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Cholesterol delivery to NS0 cells: Challenges and solutions in disposable linear low-density polyethylene-based bioreactors

Paper ID Volume ID Publish Year Pages File Format Full-Text
22360 43271 2007 10 PDF Available
Title
Cholesterol delivery to NS0 cells: Challenges and solutions in disposable linear low-density polyethylene-based bioreactors
Abstract

We report the successful cultivation of cholesterol dependent NS0 cells in linear low-density polyethylene (LLDPE) Wave Bioreactors when employing a low ratio of cyclodextrin to cholesterol additive mixture. While cultivation of NS0 cells in Wave Bioreactors was successful when using a culture medium supplemented with fetal bovine serum (FBS), cultivation with the same culture medium supplemented with cholesterol-lipid concentrate (CLC), which contains lipids and synthetic cholesterol coupled with the carrier methyl-β-cyclodextrin (mβCD), proved to be problematic. However, it was possible to cultivate NS0 cells in the medium supplemented with CLC when using conventional cultivation vessels such as disposable polycarbonate shake-flasks and glass bioreactors. A series of experiments investigating the effect of the physical conditions in Wave Bioreactors (e.g., rocking rate/angle, gas delivery mode) ruled out their likely influence, while the exposure of the cells to small squares of Wave Bioreactor film resulted in a lack of growth as in the Wave Bioreactor, suggesting an interaction between the cells, the CLC, and the LLDPE contact surface. Further experiments with both cholesterol-independent and cholesterol-dependent NS0 cells established that the concurrent presence of mβCD in the culture medium and the LLDPE film was sufficient to inhibit growth for both cell types. By reducing the excess mβCD added to the culture medium, it was possible to successfully cultivate cholesterol-dependent NS0 cells in Wave Bioreactors using a cholesterol-mβCD complex as the sole source of exogenous cholesterol. We propose that the mechanism of growth inhibition involves the extraction of cholesterol from cell membranes by the excess mβCD in the medium, followed with the irreversible adsorption or entrapment of the cholesterol-mβCD complexes to the LLDPE surface of the Wave Bioreactor. Controlling and mitigating these negative interactions enabled the routine utilization of disposable bioreactors for the cultivation of cholesterol-dependent NS0 cell lines in conjunction with an animal component-free cultivation medium.

Keywords
disposable bioreactor; NS0 cell line; cholesterol; cyclodextrin; animal component-free medium
First Page Preview
Cholesterol delivery to NS0 cells: Challenges and solutions in disposable linear low-density polyethylene-based bioreactors
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 103, Issue 1, January 2007, Pages 50–59
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering