Gene Therapy (2005) 12, 1725–1733
& 2005 Nature Publishing Group All rights reserved 0969-7128/05 $30.00
REVIEW
Gene Therapy Progress and Prospects:
In tissue engineering
J Polak1 and L Hench2
1
Tissue Engineering and Regenerative Medicine Centre, Imperial College London, UK; and 2Department of Materials, Imperial College
London, London, UK
Tissue engineering (TE) has existed for several years as
an area spanning many disciplines, including medicine
and engineering. The use of stem cells as a biological
basis for TE coupled with advances in materials science
has opened up an entirely new chapter in medicine and
holds the promise of major contributions to the repair,
Keywords: tissue engineering; regenerative; medicine; stem cells; biomaterials
In brief
Progress

Tissue engineering is focusing on the creation of
biohybrid organs.

Primary adult and foetal cells can be used for tissue
engineering.

Stem cells provide many advantages over mature
cells.

Embryonic stem cells are the source of all tissue
types.

Adult or somatic stem cells are less plastic than
embryonic stem cells but may be more appropriate
for treating some conditions.

Stem cells have great potential as therapeutic agents
and are already being used to repair tissues.

Developments in materials science are providing new
opportunities for tissue regeneration.

Bioceramics provide an ideal scaffold for engineering
of bone.

A variety of materials for tissue engineering scaffolds
have been assessed.

Clinical application of tissue engineering strategies
needs to meet a variety of challenges.

New techniques have been developed that test the
efficacy of engineered tissues in vitro.
Tissue engineering (TE) is focusing
on the creation of biohybrid organs
TE has emerged as a thriving new field of medical
science. Just a few years ago, most scientists believed that
human tissue could be replaced only with direct
Correspondence: Professor J Polak, Tissue Engineering and Regenerative
Medicine Centre, Imperial College London, Chelsea and Westminster
Campus, 369 Fulham Road, London SW10 9NH, UK
Published online 22 September 2005
www.nature.com/gt
replacement and regeneration of damaged tissues and
organs. In this article, we review the spectrum of stem
cells and scaffolds being investigated for their potential
applications in medicine.
Gene Therapy (2005) 12, 1725–1733. doi:10.1038/
sj.gt.3302651; published online 22 September 2005
Prospects

New and more efficient means to control stem cell
differentiation will emerge.

Many stem cell therapies currently at a basic stage or
being tested for safety and efficacy in animal models
will move rapidly, although to testing in humans
where robust, randomized clinical trials must be
carried out.

Understanding of mechanisms of cell engraftment
into a host tissue.

Expansion of clonally differentiated stem cells into a
cost-efficient, scalable process guaranteeing product
reproducibility/viability and satisfying regulations.
For this, growth of cells in bioreactors (growth
chambers equipped with stirrers and sensors) that
regulate the appropriate amounts of nutrients, gases
and waste products will be needed.

Design of man-made materials that support and
coordinate the growth of three-dimensional tissues
in the laboratory.

Design of appropriate delivery systems.

Design of strategies to overcome immunological
challenges and deal with regulatory and safety
issues.
transplants or full artificial parts made of plastic, metal
and/or computer chips. Now, scientists are focusing on
the creation of biohybrid organs. TE is best defined by its
goal: the design and construction in the laboratory of
living, functional components that can be used for the
maintenance, regeneration or replacement of malfunc-
tioning tissues. It is multidisciplinary, requiring expertise
from a wide range of fields including cell biology,
material sciences, engineering, physics and mathematics.
An alternative to organ replacement is the prospect
of in vivo repair or regeneration. Newts, tadpoles and
 Progress and prospects in tissue engineering
J Polak and L Hench
1726
zebrafish easily repair and regenerate damaged organs.
Research is being undertaken to study how such
regeneration is achieved and how we can harness the
body’s innate capacity for such functions. Perhaps, only
a small number of adult stem cells are needed to
stimulate the regenerative body responses, provided
that cells home to sites of injury and promote healing in
damaged tissues.
Primary adult and foetal cells can be used
for TE
A variety of cells has been used for TE. Initial work used
fully mature cells taken from adult tissue but, although
these have certain advantages including the possibility of
using autologous cells to avoid immunological problems,
they have low growth potential and a relatively high
degree of senescence. However, we and others have
demonstrated the ability of, for example, adult human
bone cells to attach, grow and differentiate when in
contact with specific man-made materials, such as
Bioglass.1,2 Fully committed cells from foetal tissue (7–
10 weeks gestation) have also been used and shown to be
a good source for TE. These cells have much greater
proliferative capacity and show less senescence than
adult cells.3,4
Stem cells provide many advantages over
mature cells
A stem cell is defined by three main criteria: self-renewal,
ability to differentiate into multiple cell types and
capability of in vivo reconstitution of a given tissue. The
most primitive is the fertilized oocyte (the zygote)
(Figure 1). Once the fertilized egg starts dividing, the
descendants of the first two divisions are the totipotent
Figure 1 Diagram showing the development and implantation of a human embryo highlighting the stage (blastocyst) at which embryonic stem cells are
derived (PGD ¼ prenatal genetic diagnosis).
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cells able to form the embryo and trophoblast. After
about 4 days, these totipotent cells form a hollow ball of
cells, the blastocyst, containing a cluster of cells called
the inner cell mass from which the embryo develops and
embryonic stem (ES) cells are derived.
Lower down in the hierarchical tree are the multipotent
cells. Most adult tissues have multipotent stem cells that
can produce a limited range of differentiated cell lineages
appropriate to their location. At the bottom of the
hierarchical tree are the unipotent stem cells, or com-
mitted progenitors, that generate one specific cell type.
Current sources of stem cells for TE include ES cells
and adult stem cells. However, concerns exist over the
use in TE of either source: for ES cells, there are ethical
considerations, the potential for tumorigenicity and a
current paucity of cell lines, while adult stem cells are
more limited in potential and often difficult to harvest
in sufficient numbers. Thus, the search continues for
an ethically conducive, easily accessible and abundant
source of stem cells.
ES cells are the source of all tissue types
Pluripotent stem cells can be divided into three different
types: embryonic germ (EG) derived from primordial
germ cells, ES cells and embryonic carcinoma (EC) cells.
These cells are widely held to be pluripotent, that is, able
to differentiate into all cells that arise from the three
germ layers but not the embryo.
The intellectual framework that led to the isolation
and derivation of ES cells began in the 1970s when
investigators discovered pluripotent cells in teratocarci-
nomas. Murine ES cells where isolated in the 1980s by
two major workers, Evans and Kaufman (Figure 2). After
17 years, ES cells were isolated successfully from human
blastocysts. Following multiple passages of ES cell
cultures, genetic and epigenetic changes can occur.5 A
unique network of transcription factors ensures self-

Figure 2 A cluster of living murine embryonic stem cells that have
differentiated in vitro spontaneously to form an embryoid body (inverted
microscopy; original magnification  200).
renewal and simultaneously suppresses differentiation
of ES cells. These include Nanog, octa-binding factor 3/4
(OCT4) and Wnt. Nanog was discovered by expression
cloning analysis and named after the mythological Celtic
land of the ever young, Tir nan Og.6 Oct4 is a member of
the Pit-Oct-Unc (POU) family of transcriptional regula-
tors restricted to early embryos.7 The Wnt family consists
of secreted and extracellular matrix (ECM)-associated
glycoproteins binding to frizzled seven-transmembrane
span receptors.8 Recently, global transcription factors for
human ES cells were identified using DNA microarray9,10
SAGE11 and cDNA library analysis.12 All demons-
trated the existence of gene clusters that are expressed
at signicantly higher levels in human ES cells compared
with fully differentiated cells. Human ES cells are
characterized by their expression of SSEA3, SSEA4,
TRA-1–60 and TRA-1–81 antigens that are now known
to be downregulated during differentiation while several
other antigens are induced.12
In preparation for the clinical application of human ES
cells, efforts are being made increasingly to (a) define the
culture media/conditions needed to derive specific cell
phenotypes and their progenitors and (b) use cultures
free of contamination with animal proteins. Questions
that need to be addressed before human ES cells can be
used clinically include:
(a) Can pure populations of cells be derived in sufficient
numbers for implantation?
(b) Can the necessary requirements of ‘good medical
practice’ be achieved (ie xeno-free)?
(c) How can potential problems of tissue rejection be
resolved?
Regarding the last question, undifferentiated me-
senchymal stem cells (MSC) do not express immunolo-
gically relevant cell surface markers and, as such, appear
to be immunoprivileged.13 For ES cells, a stem cell bank
has already been established by the Medical Research
Council in the UK, which will provide a variety of cell
Progress and prospects in tissue engineering
J Polak and L Hench
1727
lines to match the immune requirements of patient.
Somatic cell nuclear transfer (SCNT) is another method
that can provide immunocompatible cells. SCNT, or
therapeutic cloning, that has recently been used to
generate animals with a common genetic composition,
comprises the transfer of the nucleus of a somatic cell, for
example, from a patient needing TE, into an enucleated
donor oocyte. Subsequently, ES cells can be isolated from
the inner cell mass of the cloned preimplantation embryo
that carry the genome of the patient and, upon
implantation, the cells will not be rejected. Korean
workers recently reported the successful isolation of
human ES cells by this method.14,15
Politics, religious belief and the media have deter-
mined society’s current perception of the value and
medical potential of ES cells. The ethical antipathy
towards ES cells, which require the destruction of human
embryos for their derivation, has biased research
towards adult stem cells in some countries. The situation
in the UK is that the Human Embryology & Fertilization
Authority has given authorization to use and, in some
cases, create human embryos for therapeutic purposes.
Research into adult and ES cells is not sufficiently
advanced for a definitive and exclusive choice to be
made on whether one type is better or worse than the
other as a basis for developing a broad range of stem cell
therapies. Each is likely to have its own niche in therapy
or, for some conditions, a combination both of adult and
ES cells may prove best.
A variety of protocols has proved useful in driving the
differentiation of ES cells to particular lineages, including
the use of growth factor supplementation of media and
genetic modification. In our own laboratories, a robust
system has been developed for osteoblast and pneumo-
cyte (Figure 3) differentiation from murine and human
ES cells using a defined culture medium.16–18
Recent technological advances, such as RNA inter-
ference to knock down gene expression in ES cells, are
also producing enriched populations and elucidating
gene function in early development.19–21
The success of stem cell transplantation depends on
the ability of cells to interact with stromal cells and
repopulate the corresponding niches, a property of stem
cells known as homing. The mechanisms regulating
homing are not fully understood but signalling mole-
cules such as chemokines and growth factors are
involved and the expression of these increases following
tissue injury. Using the property of mammalian cells to
adapt and remodel in response to physiological/envir-
onmental cues, coculture of embryonic and other stem/
progenitor cells with mature cells or tissues is being
used increasingly to drive their differentiation towards
required lineages.22,23 Our recent work showed that
coculture with murine embryonic pulmonary mesench-
yme is an efficient means to upregulate the differentia-
tion of ES cells towards pneumocytes.24
Adult or somatic stem cells are less plastic
than ES cells but may be more appropriate
for treating some conditions
Traditionally, adult stem cells have been viewed as cells
committed to producing mature cells from the tissue
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 Progress and prospects in tissue engineering
J Polak and L Hench
1728
of origin but not cells of unrelated tissues. The first
evidence of ‘plasticity’ or ‘transdifferentiation’ stemmed
from the observation that, following gender mismatched
bone marrow transplants, some donor cells could be
found in multiple organs in the recipient, having
adopted the characteristics of cells of the organ in which
they had lodged. Various reports over the last 6 years
challenged this dogma by demonstrating that adult stem
cells, under certain microenvironmental conditions, give
rise to other cell types besides the original cell type,
indicating that they can switch cell fate. This phenom-
enon has been termed ‘stem cell plasticity’. However,
subsequent reports have generated both excitement as
well as scepticism.23,24 The concept of plasticity defies the
basic developmental biology principle of lineage restric-
tion being imparted during morphogenesis but, if
correct, the ability of adult stem cells to change fate
holds immense therapeutic potential. It is important,
therefore, that the concept of stem cell plasticity be
rigorously defined and experimentally proven. To
achieve this, the following criteria need to be met.
(i) Demonstration of a single clonal origin.
(ii) Successful engraftment in host tissue. Figure 3 Pneumocytes differentiated from human embryonic stem cells
(iii) Robust and reliable tracing of engrafted cells. and identified by their immunoreactivity for surfactant protein A (brown;
nuclei counterstained purple with haematoxylin) (original magnification
(iv) Restoration of function by engrafted cells in an  400).
animal model of injury and disease.

Bone marrow stem cells

The most studied adult stem cell, the haematopoietic
stem cell (HSC), resides in the bone marrow. The
presence of HSCs was demonstrated by the ability of
transplanted donor bone marrow stem cells to recon-
stitute the haematopoietic system of a lethally myelo-
ablated host. Both murine and human HSCs reside in
the lineage-negative fraction of bone marrow cells and,
as they express the ABCG2 transporter, populations can
be enriched by Hoechst dye exclusion. These cells are
termed side-population cells. Other stem cells are found
in the bone marrow and include MSC. Several groups
have shown the existence of a rare cell type that can be
cultured from bone marrow and other organs called the
multipotent adult progenitor cell or MAPC.25–27 These
cells, that are technically demanding to isolate, can
differentiate to osteoblasts, chondrocytes and adipocytes,
and are able to correct a particular defect in an animal
model.28 More recently, a common cell origin for
haematopoietic cells and osteoblasts has been demon-
strated in the bone marrow.29 More research needs to
be carried out to assess the relationship between MAPCs
to MSCs.
There is some evidence in animal models that bone
marrow stem cells can engraft in the lung and
differentiate to pulmonary alveolar epithelium where
they can repair injury. For the human lung, chimerism
has been demonstrated in human pulmonary epithelium,
including that of the alveoli, following transplantation of
HSCs30,31 and lung32 without evidence for engraftment of
cells from the bone marrow. However, we have recently
challenged the above by demonstrating bone marrow
origin of engrafted cells in the injured lung of female
patients who had received a male bone marrow
transplant.33
MSC are clonogenic, nonhaematopoietic cells present
in the bone marrow that can differentiate into multiple
mesoderm-type cell lineages, for example, osteoblasts,
chondrocytes, endothelial cells and also nonmesodermal
lineages, for example, neuronal-like cells. Under current
in vitro culture conditions that include foetal bovine
serum, MSC obtained from young donors can grow to
24–40 population doublings in vitro, while the prolifera-
tive potential of MSC obtained from older donors is more
compromised.34 As it is older individuals that are most
likely to require TE, this senescence reduces the applic-
ability of autologous MSC in tissue repair strategies.
Niche-specific stem cells
Adult tissues undergo continuous self-renewal and,
hence, require resident stem cells with the capacity for
multipotent differentiation. These stem cells are most
often slow-cycling and give rise to transient amplifying
cells, which impart the majority of tissue renewal in the
setting of injury. Unlike stem cells, which by definition
have unlimited proliferative capacity, transient amplify-
ing cells are restricted in their capacity to divide and
cannot proliferate indefinitely. Although stem cells and
transient amplifying cells are both self-renewing popula-
tions, the latter are thought primarily to be the major
progenitor population, which gives rise to terminally
differentiated cell types of an adult organ.
Four areas for potential clinical use of MSCs have been
explored: local implantation for localized diseases,
systemic transplantation, combining stem cell therapy
with gene therapy and use in TE protocols.
Other sources of stem cells
Many authors have recently reported the differentiation
of stem cells from other sites into multiple cell lineages
and newly identified sources include fat,35 placenta36 and
spleen.37 Such sites can be an abundant source of stem
cells; for example, a single gram of human adipose tissue

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has been reported to yield more than 70 000 pluripotent
adipose-derived adult stem cells after 24 h in culture.35
Stem cells have great potential as therapeutic
agents and are already being used to repair
tissues
Numerous studies have recently demonstrated success-
ful restoration of lost organ function by the use of a
variety of stem cells of different origins.38–47 For example,
several recent studies have highlighted the successful
treatment of acute myocardial infarction by injection of
autologous bone marrow cells.48–50
Developments in materials science are
proving new opportunities for tissue
regeneration
TE scaffolds have a dual purpose: to direct morphogen-
esis in vitro and maintain the structure and function of
the TE construct as it is integrated with the host tissues
postimplantation.51,52 An ideal scaffold for TE and
regeneration
1. is made from a material that is biocompatible, that is,
not cytotoxic;
2. acts as template for tissue growth in three dimen-
sions; for example, collagen is organized parallel
to the surface in articular cartilage and perpendicular
to the surface in the osteochondral region of an
articulating join;
3. has an interconnected macroporous network with
pore diameters in excess of 100 mm for cell penetra-
tion, tissue ingrowth, vascularization and nutrient
delivery throughout the regenerating tissue;
4. bonds to host tissue without formation of interven-
ing scar tissue;
5. exhibits a surface texture and chemistry that pro-
motes cell adhesion, adsorption of biological meta-
bolites, including growth factors;
6. influences the genes in stem cells to enhance
differentiation and proliferation of all the pheno-
types required for tissue regeneration while not
altering clonogenic or proliferative potential of the
cells;
7. resorbs at the same rate as the tissue is repaired, with
degradation products that are nontoxic and can be
easily excreted;
8. can be produced in irregular shapes to match the
tissue defect;
9. has mechanical properties sufficient to withstand
applied stresses in clinical applications;
10. has the potential to be commercially produced to the
required ISO (International Standards Organisation)
and FDA (Food and Drug Administration) regula-
tory standards at a cost suitable for routine clinical
use;
11. does not alter clonogenic and proliferative potential
of stem cells.
Numerous polymeric materials with molecularly
tailored structures and surfaces are being developed for
soft tissue applications in efforts to meet these 10
Progress and prospects in tissue engineering
J Polak and L Hench
1729
requirements.53,54 One goal is to use synthetic biomater-
ials to mimic the biological regulatory characteristics of
natural ECMs, especially to control the microenviron-
ment of stem cells55 and morphogenesis.56 Recent
advances include use of self-assembled small molecular
building blocks to produce nanofibrillar networks that
mimic the ECM,57 artificial ECM networks and ligand-
functionalized materials that induce response to cell-
secreted signals.58,59 Applications include: (a) scaffold
support of neural stem cells to enhance differentiation
into neural cells while inhibiting development of astro-
cytes59 and directing neurite outgrowth to reconstitute
lost neural tissues,60 (b) use of surface topography to
control keratinocyte function and epidermal organiza-
tion in artificial skin substitutes,61 (c) modulation of the
contractile and biosynthetic activity of chondrocytes
in TE cartilage constructs,62 (d) growth of microvascular
blood vessels by using fibroblast growth factors to
induce capillary endothelial cells to invade three-dimen-
sional (3-D) collagen or fibrin matrices to form tubules,63
control of the microenvironment of cells to promote
natural angiogenesis for vascular repair,64,65 and (e)
in vitro differentiation of progenitor cells to form
hepatocyte-like spheroid structures with some liver-like
functions.66,67
Bioceramics provide an ideal scaffold
for engineering of bone
The emphasis of our group has been to use bioactive
ceramics to produce an ideal scaffold for regeneration of
bone.1,2 Bioactive glass foam scaffolds51 meet the above
criteria as they have a hierarchical porous structure with
interconnected pores of 100–300 mm diameter, similar to
that of trabecular bone, bond to both bone and soft
connective tissues and release at controlled rates critical
concentrations of biologically active silicon and calcium
ions that upregulate seven families of genes in osteo-
progenitor cells.1,2,16 The hierarchical structure of the
bioactive foams can be tailored for tissue bonding,
resorption and delivery of dissolution products that
provide the controlled rates of release of osteogenic
stimuli,68–70 and the nanopores can be modified with
various proteins to control cell integrin–scaffold inter-
actions.71,72 The calcia–silica-based materials are easily
and economically processed to ISO standards69 and have
been used clinically for 20 years with FDA and CE
regulatory clearance. Our studies have shown that the
inorganic ionic dissolution products released during
controlled resorbtion of the gel-glass scaffolds stimulate
expression of genes and enhance osteogenesis in mouse
and human ES cells,16 adult human stem cells1 and
human foetal bone cells.2
A variety of materials for TE scaffolds have
been assessed
There is no consensus at this time as to the best material
to use in any TE application.51–57,62,65,73–79 One of the
difficulties is the plethora of materials being developed
and tested for use as scaffolds. Numerous biologically
derived polymers include: collagen, fibrin, agarose,
gelatine, alginates, hyluronan, chitosan, collagen/glyco-
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 Progress and prospects in tissue engineering
J Polak and L Hench
1730
saminoglycan. Synthetic polymers include: polylactic
acid (PLA), polyglycolic acid (PGA) PLA/PGA copoly-
mers, polycaprolactone, poly(ethylene oxide), poly(vinyl
alcohol), poly(acrylic acid), poly(propylene fumarate-
co-ethylene glycol).73,74 Ceramic materials include hydro-
xyapatite (HA), carbonate or silicon-substituted HA,
various calcium phosphates and alumina. Glasses
include various compositions in the SiO2–CaO–Na2O–
P2O5 systems. Sol–gel-derived bioactive gel-glasses and
foams include a wide range of compositions in the SiO2–
CaO–P2O5 system.68 Mixtures of many of the polymers
and ceramics have been investigated in the form of
biocomposites, too numerous to list.78
The physical form of the above materials influences
cell interactions; for example, cell viability depends on
whether polymers are in the form of bulk, fibres, weaves,
matts or hydrogels. The chemistry, surface properties,
gelling conditions, stability, degradation modes and
decomposition by-products differ depending on compo-
sition, surface modifications and texture. Uncertainties
regarding collagen, widely used as an implant material
as well as a leading candidate for TE scaffolds, illustrate
the problem. The antigenic and immunogenic responses
to collagen differ depending on source (usually bovine),
clinical use (1–8% of patients are sensitive), implantation
site, type of collagen (type I versus type II), whether the
collagen is insoluble or reconstituted soluble or solubi-
lized collagen, use of treatments to remove terminal
teleopeptides (atelocollagen), extent and method of cross-
linking (gluteraldehyde, etc), if any, and presence of
antigenic noncollagenous proteins and cell-associated
components left after processing.79 Seldom are these
factors addressed in the description and characterization
of collagens used in TE papers.
A further difficulty in comparing scaffold material
performance is the lack of standard tests. Cell pheno-
types are seldom monitored throughout a scaffold testing
programme. Most tests are carried out with established
cell lines that do not represent the progenitor cells that
will be present in a clinical environment. The distribution
of cells in various stages of the cell cycle is not
documented. Often only a few markers are analysed to
determine cell viability and phenotype, and seldom are
gene array studies carried out to determine the up- or
downregulation of genes that are critical for obtaining
and maintaining cell differentiation. Few tests are
conducted such that is possible to monitor distribution
of cell types throughout a 3-D construct. The anisotropy
of natural tissues is rarely observed in scaffold tests. The
combination of multiple cell types characteristic of
tissues and organs is seldom present in tests nor is the
presence of ECM, growth factors or angiogenisis. Thus,
conclusions regarding effectiveness of the materials are
often irrelevant to eventual clinical use.
An additional difficulty is lack of control of mechan-
ical stimuli on cell–scaffold constructs. Morphogenesis
depends on both micromechanical and microchemical
gradients. The 3-D assemblages of cells within tissues
comprise an intricate 3-D cytoskeletal network composed
of three classes of biopolymers (microfilaments, inter-
mediate filaments and microfilaments) that provide a
continual tensile prestress (tensegrity).80 The cytoskeleton
transduces mechanical stimuli into a biochemical re-
sponse. Thus, ‘cells can be switched between growth,
differentiation and death solely by varying the degree to
Gene Therapy
which the cell physically distorts its shape’.81 Effects of a
simulated microgravity environment on TE constructs of
various cell types shows the importance of micromecha-
nical stimuli on cell differentiation and gene expres-
sion.82–87 Likewise, application of mechanical stimuli
during growth has a large effect on the structure and
mechanical properties of the resultant constructs.88 Until
highly controlled tests are conducted that encompass all
of the above factors, it will be impossible to deduce
which material or combination of materials is ideal for
use in TE scaffolds for a given clinical objective.
Clinical application of TE strategies needs
to meet a variety of challenges
Although tissue engineered skin substitutes and cartilage
grafts have been used with some success for several
years, most clinical applications of TE constructs need to
be considered as experimental at this time due to the
following limitations:
(1) Cell source: At present, the only reliable cell source is
autologous cells from the patient. This source has
serious limitations of numbers of cells available and
ability of the cells to maintain a phenotype capable of
generating a viable ECM. Cloned, immortal cell lines
are capable of proliferating but usually lack the
differentiation needed for stable tissue repair.
(2) Stable 3-D constructs: All tissues and organs have a
complex interdependence of cell types with an
interconnected 3-D architecture. Most tissue engi-
neered constructs involve only one, or at most two,
cell phenotypes grown primarily in a two-dimen-
sional configuration. This compromise in structure
limits the clinical viability of the constructs.
(3) Vascularization: All tissues/organs have an interpe-
netrating network of blood vessels connected to the
circulatory system to provide nutrition and eliminate
waste products. Tissue engineered constructs at
present lack this vital network when they are
transplanted. The host tissues must quickly infiltrate
the TE graft with a blood supply or the cells will die.
A major challenge of TE is to achieve angiogenesis
rapidly after implantation and maintain a viable
nutrient supply as the construct becomes integrated.
(4) Interfacial stability: The limitations of TE constructs
listed above often result in problems at the host
tissue–graft interface. Shrinkage, infiltration by new
tissue or breakdown of the interface leads to less than
desirable clinical outcomes.
(5) Sterilization: Maintaining sterility of a TE construct
containing living cells is a serious challenge in
manufacturing, handling, storage, transport and
regulation. Most methods used for sterilization of
nonliving implants and devices, such as gamma-
irradiation and autoclaving kill cells. Sterility must
be achieved during processing and maintained until
implantation is complete.
(6) Cost: All of the above factors add to the manufactur-
ing costs and presently limit many TE applications to
exploratory patients.
(7) Survivability: Long-term survivability of TE con-
structs is uncertain. Consequently, in many cases
use is restricted to applications where no other

procedure is available, as required by ethical and
legal considerations. These ‘worse-case’ surgical
scenarios make it difficult to assess viability and
success of the new procedures.
(8) Regulatory considerations: Tissue engineered products
are subject to the same regulatory procedures as
nonliving biomaterials and devices. At present, only
few products have been produced to meet these
regulatory requirements. Costs and risk/benefit
factors are often hard to predict because of the
uncertainty of regulatory approval.
New techniques have been developed that
test the efficacy of engineered tissues in vitro
Achieving and maintaining an integrated assembly of
fully differentiated, mature cell phenotypes throughout
the in vitro and postimplantation in vivo stages are major
challenges to TE. Recent developments in two fields may
help meet these objectives:
(1) New techniques have been developed for real-time
monitoring of partial pressures of O2.89 These may be
used to provide feedback data in bioreactors in the
in vitro phase of cell growth and differentiation and
be used to monitor with minimal invasion of the
metabolic state of TE constructs during their incor-
poration phase in vivo.
(2) Our groups have developed a noninvasive biopho-
tonics system for real-time in situ monitoring of cell
viability and cell death,90–94 cell phenotype,95 cell
differentiation96–98 and cellular responses to positive
and negative stimuli, including severe toxins.96–98
The bio-Raman spectroscopic cellular fingerprints
obtained by this system do not require labels or
markers and provide data on single or clustered cells
growing in 2-D or 3-D organoids. Application of this
method using fibre optic arrays to monitor TE
constructs before and after implantation may over-
come some of the clinical limitations of TE technol-
ogy listed above.
(3) Novel spectroscopic techniques have been developed
to monitor structural changes in collagen fibril
alignment in loaded tendons and are being applied
to TE constructs.99
In conclusion, the rapidly expanding fields of stem cell
research and TE are now converging to form the new
discipline of regenerative medicine, the aim of which is
to restore normal function to organs and tissues that do
not function properly as a result of disease, traumatic
injury or birth defects. Some regenerative medicine
strategies using stem cells are already in clinic, but most
are still at the bench or animal testing stages. As new
therapeutic approaches emerge, extensive, rigorous
clinical trials must be undertaken to ensure that this
very promising area of research delivers justifiable hope
and not hype.
References
1 Hench LL, Xynos ID, Polak JM. Bioactive glasses for in situ tissue
regeneration. J Biomater Sci Polym Ed 2004; 15: 543–562.
Progress and prospects in tissue engineering
J Polak and L Hench
1731
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