Cytotechnology

, Volume 57, Issue 3, pp 251–261 | Cite as

Evaluation and optimization of hepatocyte culture media factors by design of experiments (DoE) methodology

  • Jia Dong
  • Carl-Fredrik Mandenius
  • Marc Lübberstedt
  • Thomas Urbaniak
  • Andreas K. N. Nüssler
  • Daniel Knobeloch
  • Jörg C. Gerlach
  • Katrin Zeilinger
Original Research

Abstract

Optimization of cell culture media based on statistical experimental design methodology is a widely used approach for improving cultivation conditions. We applied this methodology to refine the composition of an established culture medium for growth of a human hepatoma cell line, C3A. A selection of growth factors and nutrient supplements were systematically screened according to standard design of experiments (DoE) procedures. The results of the screening indicated that the medium additives hepatocyte growth factor, oncostatin M, and fibroblast growth factor 4 significantly influenced the metabolic activities of the C3A cell line. Surface response methodology revealed that the optimum levels for these factors were 30 ng/ml for hepatocyte growth factor and 35 ng/ml for oncostatin M. Additional experiments on primary human hepatocyte cultures showed high variance in metabolic activities between cells from different individuals, making determination of optimal levels of factors more difficult. Still, it was possible to conclude that hepatocyte growth factor, epidermal growth factor, and oncostatin M had decisive effects on the metabolic functions of primary human hepatocytes.

Keywords

Design of experiments Bioreactor C3A Primary human hepatocytes Growth factors Culture medium optimization 

Abbreviations

DexM

Dexamethasone

DoE

Design of experiments

EGF

Epidermal growth factor

FGF4

Fibroblast growth factor 4

HGF

Hepatocyte growth factor

HSA

Human serum albumin

LDH

Lactate dehydrogenase

NicA

Nicotinamide

OSM

Oncostatin M

Notes

Acknowledgments

The study was performed within the EU FP6 STREP-project Vitrocellomics. The work on primary hepatocytes was partly funded by the Federal Ministry of Education and Research (BMBF, FKZ 01GG0732 and 01GG0731 to A.N.). The authors would like to thank Dr. Ursula Müller-Vieira, Pharmacelsus GmbH, Saarbrücken, Germany, for valuable contributions.

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Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Jia Dong
    • 1
  • Carl-Fredrik Mandenius
    • 1
  • Marc Lübberstedt
    • 2
  • Thomas Urbaniak
    • 2
  • Andreas K. N. Nüssler
    • 3
  • Daniel Knobeloch
    • 2
  • Jörg C. Gerlach
    • 2
    • 4
    • 5
  • Katrin Zeilinger
    • 2
    • 4
  1. 1.Division of Biotechnology/IFMLinköping UniversityLinkopingSweden
  2. 2.Department of SurgeryCharite Universitätsmedizin BerlinBerlinGermany
  3. 3.Department of TraumatologyTU Munich, MRIMunichGermany
  4. 4.Berlin Brandenburg Center for Regenerative Therapies (BCRT)Charite Universitätsmedizin BerlinBerlinGermany
  5. 5.Departments of Surgery and Bioengineering, McGowan Institute for Regenerative MedicineUniversity of PittsburghPittsburghUSA

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