Wang Ruyue, Chen Yunlu, Wu Chenqi, Li Shujing, Liu Zhenjie, Chen Feng
College of Materials Science and Engineering, Zhejiang University of Technology, Hangzhou 310014, China.
Department of Vascular Surgery, the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310009, China.
Zhejiang Da Xue Xue Bao Yi Xue Ban. 2025 Jul 16;54(4):434-445. doi: 10.3724/zdxbyxb-2024-0698.
To fabricate an injectable composite microsphere hydrogel reinforced with silk fibroin/hyaluronic acid microspheres, achieving synergistic enhance-ment of mechanical robustness and biofunctionality.
Methacrylated hyaluronic acid (HAMA) and thiolated silk fibroin (TSF) were synthesized. Monodisperse microspheres generated via microfluidics were UV-cured (420 nm) through thiol-ene click reaction. These microspheres were embedded in a TSF/HAMA matrix to form photo-cured composites. The grafting rate of TSF and HAMA was characterized by H1-NMR; particle size distribution of microsphere hydrogels in soybean oil was observed by optical microscopy; gel point of composite microsphere hydrogels was determined by advanced extensional rheometer; microscopic morphology of microsphere hydrogels was observed by scanning electron microscopy; elemental distribution of microsphere hydrogels was detected by X-ray energy dispersive spectroscopy; tunability of composite microsphere hydrogels was observed by inverted confocal microscopy; mechanical properties of composite microsphere hydrogels were tested by compression testing; swelling ratio, degradation rate and water retention rate of composite microsphere hydrogels were measured by gravimetric method. Cytotoxicity of the composite microsphere hydrogels was determined by Calcein-AM/propidium iodide dual staining and CCK-8 assay; cell migration capability was observed by scratch assay.
The grafting rates of HAMA and TSF was 48.03% and 17.99%, respectively. Microsphere hydrogels with particle sizes of (43.3±1.2), (78.1±3.0), and (130.8±1.9) μm were prepared. The gel time of the composite microsphere hydrogels was 48-115s. The laser confocal imaging confirmed dynamic regulation characteristics of the composite microsphere hydrogels. The compressive strength of the composite microsphere hydrogels reached 22.7 kPa and maintained structural integrity at 40% strain after 20 compression cycles. The composite microsphere hydrogels exhibited differential deswelling behaviors in simulated physiological environments, and reducing microsphere particle size could significantly enhance its stability under moist conditions. The degradation rate of the composite microsphere hydrogels was (49.1±0.9)% after 200 h, and water retention rate was maintained at 40%-60% after 96 h. Biocompatibility assays confirmed >95% cell viability and unimpaired cell migration abilities.
The TSF/HAMA composite microsphere hydrogel developed in this study has characteristics of rapid fabrication, adjustable mechanical properties, enhanced environmental stability and excellent biocom-patibility, thus providing a new material solution for tissue repair and regenerative medicine.
制备一种用丝素蛋白/透明质酸微球增强的可注射复合微球水凝胶，实现机械强度和生物功能的协同增强。
合成甲基丙烯酸化透明质酸（HAMA）和巯基化丝素蛋白（TSF）。通过微流控技术产生的单分散微球经硫醇-烯点击反应进行紫外光固化（420nm）。将这些微球包埋于TSF/HAMA基质中形成光固化复合材料。通过氢核磁共振（H1-NMR）表征TSF和HAMA的接枝率；用光学显微镜观察微球水凝胶在大豆油中的粒径分布；用高级拉伸流变仪测定复合微球水凝胶的凝胶点；用扫描电子显微镜观察微球水凝胶的微观形态；用X射线能量色散光谱检测微球水凝胶的元素分布；用倒置共聚焦显微镜观察复合微球水凝胶的可调性；用压缩试验测试复合微球水凝胶的力学性能；用重量法测量复合微球水凝胶的溶胀率、降解率和保水率。通过钙黄绿素-AM/碘化丙啶双重染色和CCK-8法测定复合微球水凝胶的细胞毒性；用划痕试验观察细胞迁移能力。
HAMA和TSF的接枝率分别为48.03%和17.99%。制备了粒径为（43.3±1.2）、（78.1±3.0）和（130.8±1.9）μm的微球水凝胶。复合微球水凝胶的凝胶时间为48 - 115秒。激光共聚焦成像证实了复合微球水凝胶的动态调控特性。复合微球水凝胶的抗压强度达到22.7kPa，在20次压缩循环后，在40%应变下仍保持结构完整性。复合微球水凝胶在模拟生理环境中表现出不同的去溶胀行为，减小微球粒径可显著提高其在潮湿条件下的稳定性。复合微球水凝胶在200小时后的降解率为（49.1±0.9）%，96小时后保水率保持在40% - 60%。生物相容性检测证实细胞活力>95%，细胞迁移能力未受损。
本研究开发的TSF/HAMA复合微球水凝胶具有制备快速、机械性能可调、环境稳定性增强和生物相容性优异的特点，从而为组织修复和再生医学提供了一种新的材料解决方案。
