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Biomaterials 21 (2000) 2589}2598
Application of chitosan-based polysaccharide biomaterials
in cartilage tissue engineering: a review
J.-K. Francis Suh *, Howard W.T. Matthew
Department of Biomedical Engineering, Tulane University, Lindy Boggs Center, Suite 500, New Orleans, LA 70118, USA
Department of Chemical Engineering and Materials Science, Wyane State University, Detroit, MI, USA
Abstract
Once damaged, articular cartilage has very little capacity for spontaneous healing because of the avascular nature of the tissue.
Although many repair techniques have been proposed over the past four decades, none has sucessfully regenerated long-lasting
hyaline cartilage tissue to replace damaged cartilage. Tissue engineering approaches, such as transplantation of isolated chondrocytes,
have recently demonstrated tremendous clinical potential for regeneration of hyaline-like cartilage tissue and treatment of chondral
lesions. As such a new approach emerges, new important questions arise. One of such questions is: what kinds of biomaterials can be
used with chondrocytes to tissue-engineer aticular cartilage? The success of chondrocyte transplantation and/or the quality of
neocartilage formation strongly depend on the speci"c cell-carrier material. The present article reviews some of those biomaterials,
which have been suggested to promote chondrogenesis and to have potentials for tissue engineering of articular cartilage. A new
biomaterial, a chitosan-based polysaccharide hydrogel, is also introduced and discussed in terms of the biocompatibility with
chondrocytes. 2000 Elsevier Science Ltd. All rights reserved.
Keywords: Articular cartilage; Tissue engineering; Biomaterials; Chitosan; Polysaccharide
1. Introduction
In 1743, Hunter stated, `From Hippocrates to the
present age, it is universally allowed that ulcerated cartilage is a troublesome thing and that once destroyed, is
not repaireda [1]. Since then, a substantial amount of
research has been conducted on articular cartilage, with
signi"cant advances in our understanding of the biological repair process being made over the past four
decades. It has recently been demonstrated that articular
cartilage has a spontaneous repair response in the case of
full-thickness cartilage defects [2]. However, this repair
response is limited in terms of form and function. In
contrast, there have been no meaningful descriptions of
repair processes in partial-thickness lesions limited to the
cartilage itself [3].
While many repair techniques have been proposed
over the past four decades, none have successfully regenerated long-lasting hyaline cartilage tissue to replace
damaged cartilage [4]. In fact, most of the surgical inter-
* Corresponding author. Tel.: #1-504-865-5852; fax: #1-504-8628779.
ventions to repair damaged cartilage have been directed
toward the treatment of clinical symptoms rather than
the regeneration of hyaline cartilage, such as pain relief
and functional restoration of joint structures and articulating surfaces [5]. The concept of a tissue engineering
approach to cartilage repair was "rst proposed by Green
in 1977 [6]. In this scenario, chondrocytes grown in an ex
vivo environment would be transplanted into a cartilage
defect. Clinical application of such a tissue engineering
approach was "rst attempted by a Swedish group [7]
with fair-to-excellent clinical results in their clinical subjects. Since then, chondral repair techniques utilizing
laboratory-grown cells have attracted signi"cant attention [4].
2. Historical background in cartilage injury and repair
An initial surgical attempt to restore the normal articulating surface of joint cartilage was made with the introduction of Pridie's resurfacing technique [8,9]. This
chondral repair technique utilized the disruption of subchondral bone to induce bleeding from the bone marrow,
thus promoting the regular wound-healing mechanism in
0142-9612/00/$ - see front matter 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 1 4 2 - 9 6 1 2 ( 0 0 ) 0 0 1 2 6 - 5
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the cartilage defect site. Since Pridie's abrasion arthroplasty, several subchondral disruption techniques have
been introduced in an attempt to improve the healing
mechanisms of repaired tissue. They include subchondral
drilling [10], arthroscopic abrasion [5,11,12], and microfracture technique [13]. Several experimental animal studies on full-thickness cartilage repair have revealed that
subchondral breaching techniques created a "brin clot
formed from bleeding in the region of the cartilage defect.
This clot was subsequently in"ltrated by mesenchymal
cells, and acted as a three-dimensional sca!old for migrating progenitor cells [2]. Within six weeks, these cells gradually di!erentiated and completely "lled the defect region
with a hyaline-like repair cartilage containing signi"cantly
less proteoglycan than the normal hyaline cartilage [10].
The repair tissue at this stage was also more cellular than
the adjacent normal cartilage. Furthermore, the repair tissue showed no structural integration with residual cartilage.
As a result, there was evidence of degeneration, "ssuring in
the cartilage tissue, and even a subsequent failure to maintain
the normal appearance at a later stage [14,15]. It appears
that, in the full-thickness defect, a spontaneous repair mechanism was present for the "rst several weeks after surgical
repair, but later failed due to inadequate mechanical and
biochemical conditions in the repaired tissue [16].
3. Current concepts in cartilage tissue engineering
Recently, tissue engineering concepts have been introduced to develop cell-based repair approaches for articular cartilage [6,17,18]. Tissue engineering of articular
cartilage involves the isolation of articular chondrocytes
or their precursor cells that may be expanded in vitro and
then seeded into a biocompatible matrix, or sca!old, for
cultivation and subsequent implantation into the joint.
Certainly, the type of cell used to engineer cartilage is
critical to the long-term outcome. Di!erent cell populations
that have been investigated in the experimental studies
include matured articular chondrocytes [6,19], epiphyseal
chondrocytes [20,21], mesenchymal stem cells [22], bone
marrow stromal cells [23}25], and perichondrocytes [26].
The choice of biomaterial is also critical to the success
of such tissue engineering approaches in cartilage repair.
A variety of biomaterials, naturally occurring and synthetic, biodegradable and non-biodegradable, have been
introduced as potential cell-carrier substances for cartilage repair [27]. The naturally occurring biomaterials
include various forms of types I and II collagen-based
biomaterial, in the form sca!old matrices [28}31], gels
[22,32}34], or collagen}alginate composite gels [34].
The synthetic polymer-based biomaterials include polyglycolic acid (PGA) and poly-L-lactic acid (PLLA), or
their composite mixture [35]. In cartilage tissue engineering, PGA [18,27,36], PLLA [17,26], and PGA}PLLA
copolymers [27,37}39] have been studied for their e$cacy as chondrocyte-delivering sca!olds in vitro and in
vivo. Several investigators have also found that some
non-biodegradable polymer substances, such as polytetra#uoroethylene [40], polyethylmethacrylate [41], and
hydroxyapatite/Dacron composites [42], also facilitate
the restoration of an articular surface. However, further
studies are needed to objectively evaluate their merits in
comparison with other biomaterials.
The ideal cell-carrier substance should be the one
which most closely mimics the naturally occurring environment in the articular cartilage matrix. It has been
shown that cartilage-speci"c extracellular matrix (ECM)
components such as type II collagen and glycosaminoglycan (GAG) play a critical role in regulating expression
of the chondrocytic phenotype and in supporting chondrogenesis both in vitro and in vivo [43,44]. Otherwise,
chondrocytes may undergo de-di!erentiation and produce an inferior "brocartilaginous matrix rich in type
I collagen [45]. This inferior matrix then leads to a failure
to form hyaline cartilage [46]. Thus, we can assume that
the criteria for the choice of biomaterial in cartilage tissue
engineering include biological friendliness and biomechanical strength [27,47]. These features may provide a biochemically and biomechanically appropriate
environment necessary for engineered cells to regenerate
a long-lasting hyaline cartilage in the defect site.
4. Polysaccharide-based hydrogels
Polysaccharides form a class of materials which have
generally been underutilized in the biomaterials "eld.
Recognition of the potential utility of this class of materials is however growing and the "eld of polysaccharide
biomaterials is poised to experience rapid growth. Three
factors have speci"cally contributed to this growing recognition of polysaccharide-based biomaterials. First is
the large and growing body of information pointing to
the critical role of saccharide moieties in cell signaling
schemes and in the area of immune recognition in particular. Second has been the recent development of
powerful new synthetic techniques with the potential for
automated synthesis of biologically active oligosaccharides. These techniques may allow us to "nally decode and
exploit the language of oligosaccharide signaling. The
third factor is the explosion in tissue engineering research
and the associated need for new materials with speci"c,
controllable biological activity and biodegradability.
Apart from their biological activity, one of the more
important properties of polysaccharides in general is
their ability to form hydrogels. Hydrogel formation can
occur by a number of mechanisms and is strongly in#uenced by the types of monosaccharide involved, as well
as the presence and nature of substituent groups. Polysaccharide gel formation is generally of two types: hydrogen bonded and ionic. Hydrogen-bonded gels are typical
of molecules such as agarose (thermal gellation) and
chitosan (pH-dependent gellation), whereas ionically
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bonded gels are characteristic of alginates and carrageenans. However, the distinction is limited, since some
charged polysaccharides exhibit hydrogen-bonded gel
formation under neutral conditions.
Proteoglycans are one of the major macromolecules
found in articular cartilage. These molecules consist of
a core protein and covalently attached GAG chains. The
GAGs are long, unbranched heteropolysaccharides, consisting of repeated disaccharide units, with the general
structure: +uronic acid}amino sugar, (Fig. 1). The cartiL
lage-speci"c GAGs include chondroitin 4-sulfate (glucuronic acid and N-acetyl-galactosamine with an SO\ on
the 4-carbon position), chondroitin 6-sulfate (glucuronic
acid and N-acetyl-galactosamine with an SO\ on the
6-carbon position) and keratan sulfate (galactose and
N-acetyl-glucosamine with an SO\ on the 6-carbon posi
tion) [48,49]. The high charge density and lack of crystallinity make the sulfated GAGs highly water soluble. In
solution, these GAGs assume a #exible rod conformation, and their generally low molecular weight (5}40 kDa)
results in poor intrinsic gel formation properties. Because
of this de"ciency, the sulfated GAGs have not been
extensively studied as biomaterials for cartilage repair.
However, as described later, these molecules are capable
of forming hydrogel complexes with oppositely charged
ionic polymers, particularly the cationic polysaccharide
chitosan. This interaction may form the basis of a new
materials approach to cartilage tissue engineering.
The other important cartilage GAG is hyaluronic
acid (glucuronic acid and N-acetyl-glucosamine). This
molecule is one of the major components in synovial
#uid. Hyaluronic acid molecules are also present in cartilage matrix as the backbone structure in proteoglycan
aggregates. In general, hyaluronic acid plays a major role
as an organizer of the ECM. Furthermore, its importance
in wound healing is indicated by its transient increase
during the granulation phase [50]. Puri"ed hyaluronic
acid has been employed as a structural biomaterial because of its high molecular weight and gel forming ability.
The properties of the molecule may be broadly altered by
chemical modi"cation. For example, partial esteri"cation
of the carboxyl groups reduces the water solubility of the
polymer and increases its viscosity. Extensive esteri"cation generates materials that form water-insoluble "lms
or swellable gels [51]. Ethyl and benzyl esteri"ed hyaluronate membranes have demonstrated excellent healing responses and biodegradability in vivo. The fully
esteri"ed membranes have in vivo lifetimes of several
months, whereas the partially esteri"ed forms have been
shown to degrade within a few weeks [52].
Hyaluronic acid and derivatives have been used as
therapeutic aids in the treatment of osteoarthritis as
a means of improving lubrication of articulating surfaces
and thus reducing joint pain [53,54]. Several in vitro
culture studies have also demonstrated that hyaluronic
acid has a bene"cial e!ect by inhibiting chondrocytic
2591
chondrolysis mediated by "bronectin fragment [55]. Hyaluronic acid has also been shown to have anti-in#ammatory e!ects [56}58], as well as inhibitory e!ects on
prostaglandin synthesis [59,60], and proteoglycan release [61] and degradation [62]. Based on these "ndings,
it has been suggested that hyaluronic acid may be a good
candidate for a cell-carrier material in chondrocyte transplantation therapy [63]. Bone-marrow-derived mesenchymal cells, delivered in a hyaluronic acid hydrogel,
demonstrated good potential for cartilage resurfacing in
animal models [23,64]. However, further studies are
needed to evaluate cell}material interactions and the
mechanical characteristics of the cell}material constructs
in order to determine the e$cacy of hyaluronic-acidbased hydrogels as cell carriers for cartilage repair.
5. Chitosan as a biomaterial
Given the importance of GAGs in stimulating the
chondrogenesis [43], use of GAGs or GAG analogs as
components of a cartilage tissue sca!old appears to be
a logical approach for enhancing chondrogenesis. One
such candidate is chitosan, a partially de-acetylated derivative of chitin, found in arthropod exoskeletons. Structurally, chitosan is a linear polysaccharide consisting of
b(1P4) linked D-glucosamine residues with a variable
number of randomly located N-acetyl-glucosamine
groups. It thus shares some characteristics with various
GAGs and hyaluronic acid present in articular cartilage,
as shown in Fig. 1 [65]. Depending on the source and
preparation procedure, chitosan's average molecular
weight may range from 50 to 1000 kDa. Commercially
available preparations have degrees of deacetylation
ranging from 50 to 90%.
Chitosan is a semi-crystalline polymer and the degree
of crystallinity is a function of the degree of deacetylation.
Crystallinity is maximum for both chitin (i.e. 0%
deacetylated) and fully deacetylated (i.e. 100%) chitosan.
Minimum crystallinity is achieved at intermediate degrees of deacetylation. Because of the stable, crystalline
structure, chitosan is normally insoluble in aqueous solutions above pH 7. However, in dilute acids, the free
amino groups are protonated and the molecule becomes
fully soluble below &pH 5. The pH-dependent solubility of chitosan provides a convenient mechanism for
processing under mild conditions. Viscous solutions can
be extruded and gelled in high pH solutions or baths of
non-solvents such as methanol. Such gel "bers can be
subsequently drawn and dried to form high-strength
"bers. The polymer has been extensively studied for industrial applications based on "lm and "ber formation,
and the preparation and mechanical properties of these
forms have been reviewed previously [66}68].
Much of the potential of chitosan as a biomaterial
stems from its cationic nature and high charge density in
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Fig. 1. Molecular structures of the polysaccharide repeating units.
solution. The charge density allows chitosan to form
insoluble ionic complexes or complex coacervates with
a wide variety of water-soluble anionic polymers.
Complex formation has been documented with anionic
polysaccharides such as GAGs and alginates, as well as
synthetic polyanions such as poly(acrylic acid) [69}73].
Because the chitosan charge density is pH dependent,
transfer of these ionic complexes to physiological pH can
result in dissociation of a portion of the immobilized
polyanion. This property can be used as a technique for
local delivery of biologically active polyanions such as
the GAGs and DNA. For example, heparin released from
ionic complexes may enhance the e!ectiveness of growth
factors released by in#ammatory cells in the vicinity of an
implant [74]. In the case of DNA, complexation with
chitosan has been shown to protect plasmids from degradation by nucleases and also facilitates cellular transfection by poorly understood interactions with cell
membranes [75}77].
The N-acetylglucosamine moiety in chitosan is a structural feature also found in the GAGs. Since GAG properties include many speci"c interactions with growth
factors, receptors and adhesion proteins, this suggests
that the analogous structure in chitosan may also have
related bioactivities. In fact, chitosan oligosaccharides
have been shown to have a stimulatory e!ect on macrophages, and the e!ect has been linked to the acetylated
residues [78]. Furthermore, both chitosan and its parent
molecule, chitin, have been shown to exert chemoattractive e!ects on neutrophils in vitro and in vivo [79}81].
In vivo, chitosan is degraded by enzymatic hydrolysis.
The primary agent is lysozyme, which appears to target
acetylated residues [82]. However, there is some evidence
that some proteolytic enzymes show low levels of activity
with chitosan. The degradation products are chitosan
oligosaccharides of variable length. The degradation kinetics appear to be inversely related to the degree of
crystallinity which is controlled mainly by the degree of
deacetylation. Highly deacetylated forms (e.g. '85%)
exhibit the lowest degradation rates and may last several
months in vivo, whereas samples with lower levels of
deacetylation degrade more rapidly. This issue has been
addressed by derivatizing the molecule with side chains
of various types [83}86]. Such treatments alter molecular
chain packing and increase the amorphous fraction, thus
allowing more rapid degradation. They also inherently
a!ect both the mechanical and solubility properties.
A number of researchers have examined the tissue
response
to
various
chitosan-based
implants
[65,73,83,87}96]. In general, these materials have been
found to evoke a minimal foreign body reaction. In most
cases, no major "brous encapsulation has been observed.
Formation of normal granulation tissue, often with accelerated angiogenesis appears to be the typical course of
healing. In the short term ((7 d), a signi"cant accumulation of neutrophils in the vicinity of the implants is often
seen, but this dissipates rapidly and a chronic in#ammatory response does not develop. The stimulatory e!ects of
chitosan and chitosan fragments on immune cells mentioned above may play a role in inducing local cell
proliferation and ultimately integration of the implanted
material with the host tissue.
One of chitosan's most promising features is its excellent ability to be processed into porous structures for use
in cell transplantation and tissue regeneration. Porous
chitosan structures can be formed by freezing and
lyophilizing chitosan}acetic acid solutions in suitable
molds [97]. During the freezing process, ice crystals
nucleate from solution and grow along the lines of thermal gradients. Exclusion of the chitosan acetate salt from
the ice crystal phase and subsequent ice removal by
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J.K. Francis Suh, H.W.T. Matthew / Biomaterials 21 (2000) 2589}2598
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Fig. 2. SEM images of various forms of porous chitosan sca!olds.
lyophilization generates a porous material whose mean
pore size can be controlled by varying the freezing rate
and hence the ice crystal size (Fig. 2). Pore orientation
can be directed by controlling the geometry of thermal
gradients during freezing.
The mechanical properties of chitosan sca!olds are
mainly dependent on the pore sizes and pore orientations. Tensile testing of hydrated samples shows that
porous membranes have greatly reduced elastic moduli
(0.1}0.5 MPa) compared to non-porous chitosan membranes (5}7 MPa). The extensibility (maximum strain) of
porous membranes varied from values similar to nonporous chitosan (&30%) to greater than 100% as
a function of both pore size and orientation. The highest
extensibility was obtained with a random pore orientation structure frozen rapidly at !783C to give pores
120 lm in diameter. Porous membranes exhibited
a stress}strain curve typical of composite materials with
two distinct regions: a low-modulus region at low strains
and a transition to a 2}3-fold higher modulus at high
strains. The tensile strengths of these porous structures
were in the range of 30}60 kPa.
Chemical derivatization of chitosan provides a powerful means to promote new biological activities and to
modify its mechanical properties. The primary amino
groups on the molecule are reactive and provide a mechanism for side group attachment using a variety of mild
reaction conditions. The general e!ect of addition of
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a side chain is to disrupt the crystal structure of the
material and hence increase the amorphous fraction. This
modi"cation generates a material with lower sti!ness and
often altered solubility. Of course, the precise nature of
changes in chemical and biological properties depends
on the nature of the side group. For example, N-alkyl
derivatives can exhibit reduced solubility and micellar
aggregation in solution for alkyl lengths greater than "ve
carbons. In addition, the characteristic features of
chitosan, such as cationic, hemostatic and insoluble at
high pH, can be completely reversed by a sulfation process which can render the molecule anionic and water
soluble, and also introduce anticoagulant properties. The
variety of groups which can be attached to chitosan is
almost unlimited, and side groups can be chosen to
provide speci"c functionality, alter biological properties,
or modify physical properties.
Fig. 3. (Light micrograph, 200;) After one week, primary chondrocytes cultured on the CSA-chitosan surface maintained a spherical
morphology and demonstrated a limited rate of mitosis. A granular
surface of CSA-chitosan material is shown in the background (from
Sechriest et al. [101].
6. Chitosan}GAG composite material and its biological
interaction with articular chondrocytes
Use of chitosan as a biomaterial or drug delivery agent
has recently drawn a considerable attention in the applications for the repair of articular cartilage. Lu et al.
[98] has demonstrated that the chitosan solution injected
into the knee articular cavity of rats caused a signi"cant
increase in the density of chondrocyte in the knee articular cartilage, suggesting that the chitosan could be
potentially bene"cial to the wound healing of articular
cartilage. Mattioli-Belmonte [99] has shown that the
bone morphongenetic protein (BMP)-7, associated with
N,N-dicarboxymethyl chitosan induces or facilitates the
repair of articular cartilage lesions.
As mentioned above, chitosan is capable of forming
insoluble ionic complexes with the negatively charged
GAGs. This ionic cross-linking mechanism can be used
to immobilize chondroitin sulfates within hydrogel materials which mimic the GAG-rich ECM of the articular
chondrocyte. A recent study by Denuziere [73] has demonstrated that, when associated with various polyelectrolytic GAGs, the chitosan has a protective e!ect
against GAGs hydrolysis by their speci"c enzymes. Recent preliminary studies performed in our laboratory
have demonstrated the biocompatibility and the chondrogenic characteristics of such GAG-augmented
chitosan hydrogels [100}102]. When chondrocytes were
cultured on the chondroitin 4-sulfate (CSA)-augmented
chitosan hydrogel surfaces, the cells maintained many
morphological and functional features characteristic of
chondrocytes in normal cartilage. These promising results are brie#y summarized below.
6.1. Cell growth kinetics
Chondrocytes seeded at low density (6;10 cells/ml)
onto either the standard polystyrene culture plates or the
CSA-chitosan membrane surfaces attached to the substrate within 2 h. Over the course of one week, the majority of cells attached to the polystyrene surface assumed
a "broblast-like morphology, and the cell population
rapidly expanded. In contrast, primary chondrocytes
seeded at the same low density onto the CSA-chitosan
surface assumed a spherical morphology after attachment. After one week, chondrocytes attached to the
CSA-chitosan had largely maintained a rounded or polygonal morphology and undergone only a modest degree of
mitosis (Fig. 3). The increase in cell population of chondrocytes cultured on the polystyrene and CSA-chitosan
surfaces were 313$61 and 66$28%, respectively.
6.2. Scanning electron microscopic morphology
Scanning electron microscopic evaluation of the CSAchitosan membrane revealed a matrix material with an
irregular surface. We found no evidence that chondrocytes were embedded or partially embedded in the
hydrogel membrane. Rather, the chondrocytes adherent
to the CSA-chitosan had formed discrete points of attachment with the biomaterial through pseudopodial
extensions (Fig. 4). These microscopic "ndings suggest
that maintenance of the phenotypic morphology is due to
an interaction between cell-membrane receptors and the
CSA-rich ECM. A similar phenomenon was previously
described with primary murine articular chondrocytes
cultured on a collagen type II membrane [103].
6.3. Proteoglycan synthesis
The total S-sulfate-incorporation rate produced by
primary chondrocytes cultured on the CSA-chitosan surface (2580$460 cpm/10 cells) was approximately equal
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J.K. Francis Suh, H.W.T. Matthew / Biomaterials 21 (2000) 2589}2598
Fig. 4. SEM image of primary articular chondrocyte adherent to the
CSA-chitosan surface. The heterogeneous surface characteristics of the
biomaterial and the round morphology of the chondrocytes are shown
with discrete attachment points between the cell and the material.
to that produced by chondrocytes cultured on the standard polystyrene surface (2810$320 cpm/10 cells).
Thus, CSA-chitosan supports, but does not appear to
enhance the total amount of PG synthesized by primary
articular chondrocytes.
6.4. Collagen synthesis
After one week, chondrocytes cultured on the control
polystyrene surface (P) demonstrated a complex collagen
phenotype. The protein migration pattern shown in
Fig. 5 indicates that the collagens produced by cells in
this culture system were types I and II, type I collagen
being the predominant collagen synthesized. The presence of type I collagen is indicated by the presence of
a2(I) chains. Also present in smaller amounts were protein bands consistent with a1(V)/a1(XI) and a2(XI)
chains. Although the high molecular weight protein
bands were not identi"ed, they may represent chains of
type III collagen, type IX collagen, and/or b-chains of
type I collagen.
For chondrocytes cultured on the CSA-chitosan, on the
other hand, the predominant collagen produced by chondrocytes was type II, indicated by the presence of the
strongly stained protein band in the a1(II) position with
only faint protein bands in the a2(I) position. Chondrocytes in this culture system also demonstrated protein
bands consistent with a1(V)/a1(XI) and a2(XI) chains. The
high molecular weight protein bands in the CSA-chitosan
group were not identi"ed, but may represent pepsin-derived fragments of type IX collagen. These data indicate
that the primary chondrocytes cultured on CSA-chitosan
maintain the synthesis of cartilage-speci"c collagens.
2595
Fig. 5. SDS-PAGE analysis for collagen typing. The analysis shows
that primary articular chondrocytes cultured on polystyrene (P) produced types I and II collagens, whereas chondrocytes cultured on the
CSA-chitosan (C) predominantly produced type II collagen (from Sechriest et al. [101]).
7. Future direction and discussion
Despite extensive research on articular cartilage conducted over the past four decades, we still do not have the
de"nitive answer for a successful repair of damaged cartilage. While many new repair techniques have recently
emerged and demonstrated promising results in experimental models, few have exhibited long-term clinical
e$cacy.
Tissue engineering has recently emerged as a new interdisciplinary science to repair injured body parts and
restore their functions by using laboratory-grown tissues,
materials and arti"cial implants. It is our hope that tissue
engineering approaches with novel materials will provide
methods for developing a greater understanding of articular cartilage and associated pathologies. This understanding of the tissue pathogenesis will help us develop
new methods to enhance the repair of damaged cartilage.
The biochemical and morphologic results of this study
of primary chondrocytes cultured on CSA-chitosan
re-emphasize the favorable in#uence of articular cartilage-speci"c matrix components on the chondrocytic
phenotype in vitro. Our "ndings, coupled with the favorable biological properties of chitosan, suggest that CSAchitosan may be well suitable as a carrier-material for the
transplant of autologous chondrocytes as described by
Brittberg et al. [7] and/or as a sca!old for the tissue
engineering of cartilage-like tissue as originally described
by Vacanti et al. [104]. Future studies using chondrocytes that have undergone proliferative expansion in
vitro will explore this potential. It is also very important
that the chitosan sca!old possesses necessary mechanical
properties in order to provide seeded chondrocytes with
a proper biomechanical environment [105]. Further
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studies are necessary to investigate the material and
mechanical properties (such as porosity, compressive
elastic modulus, permeability, and viscoelastic properties) of the CSA-chitosan sca!old. A development of
a chitosan-based material that can support chondrogenesis may be signi"cant not only in terms of the quality of
neocartilage produced, but also in terms of the ability of
that tissue to integrate with the host matrix.
Along with tissue engineering as a new emerging
science for various tissue repairs, gene therapy has also
been suggested as a means of delivering sustained therapeutic levels of anti-arthritic gene products to the transplanted cells in diseased joints [58,106,107]. Genes have
been successfully introduced locally to the target cells in
the knee joints of rabbits by both ex vivo and in vivo
methods, with gene expression lasting for several weeks
[108,109]. Such gene therapy has been successfully combined with chondrocyte transplantation to investigate
the feasibility of ex vivo gene transfer to chondrocytes in
full-thickness cartilage defects [32]. Combination of
chitosan-based cartilage sca!olds with gene therapy and
the use of growth factors will hopefully improve the
long-term outcomes of cartilage repair in clinical settings.
Acknowledgements
The authors wish to acknowledge support from the
University of Pittsburgh Medical Center (JKS),
the Arthroscopic Association of North America (JKS),
the Pittsburgh Tissue Engineering Initiative (JKS), and the
National Science Foundation (HWTM: BES-9624151).
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