3D bioprinting of regenerative, corneal cell-laden inks to treat corneal blindness

3D 生物打印充满角膜细胞的再生墨水来治疗角膜失明

• 批准号: 10606474
• 负责人: Lucia Brunel
• 金额: $ 4.05万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10606474
• 关键词: 3-Dimensional; Address; Affect; Alkynes; Animals; Autologous; Azides; Behavior; Biocompatible Materials; Biological; Biomedical Engineering; Biophysics; Blindness; Cadaver; Cell Therapy; Cell secretion; Cells; Chemistry; Cicatrix; Clinical Markers; Collaborations; Collagen; Collagen Fibril; Collagen Type I; Complex; Computer-Aided Design; Contracts; Cornea; Corneal Diseases; Corneal Injury; Corneal Stroma; Cues; Custom; Defect; Deposition; Development; Doctor of Medicine; Doctor of Philosophy; Educational workshop; Encapsulated; Engineering; Epithelium; Exhibits; Eye; Eye diseases; Fostering; Foundations; Friends; Future; Geometry; Goals; Harvest; Human; Hydrogels; Immune response; Implant; Inflammation; Inflammatory Response; Ink; Keratoplasty; Laboratories; Light; Mentorship; Methods; Modeling; Monitor; Natural regeneration; Ophthalmology; Optics; Oryctolagus cuniculus; Paracrine Communication; Patient-Focused Outcomes; Patients; Pattern; Persons; Phenotype; Physicians; Process; Production; Productivity; Property; Proteins; Reaction; Regenerative Medicine; Regenerative capacity; Regenerative response; Research; Research Ethics; Research Proposals; Scientist; Series; Source; Structure; Surface; System; Technical Expertise; Techniques; Testing; Therapeutic; Thick; Time; Tissue Donors; Tissue Engineering; Tissue Grafts; Tissues; Training; Transplantation; Visual impairment; Work; Wound models; Writing; beneficiary; bioink; bioprinting; catalyst; clinical translation; conventional therapy; corneal epithelial wound healing; corneal regeneration; corneal scar; corneal surgery; cross reactivity; crosslink; curative treatments; cycloaddition; design; empowerment; global health; healing; immunohistochemical markers; implantation; improved; improved outcome; in vivo; innovation; interdisciplinary approach; materials science; mechanical properties; mesenchymal stromal cell; pre-clinical; prevent; regeneration potential; regenerative; regenerative cell; response; restoration; sight restoration; skills; symposium; translational applications; translational potential

Project Summary The demand for human donor tissue for treatment of corneal blindness far outpaces the supply, necessitating an innovative bioengineered approach for corneal regeneration. However, current laboratory-made constructs are insufficient due to lack of long-term transparency and underwhelming regenerative capacity. Recent studies suggest that cell therapies using transplanted corneal mesenchymal stromal cells (MSCs) help prevent corneal scar formation and restore corneal transparency. To propel this strategy toward clinical translation, both an effective cell delivery system and a precise understanding of their regenerative effects will be required. 3D bioprinting—in which cells and matrix components are precisely patterned—is a promising technique for creating customizable corneal constructs for implantation. We have previously demonstrated a versatile, cell-friendly 3D bioprinting platform for corneal MSCs. In my proposed research, I will apply 3D bioprinting for bioorthogonally- crosslinked collagen hydrogels with encapsulated human corneal MSCs to fabricate highly regenerative corneal stroma-like constructs. This system will allow for control over the corneal MSC microenvironment to optimize and leverage their regenerative potential. I specifically aim to overcome two common challenges in bioengineered corneal tissue that both result in loss of transparency: (1) tissue contraction over time due to cell- imposed forces, and (2) lack of cell and matrix organization that mimics the hierarchical structure of the native corneal stroma. I will test the hypotheses that (Aim 1) crosslinking collagen hydrogels with a covalent bioorthogonal chemistry increases stability against contraction from corneal MSCs without hindering their ability to secrete pro-regenerative factors and (Aim 2) aligning the collagen fibril microstructure through 3D bioprinting guides the organized deposition of nascent matrix to facilitate sustained transparency. These constructs will be evaluated in vivo (Aim 3) in a rabbit keratectomy model to assess restoration of corneal thickness, stromal integrity, and optical transparency; integration of the hydrogel into the host; re-epithelialization; surface inflammation and scarring; and phenotype of the transplanted corneal MSCs. Together, these results will be critical for understanding the effect of the surrounding 3D matrix on the regenerative capacity of corneal MSCs. My training will be supported by Sarah Heilshorn, Ph.D. (Materials Science & Engineering), an expert on biomaterials for regenerative medicine, and David Myung, M.D., Ph.D. (Ophthalmology), an attending physician who specializes in corneal surgery and diseases of the eye. In addition to expanding my scientific technical skills, my training plan includes development of mentorship, scientific writing, and presentation skills; training in research ethics; and enhancement of collaboration skills through a series of on-campus courses, workshops and seminars as well as off-campus conferences. Altogether, this research proposal will empower me to become an independent, productive research scientist as I leverage the regenerative capacity of corneal MSCs for the overarching goal of restoring vision on-demand to patients with corneal blindness.

项目摘要 用于治疗角膜失明的人类供体组织供不应求，这就需要 一种创新的生物工程角膜再生方法。然而，目前实验室制造的结构 由于缺乏长期透明度和平淡无奇的再生能力，这些资金不足。最新研究 提示使用移植的角膜间充质基质细胞(MSCs)的细胞疗法有助于预防角膜 形成疤痕，恢复角膜透明度。为了推动这一战略走向临床翻译，既有 需要有效的细胞递送系统和对其再生效果的精确了解。3D 生物打印--其中细胞和基质成分被精确地图案化--是一种很有前途的技术 用于植入的可定制角膜结构。我们之前已经展示了一种多功能的、对细胞友好的3D 角膜间充质干细胞生物打印平台。在我提出的研究中，我将把3D生物打印应用于生物正交化- 包裹人角膜间充质干细胞的交联型胶原水凝胶构建高再生角膜 基质样构造。该系统将允许对角膜MSC微环境进行控制以优化 并利用它们的再生潜力。我的具体目标是克服两个共同的挑战 生物工程的角膜组织都会导致透明度的丧失：(1)随着时间的推移，由于细胞- 施加的力量，以及(2)缺乏模仿原生组织层级结构的细胞和矩阵组织 角膜基质。我将测试以下假设：(目标1)将胶原水凝胶与共价键进行交联 生物正交化学增强抗角膜MSCs收缩的稳定性，而不会阻碍其能力 分泌促再生因子和(目标2)通过3D生物打印对准胶原原纤维的微结构 指导新生基质的有组织沉积，以促进持续的透明度。这些构造将是 兔角膜切削术模型的体内评估(AIM 3)以评估角膜厚度、基质的恢复 完整性和光学透明度；水凝胶与宿主的整合；重新上皮化；表面 炎症和瘢痕形成；移植的角膜MSCs的表型。总而言之，这些结果将是 对于了解周围3D基质对角膜MSCs再生能力的影响至关重要。 我的培训将由萨拉·海尔肖恩博士(材料科学与工程)提供支持，她是一名 再生医学生物材料和主治医师David Myung，医学博士，博士(眼科) 世界卫生组织专门从事角膜手术和眼科疾病。除了扩展我的科学技术技能外， 我的培训计划包括发展导师、科学写作和演示技能；培训 研究道德；并通过一系列校园课程、研讨会和 研讨会和校外会议。总之，这项研究提案将使我成为一名 独立、富有成效的研究科学家，因为我利用角膜MSCs的再生能力 首要目标是为角膜失明患者按需恢复视力。