3D-Bioprinting coming of age-from cells to organs

3D-Bioprinting
coming of age-
from cells to
organs
Dr. Dan Thomas
daniel.thomas@engineer.com

Introduction
• Over the past decade, annual spending on pharmaceutical
development to treat many endocrinological systems has increased
exponentially.
• Currently, preclinical studies to test the safety and efficiency of new
drugs, use laboratory animals and traditional 2D cell culture models.
Neither of these methods are completely accurate reflections of how a
drug will react in a human patient.
• A solution has emerged in the form of 3D-Bioprinting technology,
developed for the scalable, accurate and repeatable deposition of
biologically active materials. With advances in this biomanufacturing
technology, durable biological tissues for use in testing new
pharmaceutical products are now being harnessed and refined.

3D Bioprinting research
• Determine ways in which we can deposit biomaterials in 3D
to produce living tissue structures for bodily repair?
• Repair or replace large numbers of human cells, which are
biofabricated into 3D tissues in order to restore normal
human function.
• Develop future modular laboratory on a chip systems
engineered to test drugs and treatments.

Making 3-D Tissue Systems
3D Tissue
IR Curable
Scaffolds
DNA smart
glueAlginate
PLGA
Living
Cells
Collagen
Growth
Factors

Bioprinting - Cells
• In 2012 we started to engineer the
process that allowed us to deposit
high resolution scaffold structures in
3D.
• This initially began with simple
cylinders made using an alginate.
• We subsequently developed a
means to suspend and deposit cells
in-situ.

• Invention and patent
granted for the first high
precision 3-D digital fluid
dispenser and heating
system.
• This provided the team with
the first actual working high
resolution 3-D Bioprinter.

1. A few stem cells are collected from a
host and are put into a growth medium
to be cultured into many hundreds of
millions of cells.
2. After a period of time in culture, these
cells (35 million per mL) are added to a
viscous suspension with 10% Alginate, 5%
Agarose and Antibiotics.
3. Software is used to control two special high
precision extruders in 3D. This deposits the
biologically active material and a scaffold
(0.05% HYA) which also provides oxygen and
nutrients to the cells. Special protein growth
factors are added in layers to differentiate the
cells into biological differentiated layers.
4. Infra Red laser light is used to cure
artificial channel scaffolds to form a
complex solid structure. These are initially
used to provide nutrients to the cells.
3D-Bioprinting the Tissue System

Starting at the Cell

Bioprinted Cartilage Knee implant
Knee Cartilage Structure
• In order to rebuild knee cartilage then we needed to seed biogels
with Chondrocyte cells (35 million cells per mL of gel) and then
Bioprint this into a 3D structure.
• This we found was able to result in the deposition of a 3D
bioprintable material which has similar properties to real cartilage.

3D Data Acquisition and Rendering
200µm
We can use conventional computer aided design to
define our tissue reconstruction.
We take a scan, this is rebuilt into a tool path which is
converted into an instruction set to be interpreted by the 3D-
Bioprinter.

By combining composite biogels
seeded with different cell types, then
we can make heterogeneous living
tissue implants which are patient
specific.

3-D Bioprinted Trachea
The construction of
tissues constructs using
computer-aided 3-D
Bioprinting has extended
towards the bio-
manufacturing of a
tracheal ring made from
cartilage.
Bioprinting processDesign of the structure

Complex 3D-Bioprinting
• A precision syringe driver is loaded with a bio-ink made up of a
hydrogel-based (alginate, agarose, antibiotics and sucrose) biogel
containing 20 million chondrocytes per mL.
• A second syringe driver containing a special hydrogel/collagen
scaffold with 10 million endothelial cells per mL
• A software control systems is used to control the bioprinter and
instructs each of the high precision stepper motors in 3D space.
After 21-Days.

Vascular 3-D Bioprinting
Bioprinting for reconstructive tissue engineering. In this case tissues
for the use in breast reconstruction following cancer.
Once the process is finished, the structure is removed and then
placed into a bioreactor for up to two-months.
A bioreactor helps to maintain viability of tissue constructs and
gives time necessary for post process tissue maturation, fusion and
remodelling.
Bioreactor processing can be used in combination with growth
factors that promote vascularisation, angiogenesis and
innervation.

A) Hematoxylin and eosin staining of immature cartilage tissue and auricular
constructs showing condensation of cells present and (B) immunolabeling
collagen VI (shown in green) in the pericellular region of chondrocytes via
nuclear staining (shown in blue).

• By using cell polymer 3D Bioprinting then we developed a means to
create complex patient specific tissues on demand.
• These can be made permanent or can be engineered to last a specific
period of time, they dissolve as the tissue matures.
• It is this technology which we hope to scale up and allow us to make
the first transplantable ear structure and then multi-purpose
transplantable structures.
Complex Structure Bioprinting

Organ on a chip technology
The next opportunity for research is in developing
organ on a chip technology to test drugs and
treatments.
So far this has focused on making vascular structures
that can be used the analyze any side-effects of new
pharmaceutical products.

Automated ways to test drugs
• We are building new machines that can automatically dose 3D areas
of tissues with drugs.
• We also build perfusion bioreactors that test tissue structures over
periods of months for the effects of stimulation and the influence of
drugs of cell behaviour.

Extra Cellular Matrix Ghost Scaffolds
• Development of the first way to effectively engineer a complex
tissue architecture.
• This is a biologically inspired technique which uses 3D materials
to act as a scaffold and combine biological and synthetic
materials to make replacement complex architectures of the
human heart.
• This ghost structure is used to act as a scaffold for cells to grow
onto and may be a means of engineering complex tissue
architectures.

Ghosts and Extracellular Matrix
• Extracellular matrix is the biochemical support
material between all cells.
• This is a key aspect of making complex dynamic
systems which have materials with potential for cell
adhesion.
• Research is developing ways to deposit nanofibers
into complex 3D shapes.

• Bioreactor processes are important, however, the complexity of
cell communication processes to put cells in the correct place to
make a functioning organ needs extensive further study.

Complex Organs

Summary
• Going forward, 3D-Bioprinting is being explored as a method for the creation of more
advanced structures. In the longer-term, this technology offers the potential to fabricate
organised tissue constructs. This is being engineered to repair and/or replace damaged
or diseased human tissues, and directly has a bearing on developing safer and more
effective healthcare treatments.
• It also opens up the opportunity for cost effective patient specific tissue engineering to
evolve. However, fundamental obstacles include balancing scaffold properties to;
optimise resolution, cell migration, proliferation and differentiation need to be overcome,
one step at a time.
• By further engineering this process then we can produce tissues which have measurable
mechanical, metabolic and functional properties. This is from the perspective of using
shaped scaffold bioprinting technology, which produces a complex organ structure.
• The potential to produce functional tissues on demand, made in a controlled and safe
way for use in humans could one day revolutionise the future of healthcare.

Dr Daniel Thomas
3Dynamic Systems

3D-Bioprinting coming of age-from cells to organs

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