Institute of Cell Culture Technology, University of Bielefeld, 33501, Bielefeld, Germany.
Cytotechnology. 1996 Jan;21(1):61-72. doi: 10.1007/BF00364837.
Five types of dextran-based microcarriers (Dormacell(™), Pfeifer and Langen) with different concentrations of dimeric DEAE anion-exchange groups (nitrogen contents from 1.2 up to 2.9%) were tested as growth substrates for the cultivation of human umbilical vein endothelial cells (HUVECs). All microcarriers were gelatinized before use to improve cell adhesion. The one with the highest DEAE-group density was found to be most suitable for HUVEC propagation reaching final cell densities of 8×10(5) viable cells ml(-1) (95% viability) using microcarrier concentrations of 3 g l(-1). Furthermore, metabolic data of glucose/lactate and amino acid metabolism are presented in this study. The concentrations of 18 amino acids were monitored throughout cultivation. A considerable decrease of glutamine and inverse increase of glutamate was observed. Cultivation with initial glucose concentration of 16.5 mmol l(-1) resulted in high glutamine consumption rates, whereas high glucose-supplemented starting culture medium (30 mmol l(-1)) gave considerably lowered rates, indicating altered glutamine metabolism due to different glucose feeding. The glucose consumption and lactate production rates increased 2.6 fold and 3.5 fold, respectively, due to switch over from low to high glucose supplemented cultures. The rate of glucose metabolism was found not to be directly related to cell growth, because almost identical growth rates and doubling times were obtained. Considering the remaining 16 amino acids measured, serine concentrations considerably declined and glycine as well as alanine concentrations raised strongly. Most amino acid values were found insignificantly altered during 14 days of cultivation. Spinner vessel cultures served as inoculum for up scale propagation of HUVECs in membrane stirred 2 liter bioreactors. About 5×10(9) HUVECs were produced, which were used for the isolation and structural characterization of glycosphingolipids, cell membrane compounds, which are suggested to be involved in e.g. selectin-carbohydrate interaction (cell-cell adhesion), carcinogenesis and atherogenesis.
五种不同浓度二聚 DEAE 阴离子交换基团（氮含量从 1.2 到 2.9%）的葡聚糖基微载体（Dormacell（™）、Pfeifer 和 Langen）被测试作为人脐静脉内皮细胞（HUVEC）培养的生长基质。所有微载体在使用前均被明胶化以改善细胞黏附。发现具有最高 DEAE 基团密度的微载体最适合 HUVEC 繁殖，使用 3 g/L 的微载体浓度可达到 8×10(5)个活细胞/ml(-1)的最终细胞密度（95%活力）。此外，本研究还提供了葡萄糖/乳酸和氨基酸代谢的代谢数据。在整个培养过程中监测了 18 种氨基酸的浓度。观察到谷氨酰胺浓度显著降低，谷氨酸浓度反向增加。用初始葡萄糖浓度为 16.5 mmol/L 进行培养会导致谷氨酰胺消耗率高，而高葡萄糖补充起始培养基（30 mmol/L）则导致谷氨酰胺消耗率显著降低，表明由于不同的葡萄糖喂养方式，谷氨酰胺代谢发生改变。由于从低葡萄糖补充培养切换到高葡萄糖补充培养，葡萄糖消耗和乳酸生成速率分别增加了 2.6 倍和 3.5 倍。葡萄糖代谢速率与细胞生长没有直接关系，因为获得了几乎相同的生长速率和倍增时间。考虑到测量的其余 16 种氨基酸，丝氨酸浓度显著下降，甘氨酸和丙氨酸浓度强烈升高。在 14 天的培养过程中，大多数氨基酸值没有明显改变。搅拌罐培养物用作大规模生产 HUVEC 的接种物，在膜搅拌 2 升生物反应器中进行。生产了约 5×10(9)个 HUVEC，用于糖脂的分离和结构特征分析，糖脂是细胞表面化合物，被认为参与选择素-碳水化合物相互作用（细胞-细胞黏附）、癌变和动脉粥样硬化形成等过程。
