Zymography as a Research Tool in the Study of Matrix Metalloproteinase Inhibitors

Abstract

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.

Cirsiliol Suppressed Epithelial to Mesenchymal Transition in B16F10 Malignant Melanoma Cells through Alteration of the PI3K/Akt/NF-κB Signaling Pathway

“…The activity of the gelatinases as detected by zymography are well established protocols for observing the alterations of MMP-2/MMP-9, if any [52,53]. Gelatin zymography was performed to evaluate the activities of matrix metalloproteinases (MMP-2 and MMP-9) in FN-induced cells both in the presence and absence of cirsiliol according to a protocol described previously [54].…”

Inhibition of Matrix Metalloproteinase 9 Activity Promotes Synaptogenesis in the Hippocampus

“…Surprising finding of pro-MMP-9 and TIMP-1 proteins marked increase at 16 h post-Cch can be explained by the fact that TIMP-1–bound MMP-9 may indeed represent a large portion of total MMP-9 available at that time. Moreover, it has been noted that gel zymography might not be efficient enough at detecting pro-MMP-9 form, as only a fraction of pro-MMP-9 autoactivates after SDS gel electrophoresis ( Woessner 1995 ; Ren et al. 2017 ).…”

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“…Sigium cumini Myrtaceae 14.5 ± 0.28 ++ The phenolics content in leaf extracts of these plants was determined by using the folin-ciocalteau method and expressed as mg/ml gae. + sign indicates the presence of low inhibitor activity while, ++ sign indicates the presence of strong inhibitor activity andsign indicates the absence of inhibitor activity in the extract In-gel Detection of Metalloproteinase Activity from Human Saliva: Gelatin zymography is the most sensitive technique used for the detection of a wide spectrum of metalloproteinase activity from the biological sample at a very minute level 30 .…”