Regulation of matrix metalloproteinase activity during tissue remodelling

组织重塑过程中基质金属蛋白酶活性的调节

• 批准号: RGPIN-2018-04559
• 负责人: Crawford, Bryan
• 金额: $ 2.33万
• 依托单位: University of New Brunswick
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=745975
• 关键词: Regulation; matrix; metalloproteinase; activity; during

Tissues are comprised of cells and a complex matrix of proteins cells secrete and adhere to. This extracellular matrix (ECM) is constantly remodeled, most obviously during the dramatic growth and cellular rearrangements of embryonic development, but also during wound healing and as part of the process of ageing. Cells remodel the ECM using a complex family of enzymes called matrix metalloproteinases (MMPs). Because MMPs digest all of the ECM proteins, their activity must be carefully regulated or tissues would disintegrate. Not surprisingly, defects in MMP regulation are causes and/or effects of a variety of diseases, including arthritis, heart disease and cancer. But surprisingly little is known about the regulation of MMP activity in living tissues; most of what we have learned comes from experiments done on isolated cells in petri dishes, and biochemical experiments with purified proteins.Zebrafish are an excellent model organism for biomedical research and we have identified all of the MMPs and their known regulators in the zebrafish genome. Because the zebrafish embryo is transparent, and develops externally and rapidly, we can observe the effects of manipulating MMP activity relatively easily. Furthermore, we have developed techniques that take advantage of the transparency of the fish embryo to visualize MMP activity, and to detect the biochemical events that regulate these ECM-remodeling proteases in the intact embryo. We have developed a novel transgenic line of zebrafish that expresses an engineered protein, which reports on the activation of one of these MMPs. Using these embryos we will generate the first ever map' of when and where this MMP becomes activated during development. Furthermore, using these embryos we can find out if the biochemical mechanisms that are thought to regulate this MMP are actually occurring in the living animal.Invasive cells use MMPs to degrade the boundaries between tissues, allowing them to migrate through the body. This is essential for normal embryonic development, wound healing, and the immune response, but it is also how tumour cells metastasize. We would therefore like to better understand how MMPs are regulated at the interface between normally and pathologically invasive cell types. We can transplant human tumour cells into zebrafish embryos and observe them as they proliferate and spread through the embryo. Using our transgenic line and other tools, we can test hypotheses about how invasive cells use their own MMPs and/or the MMPs present in the surrounding tissues to degrade the ECM, allowing them to invade and move through the embryo. This will not only help us understand how embryos build themselves and how tissues maintain and regenerate their structure after injury, but also how MMPs become activated under pathological conditions that cause damage to tissues, and allow tumors to spread throughout the body.

组织是由细胞和细胞分泌和粘附的蛋白质的复杂基质组成的。这种细胞外基质（ECM）不断重塑，最明显的是在胚胎发育的急剧生长和细胞重排期间，但也在伤口愈合和衰老过程中。细胞使用一种称为基质金属蛋白酶（MMPs）的复杂酶家族来重塑ECM。因为MMPs能消化所有的ECM蛋白，所以必须仔细调节它们的活性，否则组织就会瓦解。不足为奇的是，MMP调节缺陷是多种疾病的原因和/或结果，包括关节炎、心脏病和癌症。但令人惊讶的是，人们对活组织中MMP活性的调控知之甚少；我们所学到的大部分知识来自于在培养皿中对分离细胞进行的实验，以及用纯化蛋白质进行的生化实验。斑马鱼是一种优秀的生物医学研究模式生物，我们已经确定了斑马鱼基因组中所有的MMPs及其已知的调节因子。由于斑马鱼胚胎是透明的，外部发育迅速，我们可以比较容易地观察到操纵MMP活性的效果。此外，我们已经开发出技术，利用鱼胚胎的透明度来可视化MMP活性，并检测在完整胚胎中调节这些ecm重塑蛋白酶的生化事件。我们已经开发了一种新的斑马鱼转基因系，表达了一种工程蛋白，报告了这些MMPs之一的激活。利用这些胚胎，我们将生成有史以来第一张MMP在发育过程中何时何地被激活的地图。此外，利用这些胚胎，我们可以发现被认为调节这种MMP的生化机制是否真的发生在活体动物身上。侵袭性细胞利用MMPs降解组织之间的边界，使它们能够在体内迁移。这对正常的胚胎发育、伤口愈合和免疫反应至关重要，但也是肿瘤细胞转移的方式。因此，我们想更好地了解MMPs是如何在正常和病理侵袭细胞类型之间的界面进行调节的。我们可以将人类肿瘤细胞移植到斑马鱼胚胎中，观察它们在胚胎中增殖和扩散的过程。利用我们的转基因细胞系和其他工具，我们可以测试关于侵袭性细胞如何利用它们自己的MMPs和/或周围组织中存在的MMPs来降解ECM，使它们能够侵入并在胚胎中移动的假设。这不仅将帮助我们了解胚胎如何自我构建，组织如何在损伤后维持和再生其结构，而且还将帮助我们了解MMPs如何在病理条件下被激活，从而导致组织损伤，并允许肿瘤在全身扩散。