Shanbhag Siddharth, Al-Sharabi Niyaz, Fritz-Wallace Katarina, Kristoffersen Einar K, Bunæs Dagmar Fosså, Romandini Mario, Mustafa Kamal, Sanz Mariano, Gruber Reinhard
Department of Clinical Dentistry, Faculty of Medicine, University of Bergen, 5009 Bergen, Norway.
Department of Immunology and Transfusion Medicine, Haukeland University Hospital, 5009 Bergen, Norway.
J Funct Biomater. 2024 Oct 9;15(10):302. doi: 10.3390/jfb15100302.
Collagen barrier membranes are frequently used in guided tissue and bone regeneration. The aim of this study was to analyze the signature of human serum proteins adsorbed onto collagen membranes using a novel protein extraction method combined with mass spectrometry. Native porcine-derived collagen membranes (Geistlich Bio-Gide, Wolhusen, Switzerland) were exposed to pooled human serum in vitro and, after thorough washing, subjected to protein extraction either in conjunction with protein enrichment or via a conventional surfactant-based method. The extracted proteins were analyzed via liquid chromatography with tandem mass spectrometry. Bioinformatic analysis of global profiling, gene ontology, and functional enrichment of the identified proteins was performed. Overall, a total of 326 adsorbed serum proteins were identified. The enrichment and conventional methods yielded similar numbers of total (315 vs. 309), exclusive (17 vs. 11), and major bone-related proteins (18 vs. 14). Most of the adsorbed proteins (n = 298) were common to both extraction groups and included several growth factors, extracellular matrix (ECM) proteins, cell adhesion molecules, and angiogenesis mediators involved in bone regeneration. Functional analyses revealed significant enrichment of ECM, exosomes, immune response, and cell growth components. Key proteins [transforming growth factor-beta 1 (TGFβ1), insulin-like growth factor binding proteins (IGFBP-5, -6, -7)] were exclusively detected with the enrichment-based method. In summary, native collagen membranes exhibited a high protein adsorption capacity in vitro. While both extraction methods were effective, the enrichment-based method showed distinct advantages in detecting specific bone-related proteins. Therefore, the use of multiple extraction methods is advisable in studies investigating protein adsorption on biomaterials.
胶原屏障膜常用于引导组织和骨再生。本研究的目的是使用一种结合质谱的新型蛋白质提取方法，分析吸附在胶原膜上的人血清蛋白特征。将天然猪源胶原膜（Geistlich Bio-Gide，瑞士沃尔胡森）在体外暴露于混合人血清中，彻底洗涤后，通过蛋白质富集或基于传统表面活性剂的方法进行蛋白质提取。通过液相色谱-串联质谱分析提取的蛋白质。对鉴定出的蛋白质进行了全局分析、基因本体论和功能富集的生物信息学分析。总体而言，共鉴定出326种吸附的血清蛋白。富集法和传统方法产生的总蛋白数量相似（315对309）、独特蛋白数量相似（17对11）以及主要骨相关蛋白数量相似（18对14）。大多数吸附蛋白（n = 298）在两个提取组中都有，包括几种生长因子、细胞外基质（ECM）蛋白、细胞粘附分子和参与骨再生的血管生成介质。功能分析显示ECM、外泌体、免疫反应和细胞生长成分显著富集。关键蛋白[转化生长因子-β1（TGFβ1）、胰岛素样生长因子结合蛋白（IGFBP-5、-6、-7）]仅通过基于富集的方法检测到。总之，天然胶原膜在体外表现出高蛋白吸附能力。虽然两种提取方法都有效，但基于富集的方法在检测特定骨相关蛋白方面具有明显优势。因此，在研究生物材料上的蛋白质吸附时，建议使用多种提取方法。
