title: Scale up cultivation of primary human umbilical vein endothelial cells on microcarriers from spinner vessels to bioreactor fermentation.
abstract: 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.
authors.affiliation: Institute of Cell Culture Technology, University of Bielefeld, 33501, Bielefeld, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Duvar
authors.forename: J
authors.initials: J
authors.lastname: Müthing
authors.forename: H
authors.initials: H
authors.lastname: Mohr
authors.forename: J
authors.initials: J
authors.lastname: Lehmann