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February 01, 2009 (Vol. 29, No. 3)

Three-Dimensional Cultivation of Cells

Rotating Bed Bioreactor Design of System Allows for 3-D Growth

  • Ex Vivo

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    Figure 1. Scalable rotating bed bioreactors

    Zellwerk provides a novel technology for 3-D cultivation of cells that differs fundamentally from conventional 2-D methods. In Z®-RP bioreactors, adherent mammalian cells, as well as other eukaryotic cells, grow three-dimensionally. This capability is a result of the rotating bed bioreactor design combined with cell type specific carriers (e.g., highly porous Sponceram® carriers for expansion of mesenchymal stem cells). The bioreactor operates in perfusion mode, which is essential to maintain cell viability in thick, tissue-like layers of cells embedded in ECM.

    Adhered cells are gently moved alternately through medium and overlay atmosphere. This “mud-flat” environment ensures that cells receive a supply of nutrients and gases and allows the removal of waste. Bioreactor vessels with different volumes are available, enabling scale up from 50 mL to 5,000 mL media volume (Figure 1).

  • cGMP Cell Processing

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    Figure 2. Z-RP cell cultivation composed of a rotating bed bioreactor 5 mL, housed in a GMP Breeder, a control unit, and corresponding software for process steering and regulation.

    Cell-therapy and tissue-engineering approaches, as well as biopharmaceutical production, require closely controlled environments for cell processing. Validatable seeding, expansion, and harvesting procedures must be established for safe and reproducible cell products. A 3-D cell culture system for advanced therapies and drug production must address these issues.

    The Z-RP cell cultivation system (rotating bed bioreactor, GMP Breeder, control unit and control software) complies with GMP standards and allows fast transfer of research results to therapeutic applications (Figure 2).

    Documented and fully automated 3-D cultivation processes can be established with the Z-RP. The Breeder provides a sterile cleanroom environment with temperature control for the bioreactor. Double UV-sterilization and other supply functions are also integrated.

    The Z-RP system is a fully functional cell culture lab concentrated into 3m2. It can be used as a tool for GMP-compliant production of autologous and allogenous cell formulations. The platform is also suitable for production of complex recombinant biopharmaceuticals.

    Three-dimensional cell culturing in a Z-RP cell culturing system can be used to expand stem cells and other primary cells. Mesenchymal human stem cells from different sources (e.g., bone marrow, umbilical cord vessels, adipose tissue) can be expanded to several mm thick layers of viable cells being embedded in self-generated ECM (Figure 3).

    Application-specific carriers (e.g., ceramic Sponceram discs with special surface characteristics due to coated ceramic materials, macroporous carrier possessing micro- and nano-roughness and porosity, respectively) allow expansion of MSCs either without further differentiation or differentiation into specialized cells, e.g., osteocytes, myocytes, or chondrocytes.

    Many other primary cells were grown in 3-D culture to functional tissue-like constructs (e.g., chondrocytes, ceratinocytes, hepatocytes, fibroblasts). In Figure 3C and 3D, results achieved with 3-D expanded hepatocytes are shown. Harvesting of cell amounts from 107 to 1010 cells can be realized using the Z-RP technology and suitable carrier. Releasing and detaching cells from ECM and the carrier surface is feasible with this system.

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    Figure 3A, 3B, 3C, and 3D

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