CSEM Scientific and Technical Report 2008

Development of Three-dimensional Cell Culture Models for Microfluidic
Cytotoxicological Platforms
L. Barbe, S. Generelli, O.T. Guenat
Development of 3-D cell culture models on microcarriers will replace traditional cell growth in Petri dishes for more in-vivo like conditions for cells
and mini-organs within microfabricated cytotoxicological platforms
Cell based assays have contributed in a major way to the
reduction of animal test systems in the drug discovery and
screening process, saving the lives of a large number of
laboratory animals. However, most cell-based assays,
although being more complex than cell-free biochemical test
systems, still represent a highly artificial cellular environment
and thus have limited predictive value for clinical efficacy.
Indeed, it is well known that many cells of normal and
malignant origin lose some of their phenotypic and functional
characteristics when grown in monolayer or suspension
culture in vitro [1] . The shortcomings of such assays thus lend
strong support to the development and evaluation of complex,
three dimensional (3-D) culture systems that are, in principle,
known to better retain cellular and organotypic histomorphological
features and to model the human tissue
environment with increasing accuracy.
The application of such 3-D culture systems is increasingly
being investigated with respect to its potential for the costefficient
optimization of preclinical and pre-animal selection of
the most active effectors from a large pool of drug candidates.
This allows the replacement of a significant number of animal
test modules. However, 3-D assays have not yet been
incorporated into mainstream drug development processes
due to the more complex methodological requirements and
due to the lack of fully automated read-out systems. Thus,
scientists are encouraged to optimize tools for such advanced
tissue-type, cell-based in vitro screening strategies [2] .
Figure 1: Human kidney cells (HEK-293) covering alginate microcarriers
(“beads”)
Most in vitro experiments with adherent human cells are
performed in 2-D cultures in which cells are plated onto plastic
surfaces treated to stimulate cell binding. Depending on their
type, cells either grow directly on the plastic, secrete ECM
components that coat the plastic to facilitate cell adhesion, or
require pre-coating of the plastic with ECM. Standard 2D
cultures in the conventional Petri dishes poorly mimic in vivo
72
conditions of the cellular environment. In fact, soluble growth
factors are present at abnormally high concentrations, 3D
cues are largely absent, oxygen tension is too high, cell–cell
interactions are rarely organized and the cell volume/medium
volume is significantly smaller than that present in vivo. In
addition, the distances between cells are larger leading to
poorer cell-cell communication, the continuous nutrient supply
and the waste removal are absent, as well as a conservation
of a set point temperature, and a minimal stress. Attempts
have been made to overcome these problems using organ
culture and various laboratory-scale bioreactors but
microsystems provide a more effective means of controlling
cell microenvironment in vitro [3] .
CSEM is in the process of developing microfluidic platforms
for cell-based assays capable of detecting the toxicity of
chemicals or drugs on targeted cells. The objective of the
present project is to provide an adequate cell culture model for
such platforms. Preliminary results using alginate microcarriers
from industrial partner Hamilton AG (Bonaduz,
Switzerland) confirmed the feasibility of culture of multiple cell
lines (see Figure 1). Different surface coatings on the bead
surface allow growth of primary cells as well as stem cells.
CSEM is currently developing fast and reliable techniques to
introduce the microcarriers inside the microfluidic device in
order to evaluate cell viability, using real time fluorescent
microscopy, and the reproducibility of this approach. The next
step will be then to correlate fluorescent measurements with a
label-free detector based on ion selective electrodes [4] .
[1] J. Friedrich, et al., Int. J. Radiat. Biol., 2007, 83 (11-12), 849-871
[2] J. Friedrich, et al., J Biomol Screen, 2007, 12 (7), 925-37
[3] J. El-Ali, et al., Nature, 2006, 442 (7101), 403-11
[4] S. Generelli, et al., Lab Chip, 2008, 8 (7),1210-1215