DataSheet1_Hybrid Hydrogel Composed of Hyaluronic Acid, Gelatin, and Extracellular Cartilage Matrix for Perforated TM Repair.doc

A novel series of composite hydrogels, built from the three components 1), hyaluronic acid methacryloyl (HAMA); 2), gelatin methacryloyl (GelMA), and 3), extracellular cartilage matrix (ECM), was prepared and studied regarding the possible utility in the surgical repair of damaged (perforated) tympanic membrane (TM). Noteworthy is component 3), which was harvested from the ribs of α-1,3-galactosidyltransferase-knockout (α-1,3 GalT-KO) pigs. The absence of α-1,3-galactosyl glycoprotein is hypothesized to prevent rejection due to foreign-body immunogenicity. The composite hydrogels were characterized by various aspects, using a variety of physicochemical techniques: aqueous swelling, structural degradation, behavior under compression, and morphology, e.g., in vitro biocompatibility was assessed by the CCK-8 and live–dead assays and through cytoskeleton staining/microscopy. Alcian blue staining and real-time PCR (RT-PCR) were performed to examine the chondrogenic induction potential of the hydrogels. Moreover, a rat TM defect model was used to evaluate the in vivo performance of the hydrogels in this particular application. Taken together, the results from this study are surprising and promising. Much further development work will be required to make the material ready for surgical use.

本研究制备并表征了一系列新型复合水凝胶，该水凝胶由三种组分构成：1）甲基丙烯酸透明质酸（hyaluronic acid methacryloyl, HAMA）；2）甲基丙烯酸明胶（gelatin methacryloyl, GelMA）；3）细胞外软骨基质（extracellular cartilage matrix, ECM），并探究其在受损（穿孔）鼓膜（tympanic membrane, TM）外科修复中的应用潜力。值得注意的是，第三种组分取自α-1,3-半乳糖基转移酶敲除（α-1,3-galactosidyltransferase-knockout, α-1,3 GalT-KO）猪的肋软骨。研究推测，该基质缺失α-1,3-半乳糖基糖蛋白，可规避异体免疫原性引发的排斥反应。本研究采用多种理化表征技术对该复合水凝胶进行多维度分析，涵盖吸水溶胀特性、结构降解行为、压缩力学性能及微观形貌；同时通过CCK-8法、活死细胞染色法及细胞骨架染色/显微成像技术评估其体外生物相容性。通过阿尔辛蓝染色与实时荧光定量聚合酶链式反应（real-time PCR, RT-PCR）检测该水凝胶的软骨诱导分化潜能。此外，本研究构建大鼠鼓膜缺损模型，以评估该水凝胶在该特定应用场景中的体内性能。综合而言，本研究所得结果既出人意料又极具应用前景，但该材料仍需开展大量后续研发工作，方能满足外科临床应用的要求。