NANO-PATTERNING OF BIOMATERIALS FOR BLOOD-VESSEL FORMATION IN ARTIFICIAL TISSUES

用于人造组织中血管形成的生物材料纳米图案化

• 批准号: 8669817
• 负责人: Patrick Benitez
• 金额: $ 0.75万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-25 至 2014-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8669817
• 关键词: Actins; Adhesions; American; Analysis of Variance; Angiopoietin-1; Animal Model; Area; Arginine; Artificial Organs; Aspartic Acid; Biocompatible Materials; Biology; Bioreactors; Blood Vessels; Cell Culture Techniques; Cell physiology; Cell-Matrix Junction; Cells; Chi-Square Tests; Chimeric Proteins; Clinic; Clinical; Coculture Techniques; Computer Analysis; Cytoplasmic Tail; Cytoskeletal Modeling; DNA; Data; Development; Elastin; Endothelial Cells; Engineering; Enzyme-Linked Immunosorbent Assay; Extracellular Matrix; Faculty; Failure; Fibronectins; Fluorescence Microscopy; Focal Adhesion Kinase 1; Glean; Glycine; Growth Factor; Human; Image Analysis; Implant; In Vitro; Integrins; Lead; Left; Length; Life; Ligands; Measures; Mechanical Stimulation; Mediating; Medical; Mentors; Mesenchymal; Metabolic; Molecular; Organ; Organ Transplantation; Oxygen; Patients; Peptide Hydrolases; Perfusion; Phosphorylation; Process; Protein Engineering; Proteins; Radial; Recombinants; Regenerative Medicine; Regression Analysis; Research; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Specific qualifier value; Speed; Staining method; Stains; Stem cells; Structure; Surface; Talin; Technology; Testing; Thick; Tissue Engineering; Tissues; Training; Translating; Translations; Transplantation; Vascular Endothelial Growth Factors; Vascularization; Work; base; blebbistatin; cell growth; cell motility; cellular engineering; clinically relevant; implantation; improved; in vivo; meetings; monolayer; nano; nanofiber; nanopattern; nanoscale; novel; polymerization; prevent; professor; receptor; regenerative therapy; scaffold; two-dimensional; vector

DESCRIPTION (provided by applicant): Patients suffering from failure of a vital organ can be treated with whole organ transplantation; transplantation alone, however, cannot meet the public's medical needs due to the limited supply of donated organs. Artificial tissues present an alternative to donated organs, but our inability to engineer functional microvessels within such constructs broadly prevents the development of clinically effective artificial tissues. Because all tissue- scale regenerative therapies require perfusion, the ability to form functional microvasculature is paramount. Upon implantation of bulk artificial tissues without microvasculature, cells on the inside die from lack of oxygen, leaving a shell of live cells about 0.2 mm thick. Artificial vascularization will prevent this by creating a volume- spanning perfusion-competent network and by enabling swift vascular integration after implantation. To engineer microvasculature, we propose a novel biomaterials strategy: nanoscale clustering of cell- matrix adhesion ligands. Previous work on 2D surfaces has shown that clustering of ligands increases growth factor sensitivity and motility via receptor clustering. Research using animal models has shown that expression of molecular disruptors of receptor clustering is associated with a decrease in both branching and maturation. Though ligand clustering and receptor clustering are related thermodynamically, it is unknown whether nanoscale ligand clustering will lead to morphologically appropriate microvasculature in a 3D, bulk biomaterial. To answer this question, we have developed a nanofibrous biomaterial that can be fabricated at a specified bulk concentration and nanoscale clustering of adhesion ligands. By mimicking the nanoscale order of the native the extracellular matrix, we expect to achieve organotypic blood-vessel structure formation in vitro. We specifically hypothesize that clustering of adhesion ligands will upregulate three essential cellular process that lead to formation of blood-vessels in vivo: (1) growth factor sensitivity, (2) cell motility, and (3) vessel branching and maturation. Growth factor sensitivity will be assessed by measuring proliferation, metabolic activity, and protease secretion. Motility, as parameterized by cell speed and persistence length, and cytoskeletal organization will be assessed by quantitative image analysis. Branching and maturation will be assessed by immunostaining for appropriate markers and computational analysis of morphological data. The biomaterials proposed here can be further developed as an implant for regenerative medicine by incorporating non- overlapping technologies such as co-culture of tissue-specific stems cells, growth factor delivery, and bioreactor/ mechanical stimulation. My mentor Sarah Heilshorn, an expert in protein-based materials engineering, and our collaborator John Cooke, a senior professor of microvascular signaling biology, have developed an appropriate training plan to accomplish this project.

描述（由申请人提供）：重要器官衰竭的患者可以通过整个器官移植进行治疗;但是，由于捐赠器官的供应有限，仅靠移植无法满足公众的医疗需求。人造组织是捐赠器官的替代品，但我们无法在这种结构中设计功能性微血管，这广泛阻碍了临床有效的人造组织的发展。因为所有 组织规模的再生治疗需要灌注，形成功能性微血管的能力是至关重要的。在植入没有微血管的大块人工组织时，内部的细胞因缺氧而死亡，留下约0.2 mm厚的活细胞壳。人工血管化将通过创建跨体积灌注能力网络和通过在植入后实现快速血管整合来防止这种情况。 为了构建微血管系统，我们提出了一种新的生物材料策略：细胞-基质粘附配体的纳米级聚集.先前在2D表面上的工作已经表明，配体的聚集通过受体聚集增加生长因子的敏感性和运动性。使用动物模型的研究表明，受体聚集的分子干扰物的表达与分支和成熟的减少有关。虽然配体聚集和受体聚集在化学上相关，但纳米级配体聚集是否会导致3D散装生物材料中形态学上合适的微血管系统尚不清楚。为了回答这个问题，我们已经开发了一种纳米纤维生物材料，可以在一个特定的体积浓度和纳米级的粘附配体集群制造。通过模拟天然细胞外基质的纳米级顺序，我们期望在体外实现器官型血管结构的形成。我们特别假设，粘附配体的聚集将上调导致血管形成的三个基本细胞过程， 体内：（1）生长因子敏感性，（2）细胞运动性，和（3）血管分支和成熟。将通过测量增殖、代谢活性和蛋白酶分泌来评估生长因子敏感性。将通过定量图像分析评估运动性（通过细胞速度和持续长度进行参数化）和细胞骨架组织。将通过适当标记物的免疫染色和形态学数据的计算分析来评估分支和成熟。本文提出的生物材料可以通过结合非重叠技术（例如组织特异性干细胞的共培养、生长因子递送和生物反应器/机械刺激）进一步开发为用于再生医学的植入物。 我的导师Sarah Heilshorn是蛋白质材料工程方面的专家，我们的合作者John Cooke是微血管信号生物学的高级教授，他们已经制定了一个合适的培训计划来完成这个项目。

项目成果

Sequence-specific crosslinking of electrospun, elastin-like protein preserves bioactivity and native-like mechanics.

• DOI: 10.1002/adhm.201200115
• 发表时间: 2013-01
• 期刊: ADVANCED HEALTHCARE MATERIALS
• 影响因子: 10
• 作者: Benitez, Patrick L.;Sweet, Jeffrey A.;Fink, Helen;Chennazhi, Krishna P.;Nair, Shantikumar V.;Enejder, Annika;Heilshorn, Sarah C.
• 通讯作者: Heilshorn, Sarah C.