Nanoscale Assembly of Bioactive Ligands to Enhance Endothelial Differentiation

生物活性配体的纳米级组装以增强内皮分化

• 批准号: 8241196
• 负责人: Wei Shen
• 金额: $ 18.09万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-01-09 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8241196
• 关键词: Address; Adhesives; Autologous; Binding Sites; Biomimetics; CD34 gene; Cells; Characteristics; Cysteine; Developmental Biology; Endothelial Cells; Engineering; Exhibits; Fibronectins; Future; Hematopoietic; Human; Integrins; Ligands; Literature; Methods; Molecular Structure; Peptides; Pluripotent Stem Cells; Positioning Attribute; Production; Proliferating; Protein Engineering; Research; Research Personnel; Site; Source; Stem cells; Stents; Structure; Surface; Technology; Testing; Tissues; Umbilical Cord Blood; Variant; Vascular Endothelial Growth Factors; Vascular Graft; Work; base; biological systems; density; design; experience; human embryonic stem cell; induced pluripotent stem cell; mimetics; nanoscale; neovascularization; polypeptide; self assembly; stem cell technology; success; surface coating; synergism

DESCRIPTION (provided by applicant): Endothelial cells have important biomedical applications ranging from enhancing the patency of engineered vascular grafts and stents to promoting neovascularization in ischemic tissues. But their limited availability hinders the success of endothelial-cell-related technologies. The advances in stem cell technology offer a unique opportunity to address this issue. In particular, endothelial cells have been derived from human pluripotent stem cells (hPSCs), which can proliferate extensively and virtually provide an unlimited cell source. The recent success in making induced PSCs (iPSCs) offers additional advantages in providing immunologically compatible autologous hPSCs and enabling "personalized" therapy in the future. The key to exploiting this opportunity to advance endothelial-cell-related technologies is our ability to guide endothelial differentiation. In currently used methods, hPSCs are differentiated into hemangioblasts, which have both hematopoietic and endothelial potentials, followed by differentiation of hemangioblasts into endothelial cells in the presence of VEGF and fibronectin(FN)-coated surfaces. VEGF and FN are both essential for efficient endothelial differentiation, and they exhibit a synergistic effect due to the unique structure of FN, which has a cell-adhesive site and a VEGF-binding site positioned in nanoscale proximity. However, naturally-derived FN has batch-to-batch variations. In addition, covalently immobilized FN has structural change that blocks the cell- adhesive ligand; physically adsorbed FN preserves the active cell-adhesive domain but does not allow precise control of surface ligand density. Therefore, cell microenvironments created with FN are not tightly controlled, hampering consistent production of endothelial cells from stem cells. This problem can be addressed by using well-controlled synthetic materials that recapitulate the essential molecular structure underlying the synergistic effect of VEGF and FN in regulating endothelial differentiation. The objective of this application is to develop synthetic materials having the essential structural characteristics underlying the synergistic effect of VEGF and FN and to use these materials to guide endothelial differentiation of human iPSC-derived hemangioblasts. Our central hypothesis is that a cell-adhesive peptide and a VEGF-mimetic peptide fused to a pair of heterodimerizing coiled-coils, respectively, can be brought into nanoscale proximity through coiled-coil self- assembly and the materials functionalized with the heterodimer, together with soluble factors, will create well- controlled cell microenvironments for efficient and reproducible endothelial differentiation of iPSC-derived hemangioblasts. The specific aims are: (1) design, synthesize, characterize, and immobilize the polypeptides that self-assemble to present a cell-adhesive peptide and a VEGF-mimetic peptide in nanoscale proximity; (2) examine endothelial differentiation of human iPSC-derived hemangioblasts on the polypeptide-functionalized substrates. Successful completion of this project will result in well-controlled, biomimetic cell microenvironments for efficient and robust endothelial differentiation of iPSC-derived hemangioblasts. PUBLIC HEALTH RELEVANCE: The proposed project aims to engineer rationally designed, well-controlled synthetic cell microenvironments to guide efficient and reproducible endothelial differentiation of hemangioblasts derived from human induced pluripotent stem cells. Such derived endothelial cells will have important biomedical applications ranging from enhancing the patency of engineered vascular grafts and stents to promoting neovascularization in ischemic tissues.

描述（由申请人提供）：内皮细胞具有重要的生物医学应用，从增强工程血管移植物和支架的通畅性到促进缺血组织中的新血管形成。但它们的可用性有限阻碍了内皮细胞相关技术的成功。干细胞技术的进步为解决这个问题提供了独特的机会。特别是，内皮细胞源自人类多能干细胞（hPSC），它可以广泛增殖，几乎提供了无限的细胞来源。最近在诱导性 PSC (iPSC) 制造方面取得的成功，为提供免疫相容的自体 hPSC 和未来实现“个性化”治疗提供了额外的优势。利用这个机会推进内皮细胞相关技术的关键是我们引导内皮分化的能力。在目前使用的方法中，hPSC 分化为具有造血和内皮潜能的成血管细胞，然后在 VEGF 和纤连蛋白 (FN) 涂层表面存在的情况下将成血管细胞分化为内皮细胞。 VEGF 和 FN 对于有效的内皮分化都是必需的，并且由于 FN 的独特结构（其具有纳米级邻近的细胞粘附位点和 VEGF 结合位点），它们表现出协同效应。然而，天然衍生的 FN 存在批次间差异。此外，共价固定的 FN 具有阻断细胞粘附配体的结构变化；物理吸附的 FN 保留了活性细胞粘附域，但不允许精确控制表面配体密度。因此，用 FN 创建的细胞微环境不受严格控制，阻碍了干细胞持续产生内皮细胞。这个问题可以通过使用控制良好的合成材料来解决，这些材料概括了 VEGF 和 FN 在调节内皮分化中协同作用的基本分子结构。本申请的目的是开发具有 VEGF 和 FN 协同作用的基本结构特征的合成材料，并使用这些材料指导人 iPSC 衍生的成血管细胞的内皮分化。我们的中心假设是，分别与一对异二聚化卷曲螺旋融合的​​细胞粘附肽和 VEGF 模拟肽可以通过卷曲螺旋自组装和用异二聚体功能化的材料实现纳米级接近。与可溶性因子一起，将为 iPSC 衍生的成血管细胞高效且可重复的内皮分化创造良好控制的细胞微环境。具体目标是：（1）设计、合成、表征和固定自组装的多肽，以纳米级接近的方式呈现细胞粘附肽和 VEGF 模拟肽； (2) 检查人 iPSC 衍生的成血管细胞在多肽功能化基质上的内皮分化。该项目的成功完成将产生良好控制的仿生细胞微环境，以实现 iPSC 衍生的成血管细胞高效、稳健的内皮分化。 公共健康相关性：拟议项目旨在设计合理设计、控制良好的合成细胞微环境，以指导源自人类诱导多能干细胞的成血管细胞的高效且可重复的内皮分化。这种衍生的内皮细胞将具有重要的生物医学应用，从增强工程血管移植物和支架的通畅性到促进缺血组织中的新血管形成。