Institute of Biothermal Science and Technology, School of Health Science and Engineering, University of Shanghai for Science and Technology, Shanghai, China.
National Institutes for Food and Drug Control, Beijing, China.
J Biomed Mater Res B Appl Biomater. 2023 Jan;111(1):62-72. doi: 10.1002/jbm.b.35132. Epub 2022 Jul 13.
Regenerative bio-scaffolds, widely used for clinical tissue reconstruction and tissue repairs, are functionally diversified and structurally complex decellularized tissue materials (e.g., extracellular matrix, ECM). ECM is naturally cross-linked and can be further selectively cross-linked upon processing. Identification, quantification and bioinformatics functional comparison of all ECM proteins are challenging for regenerative bio-scaffolds. In this study, we have applied proteomic profiling with a two-step sequential trypsinization method, and identified and quantified 300-400 constituent proteins in three commercially available regenerative bio-scaffolds (BioDesign Surgisis, ReGen tissue matrix, and ThormalGEN mesh). These proteins were classified into four categories and 14 subcategories based on their mainly biological function. The main components of regenerative bio-scaffolds were highly abundant ECM structural proteins, and the minor parts of bio-scaffolds were lowly abundant, less cross-linked, functionally more diversified proteins, especially extracellular fluid proteins that were easily solubilized by trypsin. The comparative analysis has revealed large differences in the number, type, abundance and function of identified proteins, as well as the extent of decellularization and cross-linking among regenerative bio-scaffolds. So, the proteomic profiling with a two-step sequential trypsinization method could not only provide the molecular basis to better understand the degradation process of regenerative bio-scaffolds in vivo and different clinical outcomes among various regenerative bio-scaffolds, facilitate the exploration of the response mechanisms in the host's early clinical stages of ECM-induced tissue regeneration that is still poorly understood, but also can be used for optimization of the decellularization and cross-linking process, product characterization and rational design of new ECM products.
再生生物支架广泛用于临床组织重建和组织修复，其功能多样化，结构复杂，是脱细胞组织材料（如细胞外基质，ECM）。ECM 天然交联，并可在处理后进一步进行选择性交联。对于再生生物支架而言，识别、定量和生物信息学功能比较所有 ECM 蛋白都具有挑战性。在这项研究中，我们应用两步连续胰蛋白酶消化法进行蛋白质组学分析，鉴定和定量了三种市售再生生物支架（BioDesign Surgisis、Regen Tissue Matrix 和 ThormalGEN 网片）中的 300-400 种组成蛋白。这些蛋白质根据其主要生物学功能分为四类和 14 个子类。再生生物支架的主要成分是高度丰富的 ECM 结构蛋白，生物支架的次要部分是含量较低、交联较少、功能更加多样化的蛋白质，特别是容易被胰蛋白酶溶解的细胞外液蛋白质。比较分析揭示了不同再生生物支架中鉴定蛋白的数量、类型、丰度和功能以及脱细胞和交联程度存在很大差异。因此，两步连续胰蛋白酶消化法的蛋白质组学分析不仅可以为更好地理解体内再生生物支架的降解过程以及不同再生生物支架之间的不同临床结果提供分子基础，还可以促进探索宿主在 ECM 诱导组织再生的早期临床阶段反应机制（目前仍知之甚少），并可用于优化脱细胞和交联过程、产品特性分析和新 ECM 产品的合理设计。
