Agarwal Gopal, Moes Kennedy, Schmidt Christine E
J. Crayton Pruitt Family Department of Biomedical Engineering, Herbert Wertheim College of Engineering, University of Florida, Gainesville, FL, 32611, USA.
Mater Today Bio. 2025 Jan 11;31:101483. doi: 10.1016/j.mtbio.2025.101483. eCollection 2025 Apr.
Injuries to the central nervous system (CNS) often lead to persistent inflammation and limited regeneration. This study developed a clinically relevant injectable hydrogel derived from decellularized human peripheral nerves, with mechanical properties biomimicking native CNS tissue. Using a modified Hudson method, human sciatic nerves were decellularized, effectively removing immunogenic cellular debris while retaining the extracellular matrix. Two delipidation solvents, dichloromethane: ethanol (2:1 v/v) and n-hexane: isopropanol (3:1 v/v), were evaluated, with the former achieving optimal lipid removal and better digestion. The resulting solution was crosslinked with genipin, forming an injectable hydrogel (iHPN) that gelled within 12 min at 37 °C and exhibited mechanical stiffness of approximately 400 Pa. Human astrocytes, human microglial cell clone 3 (HMC3), and mouse RAW 264.7 macrophages were cultured individually within iHPN, with lipopolysaccharide (LPS) added to mimic CNS inflammation following injury. Compared to LPS-activated cells on tissue culture plates (TCP), astrocytes within iHPN maintained a quiescent state, as evidenced by reduced GFAP expression and IL-1β secretion. RAW 264.7 and HMC3 cells in iHPN displayed an anti-inflammatory phenotype, as shown by increased CD206 and decreased CD86/CD68 expression, along with higher IL-4 and lower TNF-α/IL-1β secretion. Human SH-SY5Y neuroblastoma cells exhibited higher viability and improved neuronal differentiation in iHPN compared to TCP. Human brain neurons had higher neuronal differentiation within iHPN compared to TCP or collagen hydrogels. Overall, iHPN is a novel injectable hydrogel that has potential for minimally invasive CNS applications, such as a carrier for cell or drug delivery and/or a biomaterial to support axonal growth.
中枢神经系统（CNS）损伤常导致持续性炎症和再生受限。本研究开发了一种源自脱细胞人外周神经的具有临床相关性的可注射水凝胶，其机械性能可模拟天然CNS组织。采用改良的哈德逊方法对人坐骨神经进行脱细胞处理，有效去除免疫原性细胞碎片，同时保留细胞外基质。评估了两种脱脂溶剂，二氯甲烷：乙醇（2:1 v/v）和正己烷：异丙醇（3:1 v/v），前者实现了最佳的脂质去除效果且消化效果更好。将所得溶液与京尼平交联，形成一种可注射水凝胶（iHPN），该水凝胶在37℃下12分钟内凝胶化，机械刚度约为400 Pa。将人星形胶质细胞、人小胶质细胞克隆3（HMC3）和小鼠RAW 264.7巨噬细胞分别培养在iHPN中，并添加脂多糖（LPS）以模拟损伤后CNS炎症。与组织培养板（TCP）上LPS激活的细胞相比，iHPN中的星形胶质细胞维持静止状态，这通过GFAP表达降低和IL-1β分泌减少得以证明。iHPN中的RAW 264.7和HMC3细胞表现出抗炎表型，表现为CD206表达增加、CD86/CD68表达降低，以及IL-4分泌增加和TNF-α/IL-1β分泌减少。与TCP相比，人SH-SY5Y神经母细胞瘤细胞在iHPN中具有更高的活力和更好的神经元分化。与人脑神经元在TCP或胶原蛋白水凝胶中相比，其在iHPN中具有更高的神经元分化。总体而言，iHPN是一种新型可注射水凝胶，具有用于微创CNS应用的潜力，例如作为细胞或药物递送的载体和/或支持轴突生长的生物材料。
