A NOVEL BIOREACTOR F
OR CELLULAR ELECTRIC
Fabio Zomer Volpato
, Enrico Merzari
, Claudio Migliaresi
, Dietmar W. Hutmacher
BIOtech Research Center, University of Trento, Trento (TN), Italy
of Health and Biomedical Innovation, Queensland University of Technology, Brisbane (QLD),
In this work we present the design and development of a bioreactor for in vitro electrical
which delivers direct or alternated electrical potential difference (ΔV) to seeded scaffolds. This
manuscript presents the technical considerations regarding the development of the device and its potential in
tissue engineering, more specifically for neur
al stimulation. The equipment was built using National Instruments
(NI) CompactRIO hardware
Technical specifications of the hardware include a real
compactRIO 9075 chassis, alternated/continuous output voltage supply (+
10 V and max 1 mA
per channel) and
current input for experiment feedback. Culture chambers where produced from polycarbonate with gold
electrodes. The controlling software was designed in NI Labview environment, which allows users to control the
delivery of the voltage to f
our independent channels and can stimulate
culture chambers independently and
simultaneously. Electrical stimulation through functional electrical stimulators (FES) or bioreactors is a fairly
simple, flexible and realistic technique to translate the bi
oelectricity present in the human body to in vitro and in
dimensional cell cultures.
Electrical Stimulation, Neural Cells Stimulation
is a multidisciplinary field that unifies cell and molecular
biology, materials science, and medical procedures. It aims to tackle the problem of tissue and
organ regeneration and replacement. Tissue regeneration strategies can be either conducted
or assisted by an
phase, which may provide more stable environment
for cell survival. Commonly static cultures are used during the
with advances in tissue engineering in the past decade, scientists have start
ed to apply
dynamic conditions through the use of bioreactors for
cultures. A bioreactor can be
defined as ‘any apparatus that attempts to mimic physiological conditions in order to maintain
and encourage tissue regeneration in three
Tissue Engineering has
been exploring biomimetic approaches in order to enhance tissue formation and healing.
he physiological conditions can
enhance and guide cell adhesion, proliferation
and differentiation, increase extracellular m
atrix synthesis and growth factors secretion
Specific bioreactors designs have been developed t
o improve structure and function of
engineered tissues, such as: applying mechanical stimulus for bone and cartilage tissue
electrical stimulus for neural and muscular tissues
flow stimulus for heart
and perfusion for liver tissue
Several works have shown the benefits of
application of electrical stimuli
, spiral ganglion neuron
manuscript presents the
design and development of a bioreactor for in vitro
deliver direct or alternated electrical potential difference
ΔV) to seeded scaffolds
The design was tune
to neural stimulation; nevertheless, the
versatile platform in which it has been built allows users to simply modify and deliver a
variety of signa
we present the technical considerations regarding the development of
DESIGN AND DEVELOPMENT
The equipment was
time embedded controller
time embedded controller
9075 was connected to a NI cRIO
9075 chassis, containing an FPGA
Programmable Gate Array
of 3 Mgate reconfigurable I / O w
ith a clock frequency
of 40 MHz.
system is a versatile platform that allows users to deliver and acquire a
variety of signals, in parallel, through its embedded
FPGA. The user is able to
input and output signals, as required by each a
interchanging the I/O
are placed in the cRIO chassis.
The developed bioreactor with its culture chambers and support is presented in
The device was designed to
set of samples
Each set of samples can contain up to 4 scaffolds, as seen
, to arrive to a
total of 16 samples being stimulated contemporaneously. Two current input modules give the
current feedback of each scaffold during the experiments. The input channels were designed
allow 8 samples to work at high frequencies and 8 to work at lower frequencies but with a
high sensitivity of the input signal.
The culture chambers and its support were designed and manufactured in
polycarbonate. Gold electrodes were used in the chambers.
Such materials allow the
sterilization of the chambers via autoclave.
A summary of the hardware specifications is
National Instruments (NI) CompactRIO, [A] cRIO 9075 chassis, [B] output N9263 module, [C] and [D] input
modules N9203 and N9208, respectively.
Hardware specifications for the developed
± 10 V
Output current per channel
max 1 mA
Number of output channels
analog converter resolution
Input current per channel
max ± 20 mA
max ± 22 mA
Number of input channels
digital converter resolution
Type of stimulation
Continuous or alternated
(square, triangular or sinusoidal
Max stimulation frequenc
Designed bioreactor system depicting the
and few chambers
The controlling software was programmed in NI Labview 11.0 environment.
software architecture is expanded over three main
levels. The FPGA programming, wher
I/O signals are generated,
acquired and analyzed from the controller to the stimulation
Signals are generated at high frequency (400 MHz), while it acquires and
amplifies the measurements. The FPGA works i
n a deterministic way which allows
simultaneous operations. Thus, the software permits the user to run parallel operations.
overview of the FPGA program can be seen in
Block diagram of the FPGA program.
time microprocessor and the FPGA are synchronized in a deterministic way
by the second programming level, which is permanently downloaded in the controller.
autonomous control of the stimulation
providing a high reliability, the
possibility to manage all the working parameters and finally enabling the operator access to
the front panel, directly or remotely via
nternet network. The measured data are then
published on the loc
al network via TCP/IP
protocol. An overview of the real
can be seen in
Block diagram o
f the real
The third programming level is a user
friendly graphical interface
. Such interface is
not downloaded in the controller, instead it resides in the operator
computer. This client
interface downloads the data from the real
time via TCP/IP protocol.
trol panel allows
the user to set the desired parameters for the experiments, such as type of stimulus
(continuous or alternated), type of wave (triangular, sinusoidal or square), periods of
and rest, and the data to be saved. The data is saved
in a .txt ASCII file and stored
in the controller FTP
. The user panel can be seen in
electrical potential difference can range from
10 V in
continuous mode or with square, triangular or sinusoidal
ramps at frequencies ranging from
to 2 kHz. A number of stimulation profiles can be imposed to each independent
channel by modifying the applied voltage
offset, frequency of stim
ulation, wave type,
number of stimulations
per day, duration of each stimulation and
resting time between
, as seen in
Example of a stimulation profile.
The system was developed to work inside a standard humidified incubator at 37 °C and
. Once the scaffolds are positioned in the culture chambers, the single chambers can
be fixed at the support and placed in the incubator.
Bioelectric potentials are generated by a number of different biological processes, and
are used by cells to govern
, to conduct impulses along nerve fibres, and to
regulate muscular contraction. It results from the conversion of chemical energy
energy. Human bioelectricity is mostly present the nervous system signalling, muscular
contraction and wound healing.
is part of a project that aims to apply the latest advances in Tissue
Engineering to enhance
ome after spinal cord injuries. Nevertheless, the
applicability of such system in different areas of TE has
high potential. The developed
was built in a versatile platform which allows the adaptation to different scaffold
systems as well
as additional output signals such as current.
The research leading to these results has received funding from the European Union, Seventh
Framework Programme [
grant agreement [
Marie Curie FP7
226070, acronym “progetto Trentino”
The authors also acknowledge
BIOTOOLS s.l.r. for the cooperation on the development of the bioreactor.
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