Scaffolds for
tissue fabrication
Tissue engineering is an interdisciplinary and
multidisciplinary field. It has shown great promise in
generating living alternatives for harvested tissues
and organs for transplantation and reconstructive
surgery. Materials and fabrication technologies are
critically important for tissue engineering in designing
temporary, artificial extracellular matrices (scaffolds),
which support three-dimensional tissue formation.
This review briefly introduces the concept of tissue
engineering, and illustrates the relationship between
tissue engineering and materials science and
engineering. Important scaffold design principles are
described. The most frequently used materials and
fabrication technologies for scaffolds are reviewed.
Some exciting new developments in scaffold
materials and fabrication technologies are also
discussed.
Department of Biologic and Materials Sciences,
Department of Biomedical Engineering,
Macromolecular Science and Engineering Center,
The University of Michigan,
Ann Arbor, MI 48109-1078 USA
E-mail: mapx@umich.edu
30 May 2004
by Peter X. Ma
Imagine that one day a patient will go to a human
body shop for a prescription for a lost leg, a failing
liver, or a dysfunctional heart. In addition to the
physiological, medical, and genetic information of the
patient, the doctor will also collect three-dimensional
images of the patients remaining leg (in the case of
the lost-leg patient) with detailed anatomical
structures (bone, cartilage, tendons, ligaments, blood
vessels, muscles, nerves, skin, and so forth) and the
external shape. The doctor may collect the patients
saliva or other body fluids to extract genetic material
(DNA) and also a tiny piece of tissue or bone marrow
to obtain seeding cells for expansion.
After the patient leaves the body shop, a computer is used
by a tissue engineer to design the structure of the mirror leg
(the lost leg) based on the symmetrical remaining leg, using
various materials that simulate the extracellular matrices of
the tissues of the leg (known as scaffolds or templates).
Then, cells from a universal cell source, specifically
designed for the patient, or banked cells grown from the
patients own cells are seeded onto these scaffolds. These
engineered cell-scaffold components are then grown
separately and/or assembled in a special chamber
(bioreactor) that provides the right nutrients, regulating
molecules (such as proteins, growth factors, and
differentiation factors), physical and mechanical stimuli,
temperature, pressure, and mass transport conditions for cell
proliferation, differentiation, and tissue/organ formation.
While the tissue/organ is regenerating, the scaffolding
materials degrade and disappear, leaving nothing foreign to
the body. The regenerated leg or leg precursor will be
ISSN:1369 7021 Elsevier Ltd 2004 Open access under CC BY-NC-ND license.

surgically grafted onto the patient during the second visit to
the human body shop (Fig. 1). The engineered tissues will
have the capacity to grow, model, and remodel in concert
with the dynamic changes of the physiological environment
of the body. The grafted leg will integrate into the body. The
new leg will grow and age as the bodys natural leg. This
scenario is an example of what the field of tissue engineering
is hoping to do in the future.
Tissue engineering and scaffolds
Tissue engineering has been defined as an interdisciplinary
field that applies the principles of engineering and the life
sciences toward the development of biological substitutes
that restore, maintain, or improve tissue function1. There are
three approaches in tissue engineering1,2: (1) the use of
isolated cells or cell substitutes to replace those cells that
supply the needed function3; (2) the delivery of tissue-
inducing substances, such as growth and differentiation
factors, to targeted locations4,5; (3) growing cells in three-
dimensional scaffolds1.
The use of isolated cells or tissue-inducing substances is
considered when the defects are small and well contained. To
engineer tissues of practical size scale and predetermined
shapes, these two approaches are seriously limited. Therefore,
the third approach, i.e. growing cells in three-dimensional
scaffolds, has become increasingly active. In this approach,
scaffolds play a pivotal role. They guide cells to grow,
synthesize extracellular matrix and other biological
molecules, and facilitate the formation of functional tissues
and organs2,6.
There are a few basic requirements that have been widely
accepted for designing polymer scaffolds2. First, a scaffold
has to have high porosity and proper pore size. Second, a high
surface area is needed. Third, biodegradability is generally
required, and a proper degradation rate is needed to match
the rate of neotissue formation. Fourth, the scaffold must
have the required mechanical integrity to maintain the
predesigned tissue structure. Fifth, the scaffold should not be
toxic to the cells (i.e. biocompatible). Sixth, the scaffold
should positively interact with cells, including enhanced cell
adhesion, growth, migration, and differentiated function.
Materials for scaffolds
Polymers (macromolecules) are the primary materials for
scaffolds in various tissue engineering applications, including
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Fig. 1 Schematic diagram showing the tissue engineering concept using a hypothetical
example of leg regeneration. Scaffolding materials (temporary synthetic extracellular
matrices) are designed as a three-dimensional mirror image, on which cells grow and
regenerate the needed tissues. Because the scaffolding materials are biodegradable, they
will resorb after fulfilling the template function and leave nothing foreign in the patient.
bone and other mineralized tissues. A limited number of
inorganic materials are used in bone and mineralized tissue
engineering research. Here, the most commonly used and a
few new materials in tissue engineering are reviewed.
Materials for porous solid-state scaffolds
The materials discussed in this category can form solid, stable
porous structures to serve as predesigned three-dimensional
scaffolds. They generally do not dissolve or melt under
in vitro tissue culture conditions (in an aqueous medium) or
when implanted in vivo.
Linear aliphatic polyesters
Poly(glycolic acid) (PGA), poly(lactic acid) (PLA), and their
copolymers poly(lactic acid-co-glycolic acid) (PLGA) are a
family of linear aliphatic polyesters, which are most
frequently used in tissue engineering2,7,8. These polymers
degrade through hydrolysis of the ester bonds9.
PGA is one of the most widely used scaffolding
polymers10. Because of its relatively hydrophilic nature, PGA
degrades rapidly in aqueous solutions or in vivo, and loses
mechanical integrity between two and four weeks10,11. It has
been processed into nonwoven fibrous fabrics as one of the
most widely used scaffolds in tissue engineering today.
PLA is also widely used for scaffold fabrication. The extra
methyl group in the PLA repeating unit (compared with PGA)
makes it more hydrophobic, reduces the molecular affinity to
water, and leads to a slower hydrolysis rate. It takes many
months or even years for a PLA scaffold or implant to lose
May 2004 31
 REVIEW FEATURE
mechanical integrity in vitro or in vivo12,13. To achieve
intermediate degradation rates between PGA and PLA,
various lactic and glycolic acid ratios are used to synthesize
PLGAs. These polymers (PLA, PGA, and PLGAs) are among the
few synthetic polymers approved by the US Food and Drug
Administration (FDA) for certain human clinical applications.
There are other linear aliphatic polyesters, such as
poly(-caprolactone) (PCL)14 and poly(hydroxy butyrate)
(PHB)15, which are also used in tissue engineering research.
PCL degrades at a significantly slower rate than PLA, PGA,
and PLGA16. The slow degradation makes PCL less attractive
for general tissue engineering applications, but more
attractive for long-term implants and controlled release
applications. PCL-based copolymers have recently been
synthesized to improve degradation properties17. PHB is
made by microorganisms via fermentation18,19. PHB and
PHB-based copolymers degrade very slowly because of their
hydrophobic nature and, therefore, are less popular compared
with PGA, PLA, and PLGA for tissue engineering applications.
Other important synthetic biodegradable polymers
Poly(propylene fumarate) (PPF) can degrade through
hydrolysis of the ester bonds similar to glycolide and lactide
polymers20. Tyrosine-derived polymers have shown promising
biocompatibility21. These polymers have been used for bone
tissue engineering research20,22.
Segmented polyurethanes allow the structural variations
to achieve a range of mechanical properties23. Recent efforts
have been focused on the development of biodegradable
urethane-based polymers using less toxic diisocyanates24-26.
Polyphosphoesters27 and polyphosphazenes28,29 have been
frequently used for controlled release applications, and have
recently been explored for certain tissue engineering
applications30-32. Similarly, polyanhydrides and
poly(ortho esters) are primarily designed for controlled drug
delivery applications because of their surface erosion
properties33,34. Nevertheless, they have also been explored
for tissue engineering scaffolding applications35,36. This is not
surprising because there are some similar requirements on
biocompatibility and biodegradability in both controlled
release matrices and tissue engineering scaffolds.
Natural macromolecules
Natural polymers, such as proteins and polysaccharides, have
also been used for tissue engineering applications. Collagen is
a fibrous protein and a major natural extracellular matrix
component. It has been used for various tissue regeneration
32 May 2004
applications, especially for soft tissue repair37,38. On one
hand, collagen as a natural extracellular component has
useful biological properties desirable for tissue engineering
applications. Therefore, collagen-glycosaminoglycan (GAG)
copolymers are fabricated into scaffolds for tissue
engineering39,40. Denatured collagen (gelatin) is also
processed into porous materials for tissue repair41. On the
other hand, there are concerns over collagen because of the
potential pathogen transmission, immune reactions, poor
handling and mechanical properties, and less controlled
biodegradability.
Another category of well-known natural fibrous proteins is
silk. Silkworm silk has been used in textile production for
centuries, and has been used as nondegradable sutures for
decades because of its excellent tensile mechanical
properties42. This natural macromolecular material has
recently been introduced into the field of tissue
engineering43. Although silk is often characterized as a
nondegradable material, it can degrade in vivo via enzymatic
mechanisms. However, the degradation rate is very slow.
There is also some concern over cytotoxicity44. There is
research into the chemical modification of silk materials to
enhance biocompatibility. There are also other types of silks,
such as spider silks and even genetically engineered silks45,
which may be considered in certain applications.
Polysaccharides are another class of natural polymers.
For example, alginate46, chitosan47, and hyaluronate48 have
been used as porous solid-state tissue engineering scaffolds.
In addition to the use of relatively pure natural
macromolecules extracted from an animal or plant tissue
source, processed extracellular matrix (decellularized)
materials with multiple natural macromolecules are also used
as scaffolds for tissue engineering or repair applications. One
such example is small intestinal submucosa (SIS), which
contains type I collagen, GAGs, and some growth factors. SIS
has been used in the reconstruction of several tissue
types49,50. Similarly, other decellularized tissues, such as
urinary bladder submucosa, porcine heart valves, and human
dermis have been used for tissue engineering or repair51-53.
Again there are concerns over pathogen transmission and
immune rejection52.
Inorganic materials
In addition to the large variety of polymeric
(macromolecular) materials, certain inorganic compounds
have also been studied for bone and other mineralized tissue

engineering research54. These materials can be categorized as
porous bioactive glasses55 and calcium phosphates56. Within
the calcium phosphates, -tricalcium phosphate (-TCP),
hydroxyapatite (HAP) and its derivatives, and their
combinations are the most frequently used57-59. These
inorganic materials are widely considered to be
osteoconductive (their surface properties support osteoblastic
cells adhesion, growth, and differentiation). They are also
reported to be osteoinductive (capable of inducing bone
formation) resulting from their capacity to bind and
concentrate bone morphogenetic proteins (BMPs) in vivo56.
However, these inorganic materials are often difficult to
process into highly porous structures and are mechanically
brittle. To overcome these disadvantages, composite
materials with synthetic or natural polymers have been
explored for bone tissue engineering research60-65. Highly
porous composite scaffolds with strong mechanical properties
have been demonstrated60. Such scaffolds support three-
dimensional new bone tissue formation61.
Materials for hydrogel scaffolds
Hydrogels are cross-linked hydrophilic polymers that contain
large amounts of water without dissolution. Injectable
hydrogels are attractive candidates for certain tissue
engineering applications because of the ability to fill
irregularly shaped tissue defects, the allowance of minimally
invasive procedures such as arthroscopic surgeries, and the
ease of incorporation of cells and bioactive agents66-69.
Among the synthetic hydrogels, poly(ethylene glycol)
(PEG) is frequently studied. PEG-diacrylamide has been
synthesized, and a photopolymerization step can be
performed in contact with cells70,71. There remains some
concern over the toxicity of cross-linking agents. However, a
major limitation of PEG hydrogel for tissue engineering
scaffolds is its lack of degradability. Efforts are being made
to impart degradability to PEG by either formulating
copolymers with PLA, PGA, and PPF, or by introducing
enzymatically degradable linkages into the PEG
backbone72-76.
Semi-interpenetrating polymer networks consisting of
poly(N-isopropylacrylamide) and linear poly(acrylic acid)
chains have been developed as synthetic hydrogel scaffolds77.
Diblock copolypeptide amphiphiles containing charged and
hydrophobic segments have recently been reported to form
hydrogel scaffolds with promising thermal stability and
mechanical properties78. More studies are needed to evaluate
REVIEW FEATURE
these new synthetic hydrogels for tissue engineering
applications.
Alginates, polysaccharides from seaweeds, cross-linked
using calcium sulfate (CaSO4), have been used as cell delivery
vehicles for in vivo tissue engineering research79-81. However,
one of the major disadvantages of using CaSO4 is that
gelation kinetics are difficult to control, and the resulting
structure is not uniform. A method has recently been
developed to control the gelation rate so that cross-linking
occurs uniformly to form structurally regular and
mechanically strong three-dimensional alginate gels66.
Collagen, in addition to porous foam, has also been used
as a hydrogel for a variety of tissue repair and regeneration
studies, including blood vessel, skin, cartilage, ligaments, and
tendons82-90. Collagen has also been compounded with other
natural polymers, such as chitosan, to form composite
hydrogel scaffolds91.
Fibrin, involved in clotting, is another gel-forming fibrillar
protein that has been explored for tissue engineering
applications92-94. Fibrin gels are typically prepared by
combining fibrinogen and thrombin solutions to entrap cells
and form tissue engineering constructs. The fibrin gel has
promising properties93, but its degradation rate is rapid,
which could be challenging for certain applications.
There are also artificial proteins, such as elastin-mimicking
polypeptides developed with recombinant DNA technology,
which may serve as biomimetic hydrogel scaffolds95-97. This
novel synthetic technology may also allow the design of
chemical structure for functionalities, potentially for
synthetic hydrogels or solid-state scaffolds.
Scaffold fabrication technologies
In the body, tissues are organized into three-dimensional
structures as functional organs and organ systems6. To
engineer functional tissues and organs successfully, the
scaffolds have to be designed to facilitate cell distribution
and guide tissue regeneration in three dimensions.
Textile technologies
Earlier tissue engineering scaffolds comprising fibrous
biodegradable polymer fabrics were produced using textile
technologies. PGA, PLA, and other semicrystalline polymers
can be processed into fibers using textile technologies. One
of these scaffolds widely used in tissue engineering research
is PGA nonwoven scaffold (Fig. 2). These PGA scaffolds have
been used either alone or combined with other biodegradable
May 2004 33
 REVIEW FEATURE
polymers for the engineering of cartilage98-100, tendon101,
ureter102, intestine103, blood vessels104,105, heart valves106,
and other tissues. However, there are several limitations of
PGA nonwoven scaffolds, such as low mechanical strength,
fast degradation rate, difficulty in controlling pore shape, and
limited fiber diameter variations.
Particulate-leaching techniques
Particulate leaching is another technique that has been
widely used to fabricate scaffolds for tissue engineering
applications107,108. Briefly, salt is first ground into small
particles and those of the desired size are transferred into a
mold. A polymer solution is then cast into the salt-filled
mold. After the evaporation of the solvent, the salt crystals
are leached away using water to form the pores of the
scaffold (Fig. 3). The process is easy to carry out. The pore
size can be controlled by the size of the salt crystals and the
porosity by the salt/polymer ratio. However, certain critical
variables such as pore shape and inter-pore openings are not
controlled. To overcome these shortcomings and those of
textile technologies, new techniques are being developed.
Phase separation
A homogeneous multicomponent system, under certain
conditions, becomes thermodynamically unstable and tends
to separate into more than one phase in order to lower the
system free energy. A polymer solution separates into two
phases, a polymer-rich phase and a polymer-lean phase. After
the solvent is removed, the polymer-rich phase solidifies.
Phase-separation techniques have been used to fabricate
porous membranes for filtration and separation109. However,
the pores formed using such techniques usually have
Fig. 2 Scanning electron micrograph of a PGA nonwoven scaffold with a fiber diameter of
approximately 15 m. (From Ma2 2004 John Wiley & Sons.)
34 May 2004
Fig. 3 Scanning electron micrograph of a poly(l-lactic acid) (PLLA) foam fabricated using
the salt-leaching technique. (From Ma2 2004 John Wiley & Sons.)
diameters on the order of a few to tens of microns and are
often not uniformly distributed, which is not suitable for
tissue engineering applications. Controlled phase separation
processes, primarily thermally induced phase separation, have
recently been explored for scaffold fabrication7.
Solid-liquid phase separation
Phase separation can be achieved by lowering the
temperature to induce solvent crystallization from a polymer
solution. We define this process as a solid-liquid phase
separation (solid phase formation in a liquid phase). After the
removal of the solvent crystals (sublimation or solvent
exchange), the space originally taken by the solvent crystals
becomes pores. This technique can be used to fabricate
scaffolds from many types of polymers and polymeric
composite materials (Fig. 4)60,61.
By manipulating the phase separation conditions, various
pore structures can be achieved. For example, many tissues
(such as nerve, muscle, tendon, ligament, dentin, and so on)
have oriented tubular or fibrous bundle architectures. To
facilitate the organization and regeneration of such tissue
types, a scaffold with a high porosity and an oriented array of
open microtubules may be desirable. To achieve this goal, a
novel phase separation technique has been developed to
grow oriented rod-shaped crystals from a polymer solution.
After the removal of these rods, a parallel array of
microtubules is formed (Fig. 5a)110. This oriented tubular
scaffold has anisotropic mechanical properties similar to
fibrillar and tubular tissues, and has been shown to facilitate
cell organization into oriented tissues (Fig. 5b)110.
 REVIEW FEATURE

(a)

Fig. 4 Scanning electron micrograph of a PLLA scaffold fabricated using solid-liquid phase
separation. (From Ma et al.61 2001 John Wiley & Sons.)
(b)

Liquid-liquid phase separation

Lowering the temperature can induce the liquid-liquid phase
separation of a polymer solution with an upper critical
solution temperature. When such a process leads to the
formation of a bicontinuous structure (both the polymer-rich
and polymer-lean phases are continuous), a scaffold with an
open-pore structure is formed after the removal of the
solvent. For example, a mixture of dioxane and water has
been used for liquid-liquid phase separation to fabricate PLA
and PLGA scaffolds (Fig. 6)111-113.
Design of three-dimensional pore architecture
One of the common shortcomings of the fabrication Fig. 5 (a) Scanning electron micrograph of a PLLA scaffold with oriented microtubular
architecture, (b) MC3T3-E1 cells cultured on the PLLA scaffold for four weeks in vitro
technologies discussed above is the lack of precise control of (von Kossas silver nitrate staining). (From Ma and Zhang110 2001 John Wiley & Sons.)
the three-dimensional pore architecture of the scaffolds. To
tackle this problem, computer-assisted design and
manufacture (CAD/CAM) are being adopted114.
Such a technique was initially explored at Massachusetts
Institute of Technology115,116. One of the solid, free-form
fabrication (aka rapid prototyping) techniques, called three-
dimensional printing, was used. With such a technique,
complex-shaped objects were designed using CAD software.
Scaffolds were fabricated by ink-jet printing a binder onto
sequentially laid polymer powder layers117. However, the
smallness of the powder particles and the binder drops
(pixels) are limited (to a few hundred microns). The
accuracy of positioning the printing nozzle is also limited.
Therefore, the preciseness of the technology is seriously
Fig. 6 Scanning electron micrograph of a porous scaffold prepared from a 10% solution
limited2. Similarly, other rapid prototyping techniques of PLGA (85/15) in a mixture of dioxane and H2O. (From Ma and Zhang111 1999
such as fused deposition modeling (FDM)118 and stereo John Wiley & Sons.)

May 2004 35
 REVIEW FEATURE

(a) (b) (c) (d)

Fig. 7 Scaffold fabrication using rapid prototyped negative replica: (a) a computer-generated, three-dimensional negative replica of a scaffold with a cubical pore shape, (b) the negative
replica of the scaffold fabricated using a rapid prototyping machine, (c) a photograph of the polymer scaffold fabricated using the negative replica, and (d) a scanning electron
micrograph of the internal pore structure of the scaffold. (From Ma2 2004 John Wiley & Sons.)

lithography are also being explored for scaffold (a)

fabrication119-122.
Rapid prototyping techniques have inherent shortcomings
such as limited material selection and inadequate resolution.
In addition, the resulting constructs have structural
heterogeneity because of the pixel assembly nature of the
fabrication process. To overcome this shortcoming, a reverse
fabrication technique has been developed to fabricate a
negative replica of the scaffold2. A polymer solution is cast
into such a mold and solidified after the removal of the
solvent. The mold is then dissolved away to form the
polymer scaffold with the designed three-dimensional pore
network (Fig. 7). The scaffold is more homogeneous, but the
feature resolution is not improved. (b)
To achieve higher resolution for scaffolds with well-
controlled interconnected spherical pores, paraffin spheres
are fabricated by a dispersion method123. These paraffin
spheres are then transferred into a three-dimensional mold of
the designed shape. The spheres are bonded together through
a heat treatment process. A polymer solution is cast into the
paraffin assembly in the mold. After removal of the solvent,
the paraffin sphere assembly is dissolved away. In this way,
an interconnected spherical pore structure is created (Fig. 8).
Importantly, the features generated have significantly better
resolution than those achievable with current rapid
prototyping techniques. In addition, investment in expensive
equipment is not required, which allows the technology to be Fig. 8 Scanning electron micrographs of polymer scaffolds with interconnected spherical
easily adapted to a research, as well as an industrial, setting. pore structures prepared using paraffin spheres (porogen): (a) PLLA scaffolds, paraffin
spheres: 250-420 m, 100x; (b) PLGA (85/15), paraffin spheres: 420-500 m, 50x.
(From Ma and Choi123 2001 Mary Ann Liebert. )
Nano-featured and bioactive scaffolds
Scaffolds serve as temporary, artificial extracellular matrices features of a natural extracellular matrix in scaffold design.
to accommodate cells and support three-dimensional tissue It is now well known that many biologically functional
regeneration. Therefore, it is often beneficial to mimic certain molecules, extracellular matrix components, and cells interact

36 May 2004

at the nanoscale. Hence, nano-featured synthetic scaffold
design is one of the exciting new areas in tissue engineering.
Nano-fibrous scaffolds
Collagen is a major natural extracellular matrix component,
and possesses a fibrous structure with fiber bundles varying
in diameter from 50-500 nm124,125. To mimic the nano-
fibrous architecture, a few technologies have been developed
to engineer nano-fibrous scaffolds.
Electrospinning
Electrospinning was first introduced in early 1930s126 to
fabricate industrial or household nonwoven fabric products.
The technique has been rejuvenated over the past decade to
process biodegradable and/or biocompatible polymers
(macromolecules) into fibrous fabrics with an average fiber
diameter at micrometer or nanometer scales for tissue
engineering scaffolds. To form such fibers using
electrospinning, a polymer solution is forced through a
capillary, forming a drop of polymer solution at the tip.
A high voltage is applied between the tip and a grounded
collection target. When the electric field strength overcomes
the surface tension of the droplet, a polymer solution jet is
initiated and accelerated towards the collection target. As the
jet travels through the air, the solvent evaporates and a
nonwoven polymer fabric is formed on the target. To
generate preferential orientation or/and tubular structure, an
electrically grounded rotating drum is used as the collection
target. A few synthetic polymers (e.g. PGA, PLGA, PCL, and
synthetic polypeptides) and natural macromolecules
(e.g. collagen and fibrinogen) have been processed into
fibrous nonwoven scaffolds for tissue engineering
research127-134. However, there are challenges in using this
technique to fabricate complex three-dimensional scaffold
shapes and internal pore networks. In addition, the average
fiber diameter is usually on the larger side of the extracellular
matrix fibers, sometimes falling in the micrometer range.
Self-assembly
Self-assembly is another exciting research area, especially
where nano-sized and/or patterned biological structures are
concerned. Self-assembly is now loosely defined as the
autonomous organization of components into patterns or
structures without human intervention135. Such a strategy
has been used in developing nano-fibrous materials with
potential as tissue engineering scaffolds136.
To emulate the triple helical structure of collagen, peptide-
amphiphiles (PAs) have been synthesized137-139. A PA
REVIEW FEATURE
consists of a collagen sequence peptide head group
connected to a long-chain ester lipid. The peptide head
groups form a triple helical structure like collagen, while the
lipid tails associate via hydrophobic interactions to induce
and/or stabilize the three-dimensional structure. However,
supramolecular fiber formation has not been demonstrated.
To form supramolecular nano-fibers, more complex PAs
have been synthesized to contain several structural
domains140,141. These PAs can form nano-fibers 5-8 nm in
diameter and over 1 m in length, which are actually
cylindrical micelle tubes with hydrophobic tails in the center
and hydrophilic heads on the outside. The structure is
stabilized by disulfide bonds140. Ionic self-complementary
oligopeptides have been synthesized142,143 containing
alternating segments of ionic and hydrophobic amino acids.
These peptides have been used to form fibrous structures
with a fiber diameter of ~10 nm, on the lower side of
extracellular matrix fibers. The fibers have a -sheet
structure instead of helical one. Self-assembled nano-fiber
systems are, so far, limited to hydrogel format. To develop
such hydrogel scaffolds further, production cost, mechanical
properties, degradability, and macropore structure need
to be addressed.
Phase separation
The phase separation of a polymer solution can also be
considered a self-assembly process. Instead of assembling
small molecules, large molecules are aggregated into a new
phase from an initially homogeneous one-phase system.
A novel phase separation technique has been developed to
fabricate nano-fibrous materials from synthetic
biodegradable polymers7,111,144. For example, a poly(l-lactic
acid) (PLLA) solution is induced to phase separate and gel,
forming a polymer-rich nano-fibrous network. The solvent is
removed afterwards to fabricate the desired porous, solid-
state, nano-fibrous material (Figs. 9a and 9b)111, which has a
fiber diameter of 50-500 nm, similar to that of collagen.
To improve the three-dimensional structure of nano-
fibrous scaffolds for tissue engineering, techniques have
been developed to build predesigned macropore networks in
the nano-fibrous matrices6,144. For example, larger water-
soluble fibers (of diameter 100 m to 1 mm) are prepared
from sugar as a geometrical porogen element and
assembled into various three-dimensional structures. A PLLA
solution is then cast into this three-dimensional assembly
and is thermally induced to phase separate for nano-fibrous
May 2004 37
 REVIEW FEATURE
(a) (b)
Fig. 9 Scanning electron micrographs of nano-fibrous PLLA matrices: (a) nano-fibrous matrix prepared from PLLA/THF solution via phase separation, 500x; (b) nano-fibrous matrix
prepared from PLLA/THF solution via phase separation, 20 000x; (c) PLLA nano-fibrous scaffold with helicoidal tubular macropore network, 35x; (d) PLLA nano-fibrous scaffold with
helicoidal tubular macropore network, 250x. (From Ma and Zhang111 1999 John Wiley & Sons; Zhang and Ma6 2000 John Wiley & Sons.)
matrix formation. After solvent removal, the sugar fiber
assembly is dissolved away using water to achieve nano-
fibrous scaffolds with predesigned helicoidal tubular pore
network (Figs. 9c and 9d)6. Similarly, an interconnected
spherical pore network has been combined with phase-
separation techniques to generate nano-fibrous scaffolds with
interconnected spherical macro pores145. Such macroporous
and nano-fibrous scaffolds enhance protein adsorption and
cell adhesion146.
Nanocomposite scaffolds
Polymer/inorganic composite materials have been developed
for mineralized tissue engineering applications. To mimic the
size scale of mineral crystals in bone and other mineralized
tissues, nano-sized hydroxyapatite (nano-HAP) has been
compounded with synthetic polymers or natural
macromolecules to fabricate nano composite
scaffolds62,113,147. Nano-HAP/polymer composite scaffolds
have not only improved the mechanical properties of polymer
scaffolds, but also significantly enhanced protein adsorption
over micro-sized HAP/polymer scaffolds113. Enhanced protein
adsorption improves cell adhesion and function146.
Bioactive scaffolds
The ideal tissue engineering scaffold should positively
interact with cells, including enhanced cell adhesion, growth,
migration, and differentiated function. To achieve these
positive cell-scaffold interactions, surface or bulk
modifications of the polymers are often employed148-151.
The surface properties can be varied by either bulk or surface
modification. Bulk modification is typically realized by
copolymerization or functional group attachment to the
polymer chains before scaffold fabrication148,151,152, and
usually changes the mechanical and processing properties of
the polymers. Surface modification can be carried out after a
38 May 2004
(c) (d)
porous scaffold has been fabricated. For example, plasma
treatment alone or followed by chemical modification has
been used to modify polymer thin films and porous
scaffolds153,154. Such techniques are most effective on two-
dimensional film surfaces or very thin three-dimensional
constructs. In a complex porous three-dimensional scaffold,
the surface is not just the outside surface, but also the
internal three-dimensional surfaces. A simulated body fluid
has been used to modify the chemical composition of the
internal three-dimensional pore surfaces of polymer
scaffolds150,155. This biomimetic process has been shown to
be effective at introducing nano-sized, bone-like apatite into
the internal pore surfaces in situ, and may lead to improved
scaffolds for bone tissue engineering150,156. More three-
dimensional surface modification techniques are needed.
To program scaffolds with biological instructions, delivery
of bioactive molecules and genes has been integrated into
Fig. 10 Schematic of a biomimetic nano scaffold. The scaffold combines the novel nano-
fibrous architecture of a interconnected pore network with microspheres for controlled
release of putative regenerative factors. The nano-fibrous scaffolding design uses the
architectural features of collagen, providing a high surface area for cell attachment and
new matrix deposition, and an open structure allowing an interactive environment for
cell-cell, cell-nutrient, and cell-signal molecule interactions. The bone mineral mimicking
apatite enhances the osteoconductivity of the scaffold. The biodegradable microspheres
release regenerative factors in a controlled fashion in a targeted local environment.

the scaffold design for tissue engineering157-164. A model of a
bioactive scaffold (from our laboratory) integrating nano-
fibrous architecture, three-dimensional biomimetic surface
modification, and controlled factor release capacity is
illustrated in Fig. 10.
Conclusions
In summary, tissue engineering is one of the most exciting
interdisciplinary and multidisciplinary research areas today,
and is growing exponentially over time. Scaffold materials
and fabrication technologies play a pivotal role in tissue
engineering, and are fast evolving. This review is intended to
illustrate the important roles of materials science and
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Acknowledgments
The author wishes to acknowledge financial support from the National Institutes of Health
(DE14755 and DE15384, Biomaterials and Organogenesis Training Grants T32-DE07057
and T32-HD07505), DuPont Young Professor Award, Whitaker Foundation (RG-99-0137),
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