Cell Responsive Hydrogels to Improve Functional Recovery after Spinal Cord Injury

细胞响应水凝胶可改善脊髓损伤后的功能恢复

• 批准号: 8909603
• 负责人: Christopher Matthew Madl
• 金额: $ 3.42万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8909603
• 关键词: Alkynes; Animals; Apoptotic; Axon; Azides; Biochemical; Biological Assay; Cell Culture Techniques; Cell Survival; Cell Transplants; Cells; Chemistry; Cicatrix; Collaborations; Computer Simulation; Contusions; Cues; Development; Diffusion; Elastin; Engraftment; Ensure; Environment; Enzyme Kinetics; Extracellular Matrix; Feedback; Gel; Gelatinase A; Goals; Growth; Growth Cones; Histology; Homeostasis; Hydrogels; Immunohistochemistry; In Situ; In Situ Nick-End Labeling; Inflammation; Inflammatory; Inflammatory Response; Injury; Kinetics; Label; Lead; Life; Mechanics; Modeling; Motivation; Mus; NAPVSIPQ peptide; Natural regeneration; Nerve; Nerve Regeneration; Neurites; Neuronal Differentiation; Neurons; Neuroprotective Agents; Outcome; Oxidative Stress; Patients; Peptide Hydrolases; Peptides; Plants; Play; Process; Production; Proteins; Quality of life; Rattus; Reaction; Reactive Oxygen Species; Recovery; Recovery of Function; Rodent; Rodent Model; Role; Serine Protease; Signal Transduction; Site; Spinal cord injury; Staining method; Stains; Stem cell transplant; Support System; Therapeutic; Tissues; Transplantation; Urokinase; animal imaging; axon growth; base; behavior test; data modeling; design; differential expression; improved; nerve stem cell; neurodevelopment; neuron development; neurotrophic factor; oxidative damage; protein expression; public health relevance; response; stem cell differentiation; stem cell fate; stem cell therapy; stem cells; therapeutic target; time use; tissue regeneration

 DESCRIPTION (provided by applicant): Throughout tissue development and homeostasis, cells dynamically interact with the extracellular matrix (ECM). However, most materials developed to regulate stem cell fate and facilitate tissue regeneration are primarily cell-instructive, providing mechanical and biochemical signals to cells, and are not cell-responsive, that is they do not respond to the changes elicited in the delivered cells. The majority of materials that are cell-responsive simply degrade in response to cell-secreted proteases. A material that reacts to specific phenotypic changes elicited upon stem cell differentiation, such as those that occur during development, remains to be developed. A potential therapeutic target for such a material is spinal cord injury (SCI). SCI often results in severely debilitating conditins for patients, with limited clinically available treatment options. Nerve regeneration is limited by the body's natural inflammatory response that rapidly replaces injured spinal cord tissue with scar tissue. Furthermore, this inflammatory process results in significant oxidative damage to the surviving neurons, which further hampers regeneration. The goal of this project is to remediate the damage caused by this inflammation by delivering neural stem cells (NSCs) at the injury site within a material that is both cell-instructive, facilitating engraftment and differentiation of the delivered cells, and cell-responsive, releasing a neuroprotective peptide in response to neuronal differentiation. In Specific Aim 1, I will synthesize a material that dynamically responds to NSC differentiation by releasing a neuroprotective peptide. The peptide will be conjugated to an elastin-like protein (ELP) via a proteolytically cleavable linker using azide-alkyne "click" chemistry. Urokinase plasminogen activator (uPA) is a serine protease known to play a role in neuronal development, as it is secreted from the growth cones of axons. I hypothesize that neuronal differentiation of NSCs cultured in ELP hydrogels will result in increased uPA activity, which in turn will selectively release the neuroprotective peptide upon neuronal differentiation. In Specific Aim 2, I will develop a computational model to refine the cel-responsive material design. A reaction-diffusion model with Michaelis-Menten kinetics will be used to simulate the release of the neuroprotective peptide, and the relevant parameters will be experimentally determined. The model will be validated by culturing and differentiating NSCs in the cell-responsive ELP hydrogels and subjecting the cells to oxidative stress. In Specific Aim 3, I will deliver NSCs in the cell-responsive hydrogels to injury sites in rodent SCI contusion models and evaluate functional recovery. Material retention will be assessed with live-animal imaging, and NSC survival, engraftment, and differentiation will be assessed by histology. Recovery will be evaluated by tracing the regenerating nerves and through behavioral testing. I hypothesize that the cell-responsive material will improve the viability of the transplanted NSCs, resulting in improved functional recovery in animals treated with the cell-responsive materials.

 描述（由申请人提供）：在整个组织发育和稳态过程中，细胞与细胞外基质（ECM）动态相互作用。然而，大多数为调节干细胞命运和促进组织再生而开发的材料主要是细胞指导性的，向细胞提供机械和生化信号，并且不是细胞响应性的，也就是说，它们不对传递的细胞中引起的变化做出反应。大多数细胞响应材料只是响应细胞分泌的蛋白酶而降解。对干细胞分化引起的特定表型变化（例如发育过程中发生的变化）做出反应的材料仍有待开发。这种材料的潜在治疗目标是脊髓损伤（SCI）。脊髓损伤常常导致患者严重衰弱，临床上可用的治疗选择有限。神经再生受到限制 身体的自然炎症反应会迅速用疤痕组织取代受伤的脊髓组织。此外，这种炎症过程会对幸存的神经元造成严重的氧化损伤，从而进一步阻碍再生。该项目的目标是通过在损伤部位递送神经干细胞（NSC）来修复炎症造成的损伤，该材料既具有细胞指导性，促进所递送细胞的植入和分化，又具有细胞响应性，在损伤部位释放神经保护肽。 对神经元分化的反应。在具体目标 1 中，我将合成一种通过释放神经保护肽来动态响应 NSC 分化的材料。该肽将通过叠氮化物-炔“点击”化学通过蛋白水解可裂解接头与弹性蛋白样蛋白 (ELP) 缀合。尿激酶纤溶酶原激活剂 (uPA) 是一种丝氨酸蛋白酶，已知在神经元发育中发挥作用，因为它是从轴突的生长锥分泌的。我假设 ELP 水凝胶中培养的 NSC 的神经元分化将导致 uPA 活性增加，进而在神经元分化时选择性地释放神经保护肽。在具体目标 2 中，我将开发一个计算模型来完善细胞响应材料设计。将采用米氏动力学反应扩散模型来模拟神经保护肽的释放，并通过实验确定相关参数。该模型将通过在细胞响应性 ELP 水凝胶中培养和分化 NSC 并使细胞承受氧化应激来验证。在具体目标 3 中，我将把细胞响应水凝胶中的 NSC 递送至啮齿动物 SCI 挫伤模型的损伤部位，并评估功能恢复情况。将通过活体动物成像评估材料保留，并通过组织学评估 NSC 存活、植入和分化。将通过追踪再生神经和行为测试来评估康复情况。我假设细胞响应材料将提高移植的 NSC 的活力，从而改善接受细胞响应材料治疗的动物的功能恢复。