紊乱的生化研究提示，催化细胞外基质降解的基质金属蛋白酶（matrix metalloproteinases, MMPs）参与了颞下颌关节（TMJ）的发生，但这些研究并未完全证实其与颞下颌关节（TMJ）发生的关系。在这项研究中，我们开发了一种简单的检测系统，使用选择性荧光肽作为特定底物来检测滑液（SF）中的MMPs。我们利用mmp特异性荧光底物肽测定了SF中MMPs的活性，并研究了MMPs活性的增加与各种临床表现之间的关系。此外，为了确定对酶活性增加有贡献的MMP种类，用western bolt分析了SF中的MMPs。Westernblot分析显示，观察到的MMPs活性的增加是由MMP-2和MMP-3的激活引起的。结果，在有TMJ疼痛、关节积液和滑膜炎等炎症表现的SF患者中，MMPs活性显著高值，表明酶活性的增加是评估TMJ疾病（TMD）严重程度的有用标志。SF中活性MMP-3的存在与MMP活性的增加显著相关。这些发现提示MMP-3的激活是TMD患者早期退行性改变的标志。因此，该荧光分析系统可用于许多样品中MMPs活性的检测，如筛选。

In recent years biochemical studies on the disorder of temporomandibular joint (TMJ) have suggested that matrix metalloproteinases (MMPs) which catalyze degradation of extracellular matrix is involved in the occurrence of the TMJ, but these studies have not fully proved the relation of MMPs to the occurrence. In this study, we developed an easy assay system using the selective fluorogenic peptides as a specific substrate to detect MMPs in synovial fluid (SF). We measured MMPs activity in SF by using MMP-specific fluorogenic substrate peptides and examined about relation between increase in MMPs activity and various clinical findings. Furthermore, in order to identify MMP species contributed to the increase in enzyme activity, MMPs in SF were analyzed by western bolt. Westernblot analyses showed that the observed increase in MMPs activity was derived from activation of both MMP-2 and MMP-3. As the result, MMPs activity was significantly high value in SF from patients with inflammatory findings such as TMJ pain, joint effusion and synovitis suggesting that the increase in the enzyme activity is a useful marker for assessing the severity of TMJ disorders (TMD). The existence of active MMP-3 in SF significantly correlated with increase in activity of the MMP.These findings suggested that the activation of MMP-3 is a hallmark indicating early degenerative changes in the TMD patient. Thus, this fluorogenic assay system was useful for the detection of MMPs activity in many samples such as screening.