FRET Sensor-Modified Synthetic Hydrogels for Real-Time Monitoring of Cell-Derived Matrix Metalloproteinase Activity using Fluorescence Lifetime Imaging
FRET Sensor-Modified Synthetic Hydrogels for Real-Time Monitoring of Cell-Derived Matrix Metalloproteinase Activity using Fluorescence Lifetime Imaging
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DOI：
10.1002/adfm.202309711
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发表时间：
2024-01-30
影响因子：
19
通讯作者：
Gentleman，Eileen
中科院分区：
文献类型：
--
作者：
Yan，Ziqian;Kavanagh，Thomas;Gentleman，Eileen
基质重塑在一系列生理和病理过程中起核心作用，主要由基质金属蛋白酶(MMPs)活性驱动，基质金属蛋白酶可降解细胞外基质(ECM)蛋白。MMPs如何调节细胞和组织动力学尚不清楚，因为缺乏体内方法，许多体外策略无法提供在类似组织的3D微环境中原位酶活性的高分辨率、定量测量。在这里，基质金属蛋白酶活性的Förster共振能量转移(FRET)传感器被结合到完全合成的水凝胶中，这种凝胶模仿了天然ECM的许多特性。然后使用荧光寿命成像来提供实时的、与荧光团浓度无关的基质金属蛋白酶活性的定量，建立了一个高精度、易于适应的原位研究基质金属蛋白酶动力学的平台。然后，将包裹在水凝胶中的MCF7人乳腺癌细胞用于检测局部、亚微米水平和块状水凝胶中的基质金属蛋白酶活性。这个多用途的平台可用于一系列生物学研究，通过提供高分辨率、定量和原位读数来探索癌症转移、发展和组织修复的动力学问题。
Matrix remodeling plays central roles in a range of physiological and pathological processes and is driven predominantly by the activity of matrix metalloproteinases (MMPs), which degrade extracellular matrix (ECM) proteins. How MMPs regulate cell and tissue dynamics is not well understood as in vivo approaches are lacking and many in vitro strategies cannot provide high‐resolution, quantitative measures of enzyme activity in situ within tissue‐like 3D microenvironments. Here, a Förster resonance energy transfer (FRET) sensor of MMP activity is incorporated into fully synthetic hydrogels that mimic many properties of the native ECM. Fluorescence lifetime imaging is then used to provide a real‐time, fluorophore concentration‐independent quantification of MMP activity, establishing a highly accurate, readily adaptable platform for studying MMP dynamics in situ. MCF7 human breast cancer cells encapsulated within hydrogels are then used to detect MMP activity both locally, at the sub‐micron level, and within the bulk hydrogel. This versatile platform may find use in a range of biological studies to explore questions in the dynamics of cancer metastasis, development, and tissue repair by providing high‐resolution, quantitative, and in situ readouts of local MMP activity within native tissue‐like environments.