Nano-Ceramic for Metabolic Stem Cell Engineering

用于代谢干细胞工程的纳米陶瓷

• 批准号: 6790765
• 负责人: KYONGBUM LEE
• 金额: $ 9.98万
• 依托单位: ANGSTROM MEDICA, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-15 至 2006-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6790765
• 关键词: bioinformatics; biotechnology; bone regeneration; cell differentiation; cell population study; ceramics; genetic manipulation; hydroxyapatites; mesenchyme; nanotechnology; osteoblasts; stem cell factor; stem cells; technology /technique development; tissue /cell culture; tissue engineering; tissue support frame

DESCRIPTION (provided by applicant): The overall objective of this project is to develop a clinically and commercially viable tissue engineered bone construct using nanocrystalline ceramic scaffolds for proliferating and differentiating adipose-derived progenitor cells into cells of the osteogenic lineage. The use of a nanostructured hydroxyapatite (nano-HAP) scaffold offers several advantages for bone tissue engineering: 1) emerging evidence indicates that nanophase substrates enhance the adhesion and adhesion-dependent functions of anchorage-dependent cells; 2) the ceramic nature of nano-HAP affords chemical stability to the scaffold and therefore confers the advantage of long-term shape integrity; 3) the scaffolds are mechanically robust and can support and protect the bone tissue construct during the implantation procedure and within the body, thus allowing integration of culture and implant steps. Increasing evidence shows that human adipose tissue contains mesenchymal stem cells (MSCs) capable of differentiating into osteoblasts. Unlike bone marrow cells, adipose cells are easy to obtain and result in minimal patient discomfort. This Phase I STTR project leverages the unique material properties of nano-HAP against powerful cell property analysis and design tools drawn from metabolic engineering to identify optimal scaffold parameters and medium conditions for in vitro expansion and differentiation of adipose MSCs into osteoblasts. This work hypothesizes that the surface topography of nano-HAP promotes interactions between cell membrane and matrix proteins that result in greater stability of adherent cells, and thus improve adhesion dependent cell functions. The specific aims are: 1) Expansion of adipose-derived adult stem cells on idealized 2D nano-surface; 2) Induction of differentiated osteogenic function markers; and 3) Directed modulation of osteoblast function by surface property modification. These specific aims will be achieved using the following research design and methods. First, parallel cell culture experiments will compare adhesion and proliferation of human adipose progenitor cells on nano-HAP and other commonly used scaffolding material. These experiments will also evaluate the effects of varying nano-HAP grain and pore sizes with or without matrix protein pretreatment. Second, dose response experiments will be followed by a factorial design experiment to systematically explore the known differentiation factor space. A newly developed bioinformatics tool will be used to functionally relate differentiation factor doses and combinations with extent and speed of differentiation. Emphasis will be placed on selective differentiation into osteoblasts by controlling for differentiation into adipocytes. Finally, metabolic flux analysis will be performed to investigate the hypothesis that intermediary metabolism may be directed to support differentiated osteoblast function, including matrix protein deposition and bone formation (mineralization). Nano-HAP surface properties, metabolic flux distribution, and osteoblast activity will be functionally related using the aforementioned bioinformatics tool. This project is expected to identify specific scaffold properties, medium factors, or enzyme targets for enhancing osteoblast function through further metabolic engineering.

描述（由申请人提供）：该项目的总体目标是开发一种临床和商业上可行的组织工程骨结构，使用纳米晶体陶瓷支架将脂肪来源的祖细胞增殖和分化为成骨谱系的细胞。纳米羟基磷灰石（纳米hap）支架的使用为骨组织工程提供了几个优势：1)新证据表明纳米基质增强了锚定依赖性细胞的粘附和粘附依赖功能；2)纳米hap的陶瓷性质为支架提供了化学稳定性，因此具有长期形状完整的优势；3)支架机械坚固，可以在植入过程中和体内支持和保护骨组织结构，从而实现培养和植入步骤的整合。越来越多的证据表明，人类脂肪组织含有能够分化成成骨细胞的间充质干细胞（MSCs）。与骨髓细胞不同，脂肪细胞很容易获得，并且患者的不适最小。本一期STTR项目利用纳米hap独特的材料特性，结合代谢工程中强大的细胞特性分析和设计工具，确定脂肪间充质干细胞体外扩增和分化为成骨细胞的最佳支架参数和培养基条件。本研究假设纳米hap的表面形貌促进了细胞膜和基质蛋白之间的相互作用，从而提高了粘附细胞的稳定性，从而改善了粘附依赖性细胞的功能。具体目标是：1)在理想的二维纳米表面上扩增脂肪来源的成体干细胞；2)诱导分化成骨功能标志物；3)通过表面性质修饰定向调节成骨细胞功能。这些具体目标将通过以下研究设计和方法来实现。首先，平行细胞培养实验将比较人类脂肪祖细胞在纳米hap和其他常用支架材料上的粘附和增殖。这些实验还将评估不同纳米hap颗粒和孔隙大小对基质蛋白预处理或不预处理的影响。其次，在剂量反应实验之后进行因子设计实验，系统探索已知的分化因子空间。一种新开发的生物信息学工具将用于将分化因子剂量和组合与分化程度和速度在功能上联系起来。重点将放在通过控制向脂肪细胞的分化而选择性分化为成骨细胞。最后，将进行代谢通量分析，以研究中间代谢可能直接支持分化成骨细胞功能的假设，包括基质蛋白沉积和骨形成（矿化）。纳米hap表面特性、代谢通量分布和成骨细胞活性将使用上述生物信息学工具进行功能相关研究。该项目有望通过进一步的代谢工程来确定增强成骨细胞功能的特定支架特性、介质因子或酶靶点。