Modifying High Modulus Hydrogels for Cell Delivery: Intervertebral Disc Repair with Genipin-Crosslinked Fibrin

项目总结 背部和颈部疼痛是全球残疾的主要原因，占医疗保健费用的1350亿美元以上 花销。腰椎间盘突出症(IVD)引起的致残背痛可通过以下方法缓解 椎间盘切除术，切除突出的IVD组织的外科标准护理。虽然有效，但椎间盘切除术确实有效。 不修复因突出引起的纤维环(AF)缺陷，这可能导致加速的IVD 变性、再突出和反复疼痛。细胞接种、粘附性水凝胶是一种很有前途的策略 预防这些并发症，因为它们可以立即封闭房颤缺陷并长期输送细胞 治愈。设计这种用于IVD细胞输送的水凝胶是具有挑战性的，因为软生物材料通常使用 对于IVD损伤空间内的细胞递送风险。相反，高模数生物材料旨在 承受高强度的脊柱负荷会阻碍包裹的细胞的愈合能力。这个项目的总体目标是 研究建议是将细胞微胶囊、可降解微球的原理独一无二地结合起来 (MBS)和高模数生物材料，以设计促进IVD再生的下一代生物材料 和功能性修复。目标1将评估氧化海藻酸盐的保护能力和降解动力学 (OxAlg)MBS。目标2将表征染料木素-交联型纤维蛋白(FibGen)-OxAlg结构的作用 房颤细胞表型的大孔隙率和构建生物力学。目标3将评估生物和 FibGen-OxAlg的生物力学修复反应。我们的全球假设是OxAlg MBS将保护 来自FibGen水凝胶的AF细胞然后降解(目标1)。合成的大孔FibGen-OxAlg 构建体可促进房颤细胞增殖和细胞外基质合成，从而增强构建体 生物力学(目标2)。这种细胞传递策略将促进EX的生物和生物力学修复 体内IVD器官培养(目标3)。这项工作意义重大，因为它产生了一种容易翻译的组织 解决与房颤缺陷相关的关键临床挑战的工程策略；此方法可能是 广泛适用于其他愈合有限和机械强度较高的肌肉骨骼组织 需求，这与NIAMS的使命非常一致。这项建议具有很高的创新性，因为 存在修复和再生房颤缺损的策略，很少有发表的研究使用细胞负载的MBS作为成孔剂。 模板化的水凝胶结构，没有人在IVD修复中使用这种结构。验证这一方法的有效性 在加载的大型动物IVD器官培养系统中的生物材料策略是创新和重要的，因为 使用这种培养系统的已发表研究很少，以这种方式进行的测试将加速临床 翻译。完成拟议的目标将为候选人提供严格的多学科培训 生物材料合成、细胞微囊化、生物力学测试和IVD器官培养。辅以 建议的教学和指导经验，这一奖学金将加速一个有前途的博士的职业生涯 强烈致力于肌肉骨骼组织工程研究和教育的候选人。

PROJECT SUMMARY Back and neck pain are leading causes of global disability, which account for over $135 billion in healthcare spending. Disabling back pain caused by herniation of the intervertebral disc (IVD) can be alleviated by discectomy, the surgical standard of care that removes herniated IVD tissue. While effective, discectomy does not repair annulus fibrosus (AF) defects caused by the herniation, which can lead to accelerated IVD degeneration, reherniation and recurrent pain. Cell-seeded, adhesive hydrogels are a promising strategy to prevent these complications because they can immediately seal AF defects and deliver cells for long-term healing. Engineering such hydrogels for IVD cell delivery is challenging because soft biomaterials typically used for cell delivery risk herniating in the IVD injury space. On the contrary, high-modulus biomaterials designed to bear high-magnitude spine loads can hinder the healing capacity of encapsulated cells. The overall goal of this research proposal is to uniquely integrate principles of cellular microencapsulation, degradable microbeads (MBs) and high-modulus biomaterials to engineer next-generation biomaterials that promote IVD regeneration and functional repair. Aim 1 will assess the protective capacity and degradation kinetics of oxidized alginate (OxAlg) MBs. Aim 2 will characterize the effects of genipin-crosslinked fibrin (FibGen)-OxAlg construct macroporosity on AF cell phenotype and construct biomechanics. Aim 3 will evaluate the biological and biomechanical repair responses of FibGen-OxAlg. Our global hypotheses are that OxAlg MBs will protect AF cells from FibGen hydrogel crosslinking then degrade (Aim 1). Resultant macroporous FibGen-OxAlg constructs will promote AF cell proliferation and ECM synthesis, leading to enhanced construct biomechanics (Aim 2). This cell delivery strategy will promote biological and biomechanical repair in ex vivo IVD organ culture (Aim 3). This work is significant because it develops an easily translatable tissue engineering strategy to address the critical clinical challenges associated with AF defects; this approach may be broadly applicable to other musculoskeletal tissues that exhibit limited healing and experience high mechanical demands, which strongly aligns with the mission of NIAMS. This proposal is highly innovative because no strategies that repair and regenerate AF defects exist, few published studies use cell-laden MBs as porogens in templated hydrogel constructs, and none use such constructs in IVD repair. Validating the efficacy of this biomaterial strategy in a loaded, large animal IVD organ culture system is innovative and significant because there are few published studies using such a culture system and testing in this manner will accelerate clinical translation. Completion of the proposed aims will provide the candidate with rigorous multidisciplinary training in biomaterial synthesis, cell microencapsulation, biomechanical testing and IVD organ culture. Supplemented with the proposed teaching and mentorship experiences, this fellowship will accelerate the career of a promising PhD Candidate who is strongly committed to musculoskeletal tissue engineering research and education.