NMR ANALYSIS OF BIOARTIFICAL LIVER FROM PROGENITORS

祖细胞生物人工肝的核磁共振分析

• 批准号: 2749415
• 负责人: Jeffrey M. Macdonald
• 金额: $ 3.15万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2749415
• 关键词: bioengineering /biomedical engineering; biomaterial development /preparation; biomedical equipment development; bioreactors; cell differentiation; liver; liver cells; liver function; nuclear magnetic resonance spectroscopy; prosthesis; tissue engineering

The long-term goal of this project is to develop a liver lobule model, or bioreactor, to study all aspects of liver biology. Initial investigations will use the human hepatic cell-line, HepG2, under two growth conditions, promoting (1) growth versus (2) differentiation. If successful, the experiments will be repeated with normal hepatic progenitors isolated from human livers using multiparametric fluorescence activated cell sorting (FACS) and a known antigenic profile. The extent of differentiation will be characterized using molecular biology techniques and correlated with NMRS-visible biomarkers of metabolism. HepG2 cells will be grown in a novel coaxial bioreactor that is the same dimension as the liver lobule, and coated with extracellular matrices described by the two culture conditions. The morphology and immunohistochemistry of microtomed bioreactor sections will be analyzed at different stages of growth and compared to the normal liver lobule. In situ hybridization assays of HepG2 cells in the microtomed sections will determine the localization of mRNAs transcribing extracellular matrix proteins. Normal liver lobule metabolism will be assessed in situ by NMRS. 31P NMRS will be used to monitor cell growth. P-450 activity will be measured in the hepatocytes by monitoring p-triflouromethylanisole metabolism by 19F NMRS. Gluconeogenesis/glycogen catabolism and glycolysis/glyconeogenesis, and glutathione levels and the glycine cycle will be measured by monitoring metabolism of l- 13C- glucose/3- 13C-pyruvate and 2- 13C-glycine, respectively, using 1H [13C]-HMQC (heteronuclear multiple-quantum coherence) NMRS. Ureogenesis and glutamine synthetase activity will be measured by 1H[15N]-HMQC NMRS detection of 1 5NH4 metabolism.

该项目的长期目标是开发肝小叶 模型或生物反应器，用于研究肝脏生物学的各个方面。 初步研究将使用人类肝细胞系 HepG2， 在两种生长条件下，促进（1）生长与（2）生长 差异化。如果成功，将重复实验 使用从人类肝脏中分离出的正常肝祖细胞 多参数荧光激活细胞分选 (FACS) 和 已知的抗原谱。分化程度将是 使用分子生物学技术和相关技术进行表征 具有 NMRS 可见的代谢生物标志物。 HepG2 细胞将 在新型同轴生物反应器中生长，其尺寸与 肝小叶，并涂有描述的细胞外基质 由两种文化条件决定。形态和 生物反应器切片的免疫组织化学分析 分析不同生长阶段并与正常情况进行比较 肝小叶。 HepG2细胞的原位杂交分析 切片将确定 mRNA 的定位 转录细胞外基质蛋白。正常肝小叶 代谢将通过 NMRS 进行原位评估。将使用 31P NMRS 监测细胞生长。 P-450 活性将在 通过监测对三氟甲基苯甲醚代谢来监测肝细胞 19F核磁共振谱。 糖异生/糖原分解代谢和 糖酵解/糖异生、谷胱甘肽水平和甘氨酸 周期将通过监测 l- 13C- 的代谢来测量 分别使用 1H 葡萄糖/3- 13C-丙酮酸和 2- 13C-甘氨酸 [13C]-HMQC（异核多量子相干）NMRS。 尿素生成和谷氨酰胺合成酶活性将通过以下方式测量 1H[15N]-HMQC NMRS检测1·5NH4代谢。

项目成果

Effect of flow configuration and membrane characteristics on membrane fouling in a novel multicoaxial hollow-fiber bioartificial liver.

流动配置和膜特性对新型多同轴中空纤维生物人工肝膜污染的影响。

• DOI: 10.1111/j.1749-6632.2001.tb03845.x
• 发表时间: 2001
• 期刊: Annals of the New York Academy of Sciences
• 影响因子: 5.2
• 作者: MacDonald,JM;Wolfe,SP;Roy-Chowdhury,I;Kubota,H;Reid,LM
• 通讯作者: Reid,LM

A novel multi-coaxial hollow fiber bioreactor for adherent cell types. Part 1: hydrodynamic studies.

一种用于贴壁细胞类型的新型多同轴中空纤维生物反应器。

• DOI: 10.1002/bit.10081
• 发表时间: 2002
• 期刊: Biotechnology and bioengineering
• 影响因子: 3.8
• 作者: Wolfe,StephenP;Hsu,Edward;Reid,LolaM;Macdonald,JeffreyM
• 通讯作者: Macdonald,JeffreyM