Ren Zongli, Chen Juanjuan, Khalil Raouf A
Vascular Surgery Research Laboratory, Division of Vascular and Endovascular Surgery, Brigham and Women's Hospital and Harvard Medical School, 75 Francis Street, Boston, MA, 02115, USA.
Methods Mol Biol. 2017;1626:79-102. doi: 10.1007/978-1-4939-7111-4_8.
Matrix metalloproteinases (MMPs) are proteolytic enzymes that degrade various components of the extracellular matrix (ECM) and play a role in tissue remodeling. Changes in MMPs have been observed in cancer, connective tissue disorders, and vascular disease, and both endogenous tissue inhibitors of MMPs (TIMPs) and synthetic MMP inhibitors (MMPIs) have been evaluated as modulators of MMP activity in various biological systems. Zymography is a simple technique that is commonly used to assess MMP activity and the efficacy of MMPIs. Also, reverse zymography is a modified technique to study the activity of endogenous TIMPs. However, problems are often encountered during the zymography procedure, which could interfere with accurate assessment of MMP activity in control specimens, and thus make it difficult to determine the pathological changes in MMPs and their responsiveness to MMPIs. Simplified protocols for preparation of experimental solutions, tissue preparation, regular and reverse zymography procedures, and zymogram analysis are presented. Additional helpful tips to troubleshoot problems in the zymography technique and to enhance the quality of the zymograms should make it more feasible to determine the changes in MMPs and assess the efficacy of MMPIs in modulating MMP activity in various biological systems and pathological conditions.
基质金属蛋白酶（MMPs）是一类蛋白水解酶，可降解细胞外基质（ECM）的各种成分，并在组织重塑中发挥作用。在癌症、结缔组织疾病和血管疾病中均观察到MMPs的变化，并且基质金属蛋白酶的内源性组织抑制剂（TIMPs）和合成基质金属蛋白酶抑制剂（MMPIs）都已作为各种生物系统中MMP活性的调节剂进行了评估。酶谱法是一种常用的简单技术，用于评估MMP活性和MMPIs的疗效。此外，反向酶谱法是一种用于研究内源性TIMPs活性的改良技术。然而，在酶谱法过程中经常会遇到问题，这些问题可能会干扰对照标本中MMP活性的准确评估，从而难以确定MMPs的病理变化及其对MMPIs的反应性。本文介绍了实验溶液制备、组织制备、常规和反向酶谱法程序以及酶谱分析的简化方案。针对酶谱法技术中出现的问题进行故障排除以及提高酶谱质量的其他有用提示，应使确定MMPs的变化并评估MMPIs在各种生物系统和病理条件下调节MMP活性的疗效变得更加可行。
