新型包裹嗅鞘细胞的bFGF-HP温控型水凝胶提高脊髓损伤治疗作用研究

Spinal cord injury is a serious nervous system disease. The incidence of spinal cord injury increases year by year and the age of onset shows a trend of getting younger. Chemical drugs, though protect the remaining neurons after spinal cord injury from secondary damage, can not promote the regeneration of axons and repair damaged neurons. Olfactory ensheathing cells (OECs) have both the characteristics of astrocytes and Schwann cells. From previous studies, OECs can support neuron growth and secrete various neurotrophic factors, reduce the inflammation reaction and the destruction of cells to construct of the damaged spinal cord. Therefore, OECs can provide a suitable microenvironment for the axon regeneration. However, OECs can not penetrate into the spinal cord parenchyma after transplantation because the current OECs formulations can not maintain the long time fully contact with spinal cord damage surface. Basic fibroblast growth factor (bFGF) has dual roles in repairing damaged neurons and axons. Due to the poor chemiphysical stability and low ability to cross the blood spinal cord barrier, bFGF can not play an effective therapeutic application in clinical practice. Our team has developed a new temperature-sensitive functional material — heparin-poloxamer (HP), which has a high affinity with bFGF. Based on our previous experiments, new bFGF-HP thermo-sensitive hydrogel containing OECs (OECs-bFGF-HP hydrogel) will be prepared. Through modern pharmacology and pharmacokinetic method, the promotion effects of OECs-bFGF-HP on neuron axon growth and function restoration will be evaluated comprehensively. The therapeutic efficacy and safety of OECs-bFGF-HP in treatment of spinal cord injury will be summarized. From this study, the combinations of multiple methods — new material and its preparation, bFGF and OECs transplantation, are explored to overcome the current bottleneck in the clinical therapy of spinal cord injury.

脊髓损伤发生率逐年增高且呈年轻化趋势。脊髓损伤后化学药物治疗仅能保护残存神经元免受二次损伤，很难促进轴突再生和修复受损神经元。嗅鞘细胞（OECs）兼有星形胶质细胞与雪旺细胞的特点。研究表明，OECs支持神经元生长并分泌多种神经营养因子，减轻受损脊髓炎症反应和细胞构筑的破坏，为轴突再生提供适宜微环境。但是OECs移植后与脊髓受损面接触不佳，不能很好地迁移至脊髓实质。胶质细胞源性神经营养因子（bFGF）具有修复损伤神经元及轴突的双重作用，已得到实验动物的验证。但是bFGF稳定性差，并难以透过血-脊髓屏障，从而应用受限。本项目基于自主合成的肝素-泊洛沙姆（HP）新材料，制备包裹OECs的bFGF-HP温敏型水凝胶，通过现代药理学和药动学方法，全面评价促进神经元轴突生长和功能恢复作用，总结有效性和安全性。本课题利用新材料和制剂技术，将治疗因子和细胞移植有机结合，探索克服脊髓损伤治疗瓶颈的新技术。

脊髓损伤（SCI）后化学药物治疗仅能保护残存神经元免受二次损伤，很难促进轴突再生和修复受损神经元。本项目基于自主合成的肝素-泊洛沙姆（HP）新材料，制备包裹干细胞的FGF-HP温敏型水凝胶，探讨其对SCI的治疗作用和效果。为了更方便转化应用，我们考虑直接考察HP包载生长因子对SCI的作用。我们的研究显示HP-aFGF材料可以减少紧密连接的破坏从而保护血脊髓屏障，通过抑制ER Stress从而减少神经元细胞的凋亡，抑制胶质瘢痕的形成并具有保护髓鞘，并且具有促进轴突生长的作用。HP-aFGF材料可缓释aFGF并保护其活性，较好的克服了aFGF半衰期短、易降解等问题，同时HP具有温敏性的特点，方便给药，为SCI的临床治疗提供了新的思路。同时我们利用细胞外基质具有存储生长因子能力的特点，将bFGF与脊髓的脱细胞基质(ASC)复合，构建了可注射型、在体胶凝的bFGF-ASC-HP水凝胶。研究表明ASC表面存在大量的bFGF受体，能高效地与药物复合，显著提高bFGF对热的稳定性； bFGF与ASC形成的复合物(bFGF-ASC)并不影响HP材料的胶凝性，损伤部位局部注射bFGF-ASC-HP水溶液后能快速胶凝，形成多孔的三维结构，体外缓慢释放药物长达7天。体内SCI大鼠模型研究表明，与bFGF溶液相比，该bFGF-ASC-HP杂化水凝胶更显著地增加神经元的存活和改善大鼠的运动功能。进一步，我们还将HP水凝胶同时包载NGF和bFGF，能显著改善神经损伤后大鼠运动功能恢复，其作用机理与HP-GFs水凝胶激活PI3K/Akt、JAK/STAT3和MAPK/ERK信号通路相关。我们的研究结果为SCI治疗提供了新的治疗策略，在ACS Appl Mater Interfaces、Biomaterials等国际期刊发表论文9篇，申请发明专利3项，参与获得国家科技进步二等奖等。