This is an open access article published under an ACS AuthorChoice License, which permits

copying and redistribution of the article or any adaptations for non-commercial purposes.

pubs.acs.org/journal/abseba Perspective

Perspectives on Synthetic Materials to Guide Tissue Regeneration

for Osteochondral Defect Repair
Michael T. Frassica and Melissa A. Grunlan*

Cite This: ACS Biomater. Sci. Eng. 2020, 6, 4324−4336 Read Online

ACCESS Metrics & More Article Recommendations

See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

ABSTRACT: Regenerative engineering holds the potential to treat

clinically pervasive osteochondral defects (OCDs). In a synthetic
materials-guided approach, the scaffold’s chemical and physical
Downloaded via 223.187.112.203 on September 17, 2024 at 07:39:06 (UTC).

properties alone instruct cellular behavior in order to effect

regeneration, referred to herein as “instructive” properties. While
this alleviates the costs and off-target risks associated with exogenous
growth factors, the scaffold must be potently instructive to achieve
tissue growth. Moreover, toward achieving functionality, such a
scaffold should also recapitulate the spatial complexity of the
osteochondral tissues. Thus, in addition to the regeneration of the
articular cartilage and underlying cancellous bone, the complex
osteochondral interface, composed of calcified cartilage and
subchondral bone, should also be restored. In this Perspective, we
highlight recent synthetic-based, instructive osteochondral scaffolds that have leveraged new material chemistries as well as
innovative fabrication strategies. In particular, scaffolds with spatially complex chemical and morphological features have been
prepared with electrospinning, solvent-casting−particulate-leaching, freeze-drying, and additive manufacturing. While few synthetic
scaffolds have advanced to clinical studies to treat OCDs, these recent efforts point to the promising use of the chemical and physical
properties of synthetic materials for regeneration of osteochondral tissues.
KEYWORDS: osteochondral defect, subchondral, synthetic, materials-guided, tissue engineering

■ INTRODUCTION
Osteochondral defects (OCDs) are injuries to the cartilage of
the knee or ankle that extend from the articular cartilage into
the underlying subchondral bone (Figure 1).1 These defects,
occurring from traumatic lesions or osteochondritis dissecans,
cause severe pain and diminished mobility.2−4 Due to the
limited healing capacity of this interface,5 defects can progress
to osteoarthritis (OA) and necessitate total joint replace-
ment.6,7 As the depth of the defect often extends beyond that
of partial or full chondral defects (Figure 1), limited success is Figure 1. Schematic representation of tissue composition along the
osteochondral interface. Defect depth is denoted (left) as partial
observed for procedures such as microfracture and autologous chondral defect (1), full chondral defect (2), and osteochondral
chondrocyte implantation (ACI) due to the inability to defect (3). Cartilage layers are denoted (upper right) with
regenerate subchondral bone and articular cartilage.8,9 Thus, approximate percentage of the cartilage tissue each zone comprises.
grafting is the most common clinical option to treat OCDs, Cellularity enhances with each layer of cartilage, eventually forming a
with cylindrical autografts utilized for smaller defects (diameter transition of calcified cartilage connected to the subchondral bone
plate. Interface components are labeled (lower right) with
<3 cm2) and allografts for larger defects (diameter >3 approximate sizes.
cm2).10,11 However, both strategies suffer from a lack of graft
availability and delamination from the surrounding tissue, Received: May 19, 2020
along with risk of donor site morbidity for autografts and Accepted: July 1, 2020
disease transmission/rejection for allografts.11−13 Thus, re- Published: July 1, 2020
generative engineering has emerged as a promising option to
heal OCDs.14,15

© 2020 American Chemical Society https://dx.doi.org/10.1021/acsbiomaterials.0c00753

4324 ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

In general, regenerative engineering aims to provide a between the calcified cartilage and subchondral bone plate as
temporary three-dimensional (3D) environment or “scaffold”, well as the zones of cartilage.
in lieu of the native extracellular matrix (ECM), for recruited A broad range of biomaterials, both biologic and synthetic,
or seeded cells to proliferate and regenerate tissue.16,17 have been utilized for OCD repair based on their success in
Mesenchymal stem cells (MSCs), able to differentiate toward orthopedic and other biomedical applications, as described in
multiple cell lineages, are most widely used due to their ease of Di Luca et al., Longley et al., and Deng et al.20,21,63 Among
isolation from numerous adult tissues and their manipu- these materials, the most commonly utilized are approaches
lability.18,19 Exogenous growth factors, or cellular signaling based on proteins such as collagen and silk fibroin for their
proteins, are commonly included in scaffolds to stimulate adhesive and regenerative capacities, as recently reviewed in
regeneration of tissues, including cartilage (e.g., insulin-like the literature.20,21,63 However, because of the complex nature
growth factor-1 [IGF-1] or transforming growth factor β-1 of cell−protein interactions, we have chosen to focus on the
[TGFβ-1]) and bone (e.g., bone morphogenetic protein-2 current state of synthetic-based scaffold strategies that do not
[BMP-2] or vascular endothelial growth factor [VEGF]).20 include a protein component. This highlights the potency of
Achieving sustained, long-term release of growth factors from scaffold material properties to independently drive regener-
scaffolds during their degradation requires high loading levels ation. Synthetic materials are not without their drawbacks. For
which is associated with high costs and off-target tissue instance, bioresorbable polymers, such as poly(α-hydroxy
responses.21−23 For osteochondral tissue, the demand for acids), provide potential for adverse effects to cellular
spatiotemporal delivery of various growth factors represents an regeneration from acidic degradation products. Attempts to
additional level of complexity.24 In contrast, a material-based mitigate such effects in vivo have included reduction of implant
regenerative approach relies exclusively on scaffold chemical size and inclusion of additives.64,65 In the case of cartilage
and physical material properties to guide tissue regeneration regeneration, synthetic materials are primarily combined with
(i.e., without exogenous growth factors).25−29 As with the polysaccharides based on their unique mechanical properties as
well as chemistries that enhance chondrogenesis; therefore,
ECM of native tissue, numerous scaffold properties have been
these systems are included. Thus, within this Perspective, we
shown to direct cell behavior, including morphology (e.g., pore
provide an outlook on the recent utility of the chemical and
size and shape),30−34 modulus,35−39 chemical functional-
physical properties of synthetic scaffold systems for OCD
ity,40−43 hydrophobicity,44−46 and hydration.47−49 Moreover,
repair, with a focus on highlighting common trends and
high porosity and interconnectivity are considered beneficial potential areas for future investigation.

■
for nutrient diffusion and neotissue infiltration.50 Such
scaffolds that leverage material properties to guide tissue CONSIDERATIONS FOR
regeneration in these ways are referred to as “instruc- OSTEOCHONDRAL-INSTRUCTIVE SCAFFOLDS
tive”,27,51,52 and will be mentioned as such herein.
A materials-guided approach to treat OCDs represents a Synthetic polymer scaffolds have been widely studied for
particular challenge due to the spatial complexity of tissues decades in the field of tissue engineering. The level of control
spanning this region (Figure 1). Osteochondral tissue consists over key chemical and physical properties (e.g., degradation
of osseous and chondral tissues anchored together by a rate, pore morphology, chemical functionality, modulus, etc.)
complex layer of calcified cartilage, beneath the tidemark and allowed by variations in chemistry and fabrication provides for
an expansive scaffold library to apply to various tissue
abutting the subchondral bone plate,53,54 a feature that is key
applications. Below, we highlight key criteria and materials
to its ability to disperse normal load.55 The bone tissue is a
for synthetic scaffolds useful for the regeneration of bone,
highly vascularized, porous scaffolding of organic collagen
cartilage, and the osteochondral interface.
mineralized with inorganic hydroxyapatite (HAp), with a water
Materials-Driven Osseous Tissue Regeneration. Bone
content of ∼20−30%. The cartilage tissue is a dense ECM of tissue engineering has been extensively studied, as recently
primarily collagen and glycosaminoglycans (GAGs) but with a reviewed by Du et al.66 and Yu et al.67 For osseous tissue
much higher water content (∼75%) and a more sparse regeneration (i.e., osteogenesis), instructive scaffolds require
distribution of native cells.5,56,57 Furthermore, articular osteoconductivity, osteoinductivity, and osseointegration66−69
cartilage consists of three layers of different collagen fiber which may be achieved by their chemical and physical
orientation and cellularity (Figure 1), resulting in remarkable properties. Osteoconductivity is traditionally defined as a
biomechanical stability. In recent years, damage to this material’s ability to encourage and support new bone
interface has been identified as a major contributor to cartilage formation on its surface. This has been identified to be
defects and OCD propagation due to its key role in supporting facilitated by bioactivity (i.e., formation of a thin carbonated
the intensive loading of the joint.8,55,58 Leveraging the prior art HAp layer at the material surface) that adsorbs proteins for
on materials-guided scaffold properties associated with bone cellular attachment and proliferation.70 Scaffold chemical
and cartilage regeneration, instructive osteochondral scaffolds composition (e.g., ion releasing71 and phosphonate-contain-
have largely adopted designs with multiple phases and are ing72) and pore architecture (interconnected pore size of 200−
often formed as cylindrical autograft-sized plugs.59 Biphasic 500 μm73,74 or larger75) are associated with this trait, providing
designs have been most extensively used, presenting two growth of the mineral layer and space for neotissue infiltration.
distinct environments for differentiation of multipotent cells Osteoinductivity is broadly understood as the ability to induce
toward an osteoblastic or chondrocytic lineage. However, the bone formation via cellular differentiation to a bone-forming
simple merging of two discrete scaffolds (e.g., by mechanical lineage. Mechanisms for this action have been widely studied,
attachment, glue, or freeze-drying) creates a hard interface and including active facilitation of nutrient and waste transport via
so fails to mimic the nativelike complexity of the osteochondral vascularization,67,76 adherence of progenitor cells on the
interface.60−62 Thus, recent designs have adopted triphasic, material surface (via protein adsorption or direct integrin
multiphasic, or gradient designs to recapitulate the transition binding) and subsequent interaction with surface proper-
4325 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

ties,68,77−79 osteogenic response to calcium and phosphate drointegration. Chondroconductive scaffolds provide a mor-
ions,80−82 and concentration of osteogenic proteins by material phology similar to that of the confined chondrocytic
affinity.67,83,84 Osteoinductivity is often affiliated with a certain environment (smaller interconnected pores (<300 μm)113
chemical composition (e.g., hydrophobic and inorganic with low porosity114) to facilitate cell−cell interactions for
materials such as bioglasses, calcium phosphates, siloxanes, chondrogenesis. Chondroinduction (i.e., induced differentia-
and nanosilicates)68,85−91 and pore topography (e.g., meso- tion toward a chondrocytic lineage) is strongly related to the
pores,92,93 rough surfaces,94 and aligned fibers66). Matrix level of cellular adhesion wherein moderate adherence of
stiffness is also identified as a key factor in osteogenesis, as MSCs to a material surface is preferred and promotes a
adhered cells can respond to the local environment through a spherical cell morphology resembling that of a chondrocyte.115
variety of mechanisms.95,96 Generally, stiffer matrices have Other factors can induce this morphological change as well,
been identified as osteogenic; however, other simultaneous including encapsulation and scaffold morphology.9,116 Mecha-
cell−material interactions add complexity to a strict correlative notransduction has also been indicated in chondroinductiv-
relationship. To this point, in comparative studies, osteogenic ity117,118 and is part of a complex relationship between cell
moduli have been seen with hydrogels of 11−40 kPa36,97 or shape, matrix properties, and mechanical loading. Finally, for
>225 kPa.98 In other studies, osteogenesis has been observed chondrointegration, vertical integration of regenerated cartilage
for nonhydrogel matrices having moduli several magnitudes with underlying bone has been observed; however, lateral
higher.74,99 Finally, osseointegration refers to the ability for the integration with native cartilage tissue is more challenging.
material to integrate with the surrounding bone. In general, This can be attributed to factors such as biomechanical
this requires scaffold bioactivity for immediate bone bonding, mismatching, cell death at the defect edge, and the
an interconnected pore architecture for infiltration of neotissue antiadhesive nature of the tissue itself.119 In many cases,
and vascularization, and biodegradability that supports the bioadhesives are thus employed.120 Another key factor for
bone regeneration.74 Ideally, scaffold degradation should chondrointegration is biodegradation, where a degradation rate
parallel neotissue formation.100 that matches the rate of tissue growth and can maintain
Calcium phosphates (including HAp), resembling the mechanical support is a necessity.
inorganic phase of native bone tissue, have been the most A variety of synthetic polymers (e.g., poly(ε-caprolactone)
abundantly utilized material for bone regeneration. Such [PCL], poly(L-lactic acid) [PLLA], poly(lactide-co-glycolide)
materials are known to recruit and stimulate MSCs while [PLGA], and poly(ethylene glycol) [PEG]), sometimes in
simultaneously initiating angiogenic and chemotactic re- combination with polysaccharides (e.g., cellulose, agarose, and
sponses.71 For regenerative scaffolds, these have primarily alginate), have been explored for materials-guided cartilage
been formed into composites with polymers, typically regeneration.9,111 Due to their hydration, permeability, and
polyesters, for better control over pore morphology and associated lubricity, hydrogels have been most frequently used
toughness.101,102 More recently, bioglasses and bioceramics as materials for cartilage regeneration. PEG-DA hydrogels have
have seen use after modifications via element-doping (e.g., F−, been widely studied because of the ease of tuning cross-link
Cl−, Na+, and Mg2+)20,66 to mitigate their brittleness, to slow density, and in turn the modulus, thereby impacting cellular
their resorption rates,101,103−105 and to advance osteogenic differentiation toward chondrocytic cell types.28,48 However,
differentiation or suppress inflammation.106,107 Our group has such PEG scaffolds are limited in their ability to promote
demonstrated that bioactivity and osteoinductivity can also be sufficient ECM production, which is associated with their lack
achieved with nonbrittle materials by the incorporation of a of chemical and biological mimicry of native cartilage ECM.
silicon-based, inorganic polymer [star-polydimethylsiloxane Thus, synthetic polymers have been combined with natural
methacrylate] into poly(ethylene glycol) diacrylate (PEG- polymers (e.g., hyaluronic acid [HA], chondroitin sulfate [CS],
DA) hydrogels.88,89 The instructive behavior of this silicon- and chitosan), due to the presence of charged groups like those
based polymer is attributed to the known role that silicon plays comprising GAGs found in cartilage ECM, to form instructive
in bone mineralization and gene activation108,109 which had scaffolds.9,121,122 HA and CS have further been indicated in the
originally prompted the use of certain glasses, ceramics, and binding of the cell adhesion peptide CD44, assisting in the
nanosilicates. As described later, fabrication strategies have support of cell−cell interactions and subsequent chondro-
been utilized to maximize the potential of these materials by genesis.123,124 More recently, fibrous materials have been used
controlling morphological features of the scaffold. for articular cartilage regeneration, particularly those based on
Materials-Driven Chondral Tissue Regeneration. biodegradable polyesters such as poly(glycolic acid) [PGA],
Articular cartilage regeneration presents a significant challenge PLLA, and PCL.9,21,125 These scaffolds’ structure mimics the
in tissue engineering, as reviewed by Armiento et al.,9 with fibrous collagen of native cartilage, providing a morphologically
considerably less success compared to bone regeneration. similar environment and a compressive stiffness and strength
Cartilage is a highly hydrated, avascular tissue that displays more similar to that of cartilage. The aligned fibrous
remarkably complex biomechanical behavior characterized by morphology of these scaffolds can support a chondrocytic
high compressive stiffness, strength, and lubricity.5,110 Such phenotype,125 as previously mentioned.
complexity can be attributed to its layered nature (Figure 1), Materials-Driven Osteochondral Interface Regener-
wherein horizontally aligned collagen at the surface progresses ation. Beyond the “deep zone” of articular cartilage lies the
to vertically aligned deeper in the tissue, resulting in the osteochondral interface, consisting of calcified cartilage and the
tissue’s anisotropic feature. Furthermore, it has a remarkable subchondral bone plate, leading subsequently to cancellous
ability to expel and regain its water content (up to 80% by bone (Figure 1). Originally, instructive scaffolds regarded this
weight) during loading and articulation.111,112 Toward area as a simple interface between two unique tissues (i.e.,
materials-driven regeneration of cartilage, synthetic materials bone and cartilage). Thus, biphasic osteochondral scaffolds
should recapitulate an environment that drives chondrogenesis were formed by merging two independent scaffolds across a
through chondroconductivity, chondroinductivity, and chon- hard interface.60−62 More recently, this interface has been
4326 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

recognized for its role in the pathophysiology of OCDs and

eventual OA. Specifically, a healthy subchondral bone adapts
to the load of locomotion dispersed by the calcified cartilage
layer.8,126,127 In osteochondral tissue formation, subchondral
bone acts as a physical barrier that maintains the integrity of
neocartilage and prevents bone in-growth, thereby directly
relating it to the successful regeneration of articular
cartilage.8,58 Thus, this interface has been addressed in
osteochondral scaffolds by the incorporation of additional
layer(s) or “phase(s)” with differing material properties. For
triphasic scaffolds, the middle phase is treated as a discrete
layer with intermediate properties between bone and
cartilage.128,129 Often, “middle-ground” levels (between bone
Figure 2. Tru-Fit CB (left, A) is a biphasic PLGA copolymeric
and cartilage) of scaffold porosity,130−132 modulus,9,35,97,133
scaffold with calcium sulfate (bone phase) and PGA fibers (chondral
and material composition21,63 are used to create a transition. phase). The Agili-C (right, B) is a marine-derived aragonite (bone
Successful in vitro indications often show chondrocytic phase) scaffold with a hyaluronic acid-containing top (cartilage
proliferation in a mineralized environment and expression phase). Reproduced with permission from ref 148. Copyright 2014
phenotypical of hypertrophic chondrocytes, similar to that of Springer Berlin Heidelberg.
the calcified cartilage layer.134,135 Examples of these strategies
include diverse composites (e.g., agarose with HAp, organic sol clinical trials for osteochondral repair after initially being
gel-coated calcium polyphosphate),136,137 scaffold component utilized for bone voids; however, it has since become
orientations (e.g., arrayed platelike HAp),138 additives for unavailable.152 While preclinical in vivo studies showed
osteochondral differentiation (e.g., Mg-doped wollastonite, promise, this TruFit suffered from poor integration with
icariin, or zinc oxide),139−141 or components that promote a adjacent tissue as well as a lack of sufficient subchondral bone
proregenerative environment (e.g., strontium or copper).142,143 growth in short- and long-term studies.153−155

■ MATERIALS-GUIDED TISSUE ENGINEERING

APPROACHESCLINICAL TRIALS
More recently, the Agili-C was developed as an acellular
biphasic instructive scaffold composed of coral-derived
aragonite at the bone phase and aragonite/hyaluronic acid at
In the treatment of OCDs, nondegradable partial resurfacing the cartilage phase (Figure 2B).148 Aragonite is a calcium
devices and materials-driven regenerative devices have carbonate with ∼2% sodium, magnesium, potassium, and other
emerged in clinical trials. For partial resurfacing devices, a ions absorbed from the seawater during its formation.156
synthetic material permanently replaces damaged tissues. For Having interconnected pores ranging from 100 to 300 μm
example, Cartiva, a cryogenically fabricated poly(vinyl alcohol) (porosity >50%), this material has recently been shown to
(PVA) hydrogel, is an FDA approved therapeutic for the support osteogenic differentiation of bone-marrow-derived
treatment of OCDs in the first metatarsal phalangeal joint.144 MSCs.156,157 In addition to establishing the devices’ bio-
Biopoly, currently in clinical trials, utilizes a titanium bone compatibility and biodegradability, early studies of the device
anchor and an ultra-high-molecular-weight poly(ethylene) demonstrated its ability to recruit stem cells from the
(UHMWPE)/HA hybrid surface for treatment of osteochon- surrounding tissue, attributed to the complex, nanorough
dral defects in the knee.145 Several layered materials-guided geometry of the aragonite and CD44 binding capacity of
OCD regenerative devices are in clinical trials, including hyaluronic acid.123,158 More recent studies have shown
protein-based devices such as BioMatrix CRD (Arthrex, US), restoration of high-quality cartilage and subchondral bone in
Cartilage Repair Device (Kensey Nash Corp.), ChondroMi- a 12 month caprine model.158 This biphasic device began a
metic (Collagen Solutions, UK), and MaioRegen (Finceramic, 250+ participant premarket approval (PMA) pivotal study in
Italy).146,147 However, only two OCD devices that use a 2017 for treatment of joint surface lesions.159 Results of this
synthetic materials-guided regenerative strategy without the study will be informative regarding the regenerative potential
incorporation of proteins and/or exogenous growth factors of a biphasic scaffold to treat OCDs.
have undergone clinical trials. These two devices include the Recent Materials-Guided Approaches. A variety of
TruFit (Smith & Nephew, Andover, MA) and Agili-C instructive scaffold designs have emerged in recent years that
(CartiHeal Ltd., Israel). target healing of OCDs using a synthetic materials-guided
The TruFit Plug, an acellular biphasic scaffold, consists of a regenerative engineering approach (Table 1). Given the known
semiporous 75:25 poly(D,L-lactide-co-glycolide) (PLGA) co- influence of morphological features on directing cell
polymer with calcium sulfate incorporated in the lower bone behavior,30−34 scaffolds with spatially controlled morphology
phase and Poly(glycolic acid) (PGA) fibers in the upper and chemistry are expected to improve regeneration,
cartilage phase (Figure 2A).148,149 The two phases are merged particularly in combination with materials whose chemical
together using a small amount of solvent. Bone regeneration and physical properties are known to be instructive. In this
relies upon the osteoinductive and resorptive capacity of the section, we highlight recent examples of osteochondral
calcium sulfate.150,151 The limited osteoconductivity (i.e., scaffolds that have leveraged different fabrication strategies
limited porosity) and brittleness associated with calcium (Figure 3).
sulfate are mitigated by fabricating as a composite with Electrospinning. Electrospinning involves charging and
biodegradable PLGA. At the cartilage phase, PGA fibers are drawing a viscous polymer solution into fibers at the micro- or
incorporated with the PLGA copolymer scaffold to provide a nanometer scale onto a collector of opposite polarity.175
cartilage-mimetic fibrous environment for recruited progenitor Deposition of these fibers results in a nonwoven fibrous mesh
cells from the native tissue. The TruFit Plug advanced to with tunable porosity, interconnectivity, mechanical properties,
4327 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

Table 1. Select Recent Synthetic Materials-Guided Strategies for Osteochondral Tissue Engineeringa
no. of
phases bone cartilage interface fabrication year ref
7 PCL/30% HAp PCL PCL/(25/20/15/10/5%) SLS 2017 160
HAp
2 PECDA, PAAm, PEG-DA CSMA, PNIPAAm calcium gluconate and phase separation; thermal cross- 2017 161
alginate linking
3 PCL PCL N/A electrospinning; FDM 2017 162
2 methacrylated triblock methacrylated triblock N/A FDM 2017 163
(F127−P123)/CaP (F127−P123)
2 PVA/HAp PVA N/A Electrophoresis; freeze-drying 2017 164
2 PVA/β-TCP PVA N/A SCPL; freeze-drying 2017 165
3 PCL/30% HAp PCL PCL/15% HAp FDM 2018 166
2 PCL/10% graphene N/A N/A SCPL 2018 167
2 alginate/HAp alginate/hyaluronic acid N/A diffusion controlled, directed 2018 168
ionotropic gelation
3 PEG-DA/A6ACA/CaP PEG-DA PEG-DA freeze-drying 2018 169
2 chitosan/alginate/HAp chitosan/hyaluronic acid N/A TIPS; freeze-drying 2019 170
2 PLGA/tyramine PLGA/tyramine N/A SCPL; freeze-drying 2019 171
3 silicon−zirconium SZ2080 silicon−zirconium SZ2080 silicon−zirconium SZ2080 stereolithography 2019 172
2 PDLLA/camphene PDLLA N/A SCPL; freeze-drying 2019 173
2 β-TCP PEG-DA N/A stereolithography 2019 174
a
Abbreviations include the following: β-TCP = β-tricalcium phosphate, A6ACA = N-acryloyl 6-aminocaproic acid, CaP = calcium phosphate,
CSMA = chondroitin sulfate methacrylate, FDM = fused deposition modeling, HA = hyaluronic acid, HAp = hydroxyapatite, HRP = horseradish
peroxidase, PAAm = poly(acrylamide), PCL = poly(ε-caprolactone), PDLLA = poly(D,L-lactide acid), PECDA = poly(ε-caprolactone)-
poly(ethylene glycol)-poly(ε-caprolactone) diacrylate, PEG-DA = poly(ethylene glycol) diacrylate, PGA = poly(glycolic acid), PLGA = poly(D,L-
lactide-co-glycolide), PLLA = poly(L-lactic acid), PNIPAAm = poly(N-isopropylacrylamide), PVA = poly(vinyl alcohol), PVB = poly(vinyl
butyrate), SCPL = solvent-casting particulate-leaching, SLA = stereolithography, and SLS = selective laser sintering.

Figure 3. Synthetic materials-guided osteochondral regeneration has utilized four major fabrication techniquesSCPL, freeze-drying, additive
manufacturing, and electrospinningwith a variety of materials in configurations of biphasic, triphasic, or gradient scaffolds.

and surface area. Extensive research has demonstrated the Furthermore, bioceramics can be electrospun either as a sol−
fabrication of complex fiber shapes (e.g., solid, helical, and gel or as a composite with a polymer in solution.179
hollow) via phase separation of two materials spun from the A number of new studies have utilized electrospinning to
same solution and controlled alignment (e.g., mesh, parallel, prepare instructive osteochondral scaffolds that span from the
and patterned) by using unique counter electrode arrange- articular cartilage to the cancellous bone.176,180 Most often,
ments and movements.176 Utilization of magnetically respon- these scaffolds have been formed by combining individually
sive polymers affords the opportunity for magnetically assisted prepared, discrete electrospun layers using solvent-bonding,
fabrication, in which fibers can be aligned with direction of an freeze-drying, or other methods. For example, Yunos et al.
applied magnetic field.177,178 Electrospinning is a versatile formed a biphasic scaffold by electrospinning poly(D,L-lactide)
fabrication technique suitable for many polymers, including (PDLLA) (the cartilage phase) of varying thicknesses (up to
synthetic and natural polymers and combinations of the two. 150 μm) onto a bioglass foam (the bone phase).181 This
4328 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

scaffold was able to support chondrocytic proliferation on the Pore size can be influenced based on precise control of freezing
fibers and graded HAp formation as a function of PDLLA time, temperature, material, and other strategy-specific
thickness (during exposure to simulated body fluid). Thus, variables.188−190
while a biphasic scaffold, the formation of mineralized fibers at Freeze-drying has been utilized to create instructive scaffolds
the interface paralleled the calcified cartilage layer of the native for osteochondral regeneration having different degrees of
osteochondral interface. As this fabrication strategy advances complexity. Erickson et al. designed a biphasic scaffold by
for OCD regenerative strategies, magnetically assisted layering two distinct polymer solutions and subsequently
fabrication may help to form tighter interfaces between freeze-drying.170 The cartilage layer was a 4% chitosan and
discrete layers and complex spatially aligned fibers that can hyaluronic acid (CHA) mixture, and the bone layer consisted
be used to form more biomimetic phases. of 6% chitosan and alginate (CA) and 0.5% hydroxyapatite.
Solvent-Casting Particulate-Leaching. Solvent-casting For fabrication, the bone layer mixture was added over the
particulate-leaching (SCPL) is useful to produce materials with refrigerated cartilage layer mixture, centrifuged to remove
tunable pore sizes and shapes. In SCPL, a polymer is dissolved bubbles and create a tight interface, and then refrigerated,
in a solvent, introduced to an insoluble porogen, and then frozen, and lyophilized. Kang et al. designed a more complex
cross-linked and/or solvent evaporated. Following leaching of trilayer scaffold by a similar method but where PBS was used
the porogen, a network of pores is produced whose size and to wash out frozen crystals from pores.169 This scaffold
shape are dictated by the porogen. A wide variety of salts or consisted of a cartilage layer of PEG-DA, a transition layer of
other materials have been utilized as porogens for SCPL (e.g., PEG-DA with pore anisotropy, and a mineralized bone layer of
NaCl salt, paraffin beads, and gelatin beads) and chosen for PEG-DA, CaP, and N-acryloyl 6-aminocaproic acid (A6ACA).
properties such as their solubility, shape, size, and/or thermal The bottom two layers were fabricated by polymerizing the
stability. By first fusing the porogen template, interconnected first layer of PEG-DA and A6ACA at −20 °C for 24 h,
porosity may also be achieved. For instance, we utilized a fused followed by adding the second PEG-DA layer on top and
salt template, prepared by the addition of a small quantity of polymerizing at the same temperature. The resulting lower
water to sieved NaCl crystals, to form interconnected layer contained smaller, uniform crystals due to the 24 h
macropores of tunable size in PEG-DA hydrogels and PCL- uniform freeze, while the middle layer had columnar pore
DA shape memory polymers for osseous scaffolds.182,183 structures due to the frozen gel beneath it. The resulting
The use of SCPL to prepare instructive osteochondral bilayer scaffold was incubated in simulated body fluid for
scaffolds has largely relied on preparing biphasic or multiphasic mineralization and subjected to cell culture. Finally, to create
scaffolds whose differing pore sizes or shapes are leveraged to the final trilayer scaffold, a small amount of cell-laden PEG-DA
direct cell behavior. For example, in Mahapatra et al., a was photopolymerized on the upper surface of the bilayer
biphasic PDLLA scaffold was prepared with a small salt scaffold. This study showcased the ability to control pore
porogen for the dense layer (cartilage phase) whereas the directionality, which can mimic the anisotropic nature of
nanofibrous layer (bone phase) was created with a large salt chondral tissue. Such an approach may be utilized to prepare
porogen as well as with camphene to induce phase separation, instructive osteochondral scaffolds that mimic tissue collagen
creating two distinct layers.173 Scaffaro et al. also prepared a fiber alignment with the potential to achieve interfacial
biphasic scaffold using leachable porogens but utilized melt- complexity.
mixing rather than solvent-casting.184 Each of the mixtures of Additive Manufacturing. With the emergence of additive
polymer and porogenPLA/larger-sized salt and PCL/PEG/ manufacturing (i.e., 3D-printing), scaffolds may be prepared
smaller sized-saltwere sequentially melt-mixed and com- with a high degree of morphological and compositional
pression molded into cylindrical layers, followed by fusing the complexity through computer-aided design (CAD). The
layers together with a heat treatment and last leaching the formation of printed structures, most often in a layer-by-layer
porogens. Tang et al. created a PLGA triphasic scaffold with manner, affords a high degree of spatial control to prepare
graded pore size by utilizing gelatin-microspheres of three sizes multiple, unique zones.191 A wide variety of materials can be
to form a trilayered porogen template, fused by the addition of 3D-printed, including thermoplastics, hydrogels, polysacchar-
a small amount of ethanol/water solution as well as heat ides, ceramics, and composites merged by melting, UV-cross-
treatment.185 linking, particle fusion, and more.192 Recently, instructive
Freeze-Drying. Freeze-drying (including also thermally osteochondral scaffolds have utilized three major types of 3D
induced phase-separation [TIPS]), wherein ice acts as the printing, including stereolithography, fused deposition model-
porogen, is also utilized to fabricate porous scaffolds.186 This ing, and selective laser sintering.191
method involves the freezing of an aqueous polymeric solution Extrusion based 3D printing (i.e., fused deposition
and subsequent sublimation of the ice. Upon formation and modeling) is a process in which a melt or gel “ink” is
growth of ice, phase separation occurs as polymer chains are deposited in a layer-by layer fashion.191 This method has been
excluded and thereby form an interconnected network.187 The utilized for the generation of osteochondral scaffolds with
utility of freeze-drying is attributed to the lack of harmful spatially controlled composition and morphology. For
solvents as well as the ability to refine scaffold morphology, instance, Bittner et al.166 utilized multiple nozzles to coextrude
namely, pore directionality and pore homogeneity. For three mixtures of PCL and HAp. A triphasic scaffold was
instance, to control pore directionality, the cooling source prepared with zones of different pore sizes and HAp
can be introduced at a specific portion of the scaffold, causing concentrations: osseous phase (900 μm, 30% HAp), transition
freezing to travel from one point to another and thus phase (500 μm, 15% HAp), and cartilage phase (200 μm, 0%
producing pores oriented in that direction. Also, to influence HAp). While bioprinting has emerged as a type of fused
pore homogeneity, cooling rate can be changed, wherein faster deposition 3D printing, wherein cells are incorporated into the
rates lead to heterogeneous pore sizes due to nonuniform ice resin prior to printing, the typical inclusion of growth factors
nucleation, and slower rates lead to a uniform pore network. does not constitute a synthetic materials-driven approach.193
4329 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

However, polysaccharide printing has been achieved,194 and instructive chemical and physical properties. Herein, we have
recently, a method for bioprinting of growth-factor free highlighted how synthetic material-based scaffolds have been
materials as gradients has been reported in Idaszek et al.195 utilized in recent years for osteochondral tissue engineering.
This involved the controlled rapid mixing of multiple bioinks Early work relied on biphasic scaffolds, merged across a hard
prior to extrusion, producing gradients representing the interface, with only two synthetic materials-guided examples
transition from bone to cartilage. advancing to clinical studies. Thus, recent efforts have focused
Stereolithography (SLA), also known as optical fabrication on developing instructive, multiphasic scaffolds with spatially
or photosolidification, uses the solidification of liquid resins to complex chemical and morphological features by leveraging
form complex structures. This process typically involves a bath advanced fabrication methods. In doing so, the complex
of a liquid resin that is solidified in a layer-by-layer fashion with osteochondral interface, consisting of calcified cartilage and the
a focused laser beam. Favorably for tissue engineering scaffolds, subchondral bone plate, is better recapitulated and anticipated
this strategy consistently produces structures with higher to enhance biomechanical functionality. As exemplified herein,
resolution than traditional fused deposition modeling, as the further refinements to the chemical and morphological
determination is made by the optical spot size of the laser features, along with a greater degree of spatial control, are
rather than the extruding nozzle size.196 However, the use of a expected to enhance instructive behavior. To potentially
resin bath typically limits printing to a single material, thus
increase the potency and efficacy of such synthetic-based
hindering printing of scaffolds with multiple phases in a single
osteochondral scaffolds, the use of coculturing (i.e., controlling
step. More recently, multimaterial systems have been
culture of multiple cell lineages in separate scaffold
developed,197 which could provide a format for SLA printing
of osteochondral scaffolds. For example Wu et al. reported a zones),199−202 bioreactors (i.e., dynamic in vitro environment
scaffold composed of PEG-DA and β-TCP that was formed by to mimic native environment),203−205 and microcarriers (i.e.,
multimaterial mask projection stereolithography, a process that 100−400 μm particles that facilitate cell growth/expan-
effectively allows the use of multiple materials in one sion)202,206,207 may be employed. Microfluidic advancements
scaffold.174 This was accomplished by a bottom-up process represent significant promise in increased understanding of the
designed to print with “material A” (β-TCP), wash the scaffold microenvironments necessary for osteochondral regenera-
to remove liquid resin, and then print with “material B” (PEG- tion.208 Mechanically robust double network (DN) hydrogels
DA), resulting in a well-integrated biphasic scaffold. Another may provide an opportunity to produce osteochondral
promising technology in stereolithography is the use of two- scaffolds with surfaces that can withstand articulating forces
photon polymerization. This technology induces resin prior to tissue regeneration.57,209−213 New synthetic materials-
polymerization by the absorption of two photons using a guided osteochondral repair strategies are constantly emerging,
femtosecond laser, allowing for an extremely high precision and those that further address the spatial and compositional
cure. This permits a fabrication feature size as small as 100 nm. complexities of the native tissue will significantly advance the
Strategies using this method have shown promise in tissue potential of the field to treat OCDs. While protein-based
engineering.198 Recently, Maciulaitis et al. utilized two-photon scaffolds have been widely studied and are known to be
polymerization to form an osteochondral repair scaffold of a instructive, scaffolds that exclude proteins rely solely on
material deemed SZ2080 [consisting of 20% “inorganics” (i.e., material properties to direct regeneration and therefore isolate
methacryloxypropyl trimethoxysilane and zirconium propox- the potency of these cues such that they can be leveraged more
ide) and 80% organics (i.e., 2-(dimethylamino)ethyl meth- effectively.

■
acrylate)].172 The scaffold was designed with hexagonal pores,
set at a diameter of 100 μm and height of 51 μm, displaying AUTHOR INFORMATION
extremely precise morphological control.
Selective laser sintering (SLS) is an additive manufacturing Corresponding Author
strategy that utilizes a high-power laser to fuse particles of a set Melissa A. Grunlan − Department of Biomedical Engineering,
size and composition. This technique is conducive for gradient Department of Materials Science & Engineering, and
production as it is completed layer-by-layer, wherein a layer of Department of Chemistry, Texas A&M University, College
particle powder is added and then fused, and the process is Station, Texas 77843-2120, United States; orcid.org/0000-
repeated. Furthermore, the complex surfaces and pore shapes 0002-5428-0461; Email: [email protected]
in these scaffolds are more like that of native tissue, rather than
the geometrical and consistent alignment of extrusion printed Author
scaffolds. Du et al. highlights the use of SLS to fabricate a Michael T. Frassica − Department of Biomedical Engineering,
unique gradient-based scaffold of PCL and HAp.160 This study Texas A&M University, College Station, Texas 77843-2120,
utilized PCL microspheres and PCL microspheres modified United States
with HAp nanoparticles, both with diameters of ∼100 μm. Complete contact information is available at:
Decreasing ratios of PCL/HAp to PCL microspheres were https://pubs.acs.org/10.1021/acsbiomaterials.0c00753
added manually at designated positions along the fabrication
process. Thus, a gradient of HAp was developed, from 30% Notes
modified to 0% at 5% increments making for seven seamless
The authors declare no competing financial interest.
layers with interconnected pore sizes in the range 400−500
μm.

■ CONCLUSIONS
■ ACKNOWLEDGMENTS
Funding from the Texas A&M Engineering Experiment Station
Synthetic polymer scaffolds have the potential to guide the is acknowledged. M.T.F. thanks the Texas A&M University
regeneration of complex osteochondral tissues through their Office of Graduate and Profession Studies (OGAPS).
4330 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering

■
pubs.acs.org/journal/abseba Perspective

ABBREVIATIONS repair: current problems in the surgical management. Knee Surgery,

Sports Traumatology, Arthroscopy 2010, 18 (4), 434−447.
OCD, osteochondral defect (9) Armiento, A. R.; Stoddart, M. J.; Alini, M.; Eglin, D. Biomaterials
OA, osteoarthritis for Articular Cartilage Tissue Engineering: Learning from Biology.
ACI, autologous chondrocyte implantation Acta Biomater. 2018, 65, 1−20.
3D, three-dimensional (10) Gomoll, A. H.; Farr, J.; Gillogly, S. D.; Kercher, J. S.; Minas, T.
ECM, extracellular matrix Surgical management of articular cartilage defects of the knee.
MSC, mesenchymal stem cell Instructional course lectures 2011, 60, 461−483.
IGF-1, insulin-like growth factor-1 (11) Camp, C. L.; Stuart, M. J.; Krych, A. J. Current concepts of
TGFβ-1, transforming growth factor β-1 articular cartilage restoration techniques in the knee. Sports Health
BMP-2, bone morphogenetic protein-2 2014, 6 (3), 265−73.
VEGF, vascular endothelial growth factor (12) Hangody, L.; Fules, P. Autologous osteochondral mosaicplasty
HAp, hydroxyapatite for the treatment of full-thickness defects of weight-bearing joints: ten
GAG, glycosaminoglycan years of experimental and clinical experience. Journal of Bone and Joint
β-TCP, β-tricalcium phosphate Surgery 2003, 85-A (S2), 25−32.
PEG-DA, poly(ethylene glycol) diacrylate (13) Demange, M.; Gomoll, A. H. The use of osteochondral
allografts in the management of cartilage defects. Current Reviews in
PEG-DMA, poly(ethylene glycol) dimethacrylate
Musculoskeletal Medicine 2012, 5 (3), 229−235.
PCL, poly(ε-caprolactone)
(14) Ansari, S.; Khorshidi, S.; Karkhaneh, A. Engineering of
PLLA, poly(L-lactic acid) Gradient Osteochondral Tissue: From Nature to Lab. Acta Biomater.
PLGA, poly(D,L-lactide-co-glycolide) 2019, 87, 41−54.
HA, hyaluronic acid (15) Nukavarapu, S. P.; Dorcemus, D. L. Osteochondral tissue
CS, chondroitin sulfate engineering: Current strategies and challenges. Biotechnol. Adv. 2013,
PGA, poly(glycolic acid) 31 (5), 706−721.
PVA, poly(vinyl alcohol) (16) Hutmacher, D. W. Scaffolds in tissue engineering bone and
UHMWPE, ultra-high-molecular-weight poly(ethylene) cartilage. Biomaterials 2000, 21 (24), 2529−2543.
PMA, premarket approval (17) Park, K. M.; Shin, Y. M.; Kim, K.; Shin, H. Tissue Engineering
PVB, poly(vinyl butyrate) and Regenerative Medicine 2017: A Year in Review. Tissue Eng., Part
FDM, fused deposition modeling B 2018, 24 (5), 327−344.
SCPL, solvent-casting particulate-leaching (18) Caplan, A. I. Adult mesenchymal stem cells for tissue
PECDA, poly(ε-caprolactone)-poly(ethylene glycol)-poly- engineering versus regenerative medicine. J. Cell. Physiol. 2007, 213
(ε-caprolactone) diacrylate (2), 341−347.
PAAm, poly(acrylamide) (19) Rosenbaum, A. J.; Grande, D. A.; Dines, J. S. The use of
SLA, stereolithography mesenchymal stem cells in tissue engineering: A global assessment.
SLS, selective laser sintering Organogenesis 2008, 4 (1), 23−27.
(20) Deng, C.; Chang, J.; Wu, C. Bioactive scaffolds for
A6ACA, N-acryloyl 6-aminocaproic acid
osteochondral regeneration. Journal of Orthopaedic Translation 2019,
CSMA, chondroitin sulfate methacrylate 17, 15−25.
PNIPAAm, poly(N-isopropylacrylamide) (21) Di Luca, A.; Van Blitterswijk, C.; Moroni, L. The osteochondral
CaP, calcium phosphate interface as a gradient tissue: from development to the fabrication of
PDLLA, poly(D,L-lactide acid) gradient scaffolds for regenerative medicine. Birth Defects Res., Part C
CAD, computer-aided design 2015, 105 (1), 34−52.

■
(22) Yan, L.-P.; Oliveira, J. M.; Oliveira, A. L.; Reis, R. L. Current
REFERENCES Concepts and Challenges in Osteochondral Tissue Engineering and
Regenerative Medicine. ACS Biomater. Sci. Eng. 2015, 1 (4), 183−
(1) Buckwalter, J. A. Articular cartilage: injuries and potential for
200.
healing. Journal of orthopaedic and sports physical therapy 1998, 28 (4), (23) Babensee, J. E.; McIntire, L. V.; Mikos, A. G. Growth Factor
192−202. Delivery for Tissue Engineering. Pharm. Res. 2000, 17 (5), 497−504.
(2) Ambra, L. F.; de Girolamo, L.; Mosier, B.; Gomoll, A. H. (24) Ikada, Y. Challenges in tissue engineering. J. R. Soc., Interface
Review: interventions for cartilage disease: current state-of-the-art and 2006, 3, 589−601.
emerging technologies. Arthritis Rheumatol. 2017, 69 (7), 1363−1373. (25) Dutta, R. C.; Dutta, A. K. Cell-interactive 3D-scaffold; advances
(3) Bruns, J.; Werner, M.; Habermann, C. Osteochondritis and applications. Biotechnol. Adv. 2009, 27, 334−339.
Dissecans: Etiology, Pathology, and Imaging with a Special Focus (26) Kleinman, H. K.; Philp, D.; Hoffman, M. P. Role of the
on the Knee Joint. Cartilage 2018, 9 (4), 346−362. extracellular matrix in morphogenesis. Curr. Opin. Biotechnol. 2003, 14
(4) Ytrehus, B.; Carlson, C. S.; Ekman, S. Etiology and Pathogenesis (5), 526−532.
of Osteochondrosis. Vet. Pathol. 2007, 44 (4), 429−448. (27) Lutolf, M. P.; Hubbell, J. A. Synthetic biomaterials as
(5) Sophia Fox, A. J.; Bedi, A.; Rodeo, S. A. The Basic Science of instructive extracellular microenvironments for morphogenesis in
Articular Cartilage: Structure, Composition, and Function. Sports tissue engineering. Nat. Biotechnol. 2005, 23, 47−55.
Health 2009, 1 (6), 461−468. (28) Brandl, F.; Sommer, F.; Goepferich, A. Rational design of
(6) Lefkoe, T. P.; Trafton, P. G.; Ehrlich, M. G.; Walsh, W. R.; hydrogels for tissue engineering: Impact of physical factors on cell
Dennehy, D. T.; Barrach, H.-J.; Akelman, E. An Experimental Model behavior. Biomaterials 2007, 28, 134−6.
of Femoral Condylar Defect Leading to Osteoarthrosis. Journal of (29) Pennesi, C.; Scaglione, S.; Gionnoni, P.; Quarto, R. Regulatory
Orthopaedic Trauma 1993, 7 (5), 458−467. influence of scaffolds on cell behavior: How cells decode biomaterials.
(7) Sellards, R. A.; Nho, S. J.; Cole, B. J. Chondral injuries. Curr. Curr. Pharm. Biotechnol. 2011, 12, 151−159.
Opin. Rheumatol. 2002, 14 (2), 134−141. (30) Nichol, J. W.; Khademhosseini, A. Modular tissue engineering:
(8) Gomoll, A. H.; Madry, H.; Knutsen, G.; van Dijk, N.; Seil, R.; Engineering biological tissues from the bottom up. Soft Matter 2009,
Brittberg, M.; Kon, E. The subchondral bone in articular cartilage 5, 1312.

4331 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

(31) Khetan, S.; Burdick, J. A. Patterning hydrogels in three spontaneous osteochondral repair in cartilage tissue engineering.
dimensions towards controlling cellular interactions. Soft Matter 2011, Regenerative Biomaterials 2015, 2 (1), 9−19.
7, 830−838. (51) Dhandayuthapani, B.; Yoshida, Y.; Maekawa, T.; Kumar, D. S.
(32) Woodfield, T. B. F.; Moroni, L.; Malda, J. Combinatorial Polymeric Scaffolds in Tissue Engineering Application: A Review. Int.
approaches to controlling cell behaviour and tissue formation via J. Polym. Sci. 2011, 2011, 290602.
rapid-prototyping and smart scaffold design. Comb. Chem. High (52) Bongio, M.; van den Beucken, J. J. J. P.; Leeuwenburgh, S. C.
Throughput Screening 2009, 12, 562−579. G.; Jansen, J. A. Development of bone substitute materials: from
(33) Khademhosseini, A.; Langer, R. Microengineered hydrogels for ‘biocompatible’ to ‘instructive’. J. Mater. Chem. 2010, 20 (40), 8747−
tissue engineering. Biomaterials 2007, 28, 5087. 8759.
(34) McGlohorn, J. B.; Holder, J.; Grimes, L. W.; Thomas, C. B.; (53) Hunziker, E. B.; Quinn, T. M.; Häuselmann, H. J. Quantitative
Burg, K. J. L. Evaluation of smooth muscle cell response using two structural organization of normal adult human articular cartilage.
types of porous polylactide scaffolds with differing pore topography. Osteoarthritis and Cartilage 2002, 10 (7), 564−572.
Tissue Eng. 2004, 10, 505−514. (54) Lyons, T. J.; Stoddart, R. W.; McClure, S. F.; McClure, J. The
(35) Discher, D. E.; Janmey, P.; Wang, Y.-l. Tissue cells feel and tidemark of the chondro-osseous junction of the normal human knee
respond to the stiffness of their substrate. Science 2005, 310, 1139− joint. J. Mol. Histol. 2005, 36 (3), 207−215.
1143. (55) Lepage, S. I. M.; Robson, N.; Gilmore, H.; Davis, O.; Hooper,
(36) Engler, A. J.; Sen, S.; Sweeney, H. L.; Discher, D. E. Matrix A.; St. John, S.; Kamesan, V.; Gelis, P.; Carvajal, D.; Hurtig, M.; Koch,
elasticity directs stem cell lineage specification. Cell 2006, 126, 677− T. G. Beyond Cartilage Repair: The Role of the Osteochondral Unit
689. in Joint Health and Disease. Tissue Eng., Part B 2019, 25 (2), 114−
(37) Liao, H.; Munoz-Pinto, D.; Qu, X.; Hou, Y.; Grunlan, M. A.; 125.
Hahn, M. S. Influence of hydrogel mechanical properties and mesh (56) Woodruff, M. A.; Lange, C.; Reichert, J.; Berner, A.; Chen, F.;
size on vocal fold fibroblast extracellular matrix production and Fratzl, P.; Schantz, J.-T.; Hutmacher, D. W. Bone tissue engineering:
phenotype. Acta Biomater. 2008, 4, 1161−1171. from bench to bedside. Mater. Today 2012, 15 (10), 430−435.
(38) Dado, D.; Levenberg, S. Cell-scaffold mechanical interplay (57) Means, A. K.; Grunlan, M. A. Modern Strategies To Achieve
within engineered tissues. Semin. Cell Dev. Biol. 2009, 20, 656−664. Tissue-Mimetic, Mechanically Robust Hydrogels. ACS Macro Lett.
(39) Lin, S.; Sangaj, N.; Razafiarison, T.; Zhang, C.; Varghese, S. 2019, 8 (6), 705−713.
Influence of physical properties of biomaterials on cellular behavior. (58) Madry, H.; Orth, P.; Cucchiarini, M. Role of the Subchondral
Pharm. Res. 2011, 28, 1422−1430. Bone in Articular Cartilage Degeneration and Repair. Journal of the
(40) Benoit, D. S. W.; Schwartz, M. P.; Durney, A. R.; Anseth, K. S. American Academy of Orthopaedic Surgeons 2016, 24 (4), e45−e46.
Small functional groups for controlled differentiation of hydrogel- (59) Jeon, J. E.; Vaquette, C.; Klein, T. J.; Hutmacher, D. W.
encapsulated human mesenchymal stem cells. Nat. Mater. 2008, 7, Perspectives in Multiphasic Osteochondral Tissue Engineering. Anat.
816−823. Rec. 2014, 297 (1), 26−35.
(41) Escobar Ivirico, J. L.; Salmeron Sanchez, M.; Gomez Ribelles, J. (60) Schek, R. M.; Taboas, J. M.; Segvich, S. J.; Hollister, S. J.;
L.; Monleon Pradas, M.; Soria, J. M.; Gomes, M. E.; Reis, R. L.; Krebsbach, P. H. Engineered osteochondral grafts using biphasic
Mano, J. F. Proliferation and differentiation of goat bone marrow composite solid free-form fabricated scaffolds. Tissue Eng. 2004, 10
stromal cells in 3D scaffolds with tunable hydrophilicity. J. Biomed. (9−10), 1376−85.
Mater. Res., Part B 2009, 91B, 277−286. (61) Oliveira, J. M.; Rodrigues, M. T.; Silva, S. S.; Malafaya, P. B.;
(42) Song, J. H.; Yoon, B. H.; Kim, H. E.; Kim, H. W. Bioactive and Gomes, M. E.; Viegas, C. A.; Dias, I. R.; Azevedo, J. T.; Mano, J. F.;
degradable hybridized nanofibers of gelatin-siloxane for bone Reis, R. L. Novel hydroxyapatite/chitosan bilayered scaffold for
regeneration. J. Biomed. Mater. Res., Part A 2008, 84 (4), 875−884. osteochondral tissue-engineering applications: Scaffold design and its
(43) Ning, C. Q.; Mehta, J.; El-Ghannam, A. Effect of silica on the performance when seeded with goat bone marrow stromal cells.
bioactivity of calcium phosphate composites in vitro. J. Mater. Sci.: Biomaterials 2006, 27 (36), 6123−6137.
Mater. Med. 2005, 16, 355−360. (62) Nooeaid, P.; Salih, V.; Beier, J. P.; Boccaccini, A. R.
(44) Ayala, R.; Zhang, C.; Yang, D.; Hwang, Y.; Aung, A.; Shroff, S. Osteochondral tissue engineering: scaffolds, stem cells and
S.; Arce, F. T.; Lal, R.; Arya, G.; Varghese, S. Engineering the cell- applications. Journal of Cellular and Molecular Medicine 2012, 16
material interface for controlling stem cell adhesion, migration, and (10), 2247−2270.
differentiation. Biomaterials 2011, 32, 3700−3711. (63) Longley, R.; Ferreira, A. M.; Gentile, P. Recent Approaches to
(45) Jansen, E. J.; Sladek, R. E.; Bahar, H.; Yaffe, A.; Gijbels, M. J.; the Manufacturing of Biomimetic Multi-Phasic Scaffolds for
Kuijer, R.; Bulstra, S. K.; Guldemond, N. A.; Binderman, I.; Koole, L. Osteochondral Regeneration. Int. J. Mol. Sci. 2018, 19 (6), 1755.
H. Hydrophobicity as a design criterion for polymer scaffolds in bone (64) Hacker, M. C.; Mikos, A. G. Synthetic Polymers. In Principles of
tissue engineering. Biomaterials 2005, 26 (21), 4423−31. Regenerative Medicine, 2nd ed.; Academic Press: San Diego, 2011; pp
(46) Fisher, J. P.; Lalani, Z.; Bossano, C. M.; Brey, E. M.; Demian, 587−622. DOI: 10.1016/B978-0-12-381422-7.10033-1.
N.; Johnston, C. M.; Dean, D.; Jansen, J. A.; Wong, M. E.; Mikos, A. (65) Sheikh, Z.; Najeeb, S.; Khurshid, Z.; Verma, V.; Rashid, H.;
G. Effect of biomaterial properties on bone healing in a rabbit tooth Glogauer, M. Biodegradable Materials for Bone Repair and Tissue
extration socket model. J. Biomed. Mater. Res. 2004, 68, 428−438. Engineering Applications. Materials 2015, 8 (9), 5744−5794.
(47) Anseth, K. S.; Bowman, C. N.; Brannon-Peppas, L. Mechanical (66) Du, Y.; Guo, J. L.; Wang, J.; Mikos, A. G.; Zhang, S.
properties of hydrogels and their experimental determination. Hierarchically Designed Bone Scaffolds: From Internal Cues to
Biomaterials 1996, 17 (17), 1647−1657. External Stimuli. Biomaterials 2019, 218, 119334.
(48) Drury, J. L.; Mooney, D. G. Hydrogels for tissue engineering: (67) Yu, X.; Tang, X.; Gohil, S. V.; Laurencin, C. T. Biomaterials for
Scaffold design variables and applications. Biomaterials 2003, 24, Bone Regenerative Engineering. Adv. Healthcare Mater. 2015, 4 (9),
4337−4351. 1268−1285.
(49) Park, H.; Guo, X.; Temenoff, J. S.; Tabata, Y.; Caplan, A. I.; (68) Stevens, M. M. Biomaterials for bone tissue engineering. Mater.
Kurtis Kasper, F.; Mikos, A. G. Effect of swelling ratio of injectable Today 2008, 11 (5), 18−25.
hydrogel composites on chondrogenic differentiation of encapsulated (69) Albrektsson, T.; Johansson, C. Osteoinduction, osteoconduc-
rabbit marrow mesenchymal stem cells in vitro. Biomacromolecules tion and osseointegration. European Spine Journal 2001, 10 (2), S96−
2009, 10, 541−546. S101.
(50) Pan, Z.; Duan, P.; Liu, X.; Wang, H.; Cao, L.; He, Y.; Dong, J.; (70) LeGeros, R. Z. Calcium Phosphate-Based Osteoinductive
Ding, J. Effect of porosities of bilayered porous scaffolds on Materials. Chem. Rev. 2008, 108 (11), 4742−4753.

4332 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

(71) Tang, Z.; Li, X.; Tan, Y.; Fan, H.; Zhang, X. The material and (91) Carrow, J. K.; Di Luca, A.; Dolatshahi-Pirouz, A.; Moroni, L.;
biological characteristics of osteoinductive calcium phosphate Gaharwar, A. K. 3D-printed bioactive scaffolds from nanosilicates and
ceramics. Regenerative biomaterials 2018, 5 (1), 43−59. PEOT/PBT for bone tissue engineering. Regenerative Biomaterials
(72) Kretlow, J. D.; Mikos, A. G. Review: Mineralization of Synthetic 2019, 6 (1), 29−37.
Polymer Scaffolds for Bone Tissue Engineering. Tissue Eng. 2007, 13 (92) Zhang, X.; Zeng, D.; Li, N.; Wen, J.; Jiang, X.; Liu, C.; Li, Y.
(5), 927−938. Functionalized mesoporous bioactive glass scaffolds for enhanced
(73) Di Luca, A.; Ostrowska, B.; Lorenzo-Moldero, I.; Lepedda, A.; bone tissue regeneration. Sci. Rep. 2016, 6, 19361.
Swieszkowski, W.; Van Blitterswijk, C.; Moroni, L. Gradients in pore (93) Zhang, K.; Fan, Y.; Dunne, N.; Li, X. Effect of microporosity on
size enhance the osteogenic differentiation of human mesenchymal scaffolds for bone tissue engineering. Regenerative Biomaterials 2018, 5
stromal cells in three-dimensional scaffolds. Sci. Rep. 2016, 6, 22898. (2), 115−124.
(74) Roseti, L.; Parisi, V.; Petretta, M.; Cavallo, C.; Desando, G.; (94) Samavedi, S.; Whittington, A. R.; Goldstein, A. S. Calcium
Bartolotti, I.; Grigolo, B. Scaffolds for Bone Tissue Engineering: State phosphate ceramics in bone tissue engineering: A review of properties
of the art and new perspectives. Mater. Sci. Eng., C 2017, 78, 1246− and their influence on cell behavior. Acta Biomater. 2013, 9 (9),
1262. 8037−8045.
(75) Weber, F. E. Reconsidering Osteoconduction in the Era of (95) Breuls, R. G. M.; Jiya, T. U.; Smit, T. H. Scaffold stiffness
Additive Manufacturing. Tissue Eng., Part B 2019, 25 (5), 375−386. influences cell behavior: opportunities for skeletal tissue engineering.
(76) Habibovic, P.; Sees, T. M.; van den Doel, M. A.; van Open Orthop J. 2008, 2, 103−109.
Blitterswijk, C. A.; de Groot, K. Osteoinduction by biomaterials (96) Discher, D. E.; Mooney, D. J.; Zandstra, P. W. Growth factors,
Physicochemical and structural influences. J. Biomed. Mater. Res., Part matrices, and forces combine and control stem cells. Science 2009,
A 2006, 77A (4), 747−762. 324, 1673−1677.
(77) Harburger, D. S.; Calderwood, D. A. Integrin signalling at a (97) Huebsch, N.; Arany, P. R.; Mao, A. S.; Shvartsman, D.; Ali, O.
glance. J. Cell Sci. 2009, 122 (2), 159. A.; Bencherif, S. A.; Rivera-Feliciano, J.; Mooney, D. J. Harnessing
(78) Maia, F. R.; Bidarra, S. J.; Granja, P. L.; Barrias, C. C. traction-mediated manipulation of the cell/matrix interface to control
Functionalization of biomaterials with small osteoinductive moieties. stem-cell fate. Nat. Mater. 2010, 9, 518.
Acta Biomater. 2013, 9 (11), 8773−8789. (98) Chatterjee, K.; Lin-Gibson, S.; Wallace, W. E.; Parekh, S. H.;
(79) Habibovic, P.; de Groot, K. Osteoinductive biomaterials Lee, Y. J.; Cicerone, M. T.; Young, M. F.; Simon, C. G. The effect of
properties and relevance in bone repair. J. Tissue Eng. Regener. Med. 3D hydrogel scaffold modulus on osteoblast differentiation and
2007, 1 (1), 25−32. mineralization revealed by combinatorial screening. Biomaterials
(80) Breitwieser, G. E. Extracellular calcium as an integrator of tissue
2010, 31 (19), 5051−5062.
function. Int. J. Biochem. Cell Biol. 2008, 40 (8), 1467−1480. (99) Turnbull, G.; Clarke, J.; Picard, F.; Riches, P.; Jia, L.; Han, F.;
(81) Barradas, A. M. C.; Fernandes, H. A. M.; Groen, N.; Chai, Y.
Li, B.; Shu, W. 3D bioactive composite scaffolds for bone tissue
C.; Schrooten, J.; van de Peppel, J.; van Leeuwen, J. P. T. M.; van
engineering. Bioactive Materials 2018, 3 (3), 278−314.
Blitterswijk, C. A.; de Boer, J. A calcium-induced signaling cascade
(100) Velasco, M. A.; Narváez-Tovar, C. A.; Garzón-Alvarado, D. A.
leading to osteogenic differentiation of human bone marrow-derived
Design, materials, and mechanobiology of biodegradable scaffolds for
mesenchymal stromal cells. Biomaterials 2012, 33 (11), 3205−3215.
bone tissue engineering. BioMed Res. Int. 2015, 2015, 729076.
(82) Chai, Y. C.; Carlier, A.; Bolander, J.; Roberts, S. J.; Geris, L.;
(101) Amini, A. R.; Laurencin, C. T.; Nukavarapu, S. P. Bone Tissue
Schrooten, J.; Van Oosterwyck, H.; Luyten, F. P. Current views on
Engineering: Recent Advances and Challenges. Critical Reviews in
calcium phosphate osteogenicity and the translation into effective
bone regeneration strategies. Acta Biomater. 2012, 8 (11), 3876− Biomedical Engineering 2012, 40 (5), 363−408.
(102) Seo, S.-J.; Mahapatra, C.; Singh, R. K.; Knowles, J. C.; Kim,
3887.
(83) Aquino-Martínez, R.; Monroe, D. G.; Ventura, F. Calcium H.-W. Strategies for osteochondral repair: Focus on scaffolds. J. Tissue
mimics the chemotactic effect of conditioned media and is an effective Eng. 2014, 5, 204173141454185.
inducer of bone regeneration. PLoS One 2019, 14 (1), No. e0210301. (103) Fu, Q.; Saiz, E.; Rahaman, M. N.; Tomsia, A. P. Bioactive glass
(84) Malhotra, A.; Habibovic, P. Calcium Phosphates and scaffolds for bone tissue engineering: state of the art and future
Angiogenesis: Implications and Advances for Bone Regeneration. perspectives. Mater. Sci. Eng., C 2011, 31 (7), 1245−1256.
Trends Biotechnol. 2016, 34 (12), 983−992. (104) Bianco, A.; Cacciotti, I.; Lombardi, M.; Montanaro, L.;
(85) Dinarvand, P.; Seyedjafari, E.; Shafiee, A.; Babaei Jandaghi, A.; Bemporad, E.; Sebastiani, M. F-substituted hydroxyapatite nano-
Doostmohammadi, A.; Fathi, M. H.; Farhadian, S.; Soleimani, M. powders: Thermal stability, sintering behaviour and mechanical
New Approach to Bone Tissue Engineering: Simultaneous Applica- properties. Ceram. Int. 2010, 36 (1), 313−322.
tion of Hydroxyapatite and Bioactive Glass Coated on a Poly(l-lactic (105) Dorozhkin, S. V. Calcium Orthophosphates in Nature,
acid) Scaffold. ACS Appl. Mater. Interfaces 2011, 3 (11), 4518−4524. Biology and Medicine. Materials 2009, 2 (2), 399−498.
(86) Rezwan, K.; Chen, Q. Z.; Blaker, J. J.; Boccaccini, A. R. (106) Velard, F.; Laurent-Maquin, D.; Braux, J.; Guillaume, C.;
Biodegradable and bioactive porous polymer/inorganic composite Bouthors, S.; Jallot, E.; Nedelec, J.-M.; Belaaouaj, A.; Laquerriere, P.
scaffolds for bone tissue engineering. Biomaterials 2006, 27, 3413− The effect of zinc on hydroxyapatite-mediated activation of human
3431. polymorphonuclear neutrophils and bone implant-associated acute
(87) Wang, W.; Yeung, K. W. K. Bone grafts and biomaterials inflammation. Biomaterials 2010, 31 (8), 2001−2009.
substitutes for bone defect repair: A review. Bioactive Materials 2017, (107) Honda, M.; Kikushima, K.; Kawanobe, Y.; Konishi, T.;
2 (4), 224−247. Mizumoto, M.; Aizawa, M. Enhanced early osteogenic differentiation
(88) Munoz-Pinto, D. J.; Jimenez-Vergara, A. C.; Hou, Y.; Hayenga, by silicon-substituted hydroxyapatite ceramics fabricated via ultra-
H. N.; Rivas, A.; Grunlan, M.; Hahn, M. S. Osteogenic potential of sonic spray pyrolysis route. J. Mater. Sci.: Mater. Med. 2012, 23 (12),
poly(ethylene glycol)-poly(dimethylsiloxane) hybrid hydrogels. Tissue 2923−2932.
Eng., Part A 2012, 18 (15−16), 1710−9. (108) Kim, E.-J.; Bu, S.-Y.; Sung, M.-K.; Choi, M.-K. Effects of
(89) Frassica, M. T.; Jones, S. K.; Diaz-Rodriguez, P.; Hahn, M. S.; Silicon on Osteoblast Activity and Bone Mineralization of MC3T3-E1
Grunlan, M. A. Incorporation of a silicon-based polymer to PEG-DA Cells. Biol. Trace Elem. Res. 2013, 152 (1), 105−112.
templated hydrogel scaffolds for bioactivity and osteoinductivity. Acta (109) Beck, G. R.; Ha, S.-W.; Camalier, C. E.; Yamaguchi, M.; Li, Y.;
Biomater. 2019, 99, 100. Lee, J.-K.; Weitzmann, M. N. Bioactive silica based nanoparticles
(90) Kerativitayanan, P.; Gaharwar, A. K. Elastomeric and stimulate bone forming osteoblasts, suppress bone esorbing
mechanically stiff nanocomposites from poly(glycerol sebacate) and osteoclasts, and enhance bone mineral density in vivo. Nanomedicine
bioactive nanosilicates. Acta Biomater. 2015, 26, 34−44. 2012, 8 (6), 793−803.

4333 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

(110) Becerra, J.; Andrades, J. A.; Guerado, E.; Zamora-Navas, P.; (129) Xiao, H.; Huang, W.; Xiong, K.; Ruan, S.; Yuan, C.; Mo, G.;
López-Puertas, J. M.; Reddi, A. H. Articular Cartilage: Structure and Tian, R.; Zhou, S.; She, R.; Ye, P.; Liu, B.; Deng, J. Osteochondral
Regeneration. Tissue Eng., Part B 2010, 16 (6), 617−627. repair using scaffolds with gradient pore sizes constructed with silk
(111) Walter, S. G.; Ossendorff, R.; Schildberg, F. A. Articular fibroin, chitosan, and nano-hydroxyapatite. Int. J. Nanomed. 2019, 14,
cartilage regeneration and tissue engineering models: a systematic 2011−2027.
review. Archives of Orthopaedic and Trauma Surgery 2019, 139 (3), (130) Hu, J.; Feng, K.; Liu, X.; Ma, P. X. Chondrogenic and
305−316. osteogenic differentiations of human bone marrow-derived mesen-
(112) Burris, D. L.; Ramsey, L.; Graham, B. T.; Price, C.; Moore, A. chymal stem cells on a nanofibrous scaffold with designed pore
C. How Sliding and Hydrodynamics Contribute to Articular Cartilage network. Biomaterials 2009, 30 (28), 5061−5067.
Fluid and Lubrication Recovery. Tribol. Lett. 2019, 67 (2), 46. (131) Murphy, C. M.; Haugh, M. G.; O’Brien, F. J. The effect of
(113) Bružauskaitė, I.; Bironaitė, D.; Bagdonas, E.; Bernotienė, E. mean pore size on cell attachment, proliferation and migration in
Scaffolds and cells for tissue regeneration: different scaffold pore collagen−glycosaminoglycan scaffolds for bone tissue engineering.
sizesdifferent cell effects. Cytotechnology 2016, 68 (3), 355−369. Biomaterials 2010, 31 (3), 461−466.
(114) Nava, M. M.; Draghi, L.; Giordano, C.; Pietrabissa, R. The (132) Oh, S. H.; Kim, T. H.; Im, G. I.; Lee, J. H. Investigation of
effect of scaffold pore size in cartilage tissue engineering. J. Appl. Pore Size Effect on Chondrogenic Differentiation of Adipose Stem
Biomater. Funct. Mater. 2016, 14 (3), e223−e229. Cells Using a Pore Size Gradient Scaffold. Biomacromolecules 2010, 11
(115) Gao, L.; McBeath, R.; Chen, C. S. Stem Cell Shape Regulates (8), 1948−1955.
a Chondrogenic Versus Myogenic Fate Through Rac1 and N- (133) Engler, A. J.; Tse, J. R. Stiffness gradients mimicking in vivo
Cadherin. Stem Cells 2010, 28 (3), 564−572. tissue variation regulate mesenchymal stem cell fate. PLoS One 2011,
(116) Panadero, J. A.; Lanceros-Mendez, S.; Ribelles, J. L. G. 6, No. e15978.
Differentiation of mesenchymal stem cells for cartilage tissue (134) Chen, K.; Shi, P.; Teh, T. K. H.; Toh, S. L.; Goh, J. C. H. In
engineering: Individual and synergetic effects of three-dimensional vitro generation of a multilayered osteochondral construct with an
environment and mechanical loading. Acta Biomater. 2016, 33, 1−12. osteochondral interface using rabbit bone marrow stromal cells and a
(117) Muhammad, H.; Rais, Y.; Miosge, N.; Ornan, E. M. The silk peptide-based scaffold. J. Tissue Eng. Regener. Med. 2016, 10 (4),
primary cilium as a dual sensor of mechanochemical signals in 284−293.
chondrocytes. Cell. Mol. Life Sci. 2012, 69 (13), 2101−2107. (135) Diaz-Rodriguez, P.; Erndt-Marino, J. D.; Gharat, T.; Munoz
(118) Uzieliene, I.; Bernotas, P.; Mobasheri, A.; Bernotiene, E. The Pinto, D. J.; Samavedi, S.; Bearden, R.; Grunlan, M. A.; Saunders, W.
Role of Physical Stimuli on Calcium Channels in Chondrogenic B.; Hahn, M. S. Toward zonally tailored scaffolds for osteochondral
Differentiation of Mesenchymal Stem Cells. Int. J. Mol. Sci. 2018, 19 differentiation of synovial mesenchymal stem cells. J. Biomed. Mater.
(10), 2998. Res., Part B 2019, 107 (6), 2019−2029.
(119) Huey, D. J.; Hu, J. C.; Athanasiou, K. A. Unlike Bone, (136) Khanarian, N. T.; Haney, N. M.; Burga, R. A.; Lu, H. H. A
Cartilage Regeneration Remains Elusive. Science 2012, 338 (6109), functional agarose-hydroxyapatite scaffold for osteochondral interface
917. regeneration. Biomaterials 2012, 33 (21), 5247−5258.
(120) Mehdizadeh, M.; Yang, J. Design Strategies and Applications (137) Lee, W. D.; Gawri, R.; Pilliar, R. M.; Stanford, W. L.; Kandel,
of Tissue Bioadhesives. Macromol. Biosci. 2013, 13 (3), 271−288. R. A. Sol gel-derived hydroxyapatite films over porous calcium
(121) Yoo, H. S.; Lee, E. A.; Yoon, J. J.; Park, T. G. Hyaluronic acid polyphosphate substrates for improved tissue engineering of
modified biodegradable scaffolds for cartilage tissue engineering. osteochondral-like constructs. Acta Biomater. 2017, 62, 352−361.
Biomaterials 2005, 26 (14), 1925−1933. (138) Komuro, H.; Wint, W. Y.; Horiuchi, N.; Nozaki, K.; Sasano,
(122) Wang, D.-A.; Varghese, S.; Sharma, B.; Strehin, I.; Fermanian, T.; Miyashin, M.; Yamashita, K.; Nagai, A. An oriented hydroxyapatite
S.; Gorham, J.; Fairbrother, D. H.; Cascio, B.; Elisseeff, J. H. film with arrayed plate-like particles enhance chondrogenic differ-
Multifunctional chondroitin sulphate for cartilage tissue−biomaterial entiation of ATDC5 cells. J. Biomed. Mater. Res., Part A 2020, 108,
integration. Nat. Mater. 2007, 6, 385. 537.
(123) Wu, S.-C.; Chen, C.-H.; Chang, J.-K.; Fu, Y.-C.; Wang, C.-K.; (139) Yang, J.; Liu, Y.; He, L.; Wang, Q.; Wang, L.; Yuan, T.; Xiao,
Eswaramoorthy, R.; Lin, Y.-S.; Wang, Y.-H.; Lin, S.-Y.; Wang, G.-J.; Y.; Fan, Y.; Zhang, X. Icariin conjugated hyaluronic acid/collagen
Ho, M.-L. Hyaluronan initiates chondrogenesis mainly via CD44 in hydrogel for osteochondral interface restoration. Acta Biomater. 2018,
human adipose-derived stem cells. J. Appl. Physiol. 2013, 114 (11), 74, 156−167.
1610−1618. (140) Khader, A.; Arinzeh, T. L. Biodegradable zinc oxide composite
(124) Ruffell, B.; Poon, G. F. T.; Lee, S. S. M.; Brown, K. L.; Tjew, scaffolds promote osteochondral differentiation of mesenchymal stem
S.-L.; Cooper, J.; Johnson, P. Differential Use of Chondroitin Sulfate cells. Biotechnol. Bioeng. 2020, 117, 194.
to Regulate Hyaluronan Binding by Receptor CD44 in Inflammatory (141) Yu, X.; Zhao, T.; Qi, Y.; Luo, J.; Fang, J.; Yang, X.; Liu, X.; Xu,
and Interleukin 4-activated Macrophages. J. Biol. Chem. 2011, 286 T.; Yang, Q.; Gou, Z.; Dai, X. In vitro Chondrocyte Responses in Mg-
(22), 19179−19190. doped Wollastonite/Hydrogel Composite Scaffolds for Osteochon-
(125) Kazemnejad, S.; Khanmohammadi, M.; Baheiraei, N.; Arasteh, dral Interface Regeneration. Sci. Rep. 2018, 8 (1), 17911.
S. Current State of Cartilage Tissue Engineering using Nanofibrous (142) Wang, C.; Chen, B.; Wang, W.; Zhang, X.; Hu, T.; He, Y.; Lin,
Scaffolds and Stem Cells. Avicenna J. Med. Biotechnol 2017, 9 (2), 50− K.; Liu, X. Strontium released bi-lineage scaffolds with immunomo-
65. dulatory properties induce a pro-regenerative environment for
(126) Fell, N. L. A.; Lawless, B. M.; Cox, S. C.; Cooke, M. E.; osteochondral regeneration. Mater. Sci. Eng., C 2019, 103, 109833.
Eisenstein, N. M.; Shepherd, D. E. T.; Espino, D. M. The role of (143) Lin, R.; Deng, C.; Li, X.; Liu, Y.; Zhang, M.; Qin, C.; Yao, Q.;
subchondral bone, and its histomorphology, on the dynamic Wang, L.; Wu, C. Copper-incorporated bioactive glass-ceramics
viscoelasticity of cartilage, bone and osteochondral cores. Osteo- inducing anti-inflammatory phenotype and regeneration of cartilage/
arthritis and Cartilage 2019, 27 (3), 535−543. bone interface. Theranostics 2019, 9 (21), 6300−6313.
(127) Stewart, H. L.; Kawcak, C. E. The Importance of Subchondral (144) Baker, M. I.; Walsh, S. P.; Schwartz, Z.; Boyan, B. D. A review
Bone in the Pathophysiology of Osteoarthritis. Frontiers in Veterinary of polyvinyl alcohol and its uses in cartilage and orthopedic
Science 2018, 5, 178. applications. J. Biomed. Mater. Res., Part B 2012, 100B (5), 1451−
(128) Yousefi, A.-M.; Hoque, M. E.; Prasad, R. G. S. V.; Uth, N. 1457.
Current strategies in multiphasic scaffold design for osteochondral (145) Nathwani, D.; McNicholas, M.; Hart, A.; Miles, J.; Bobic, V.
tissue engineering: A review. J. Biomed. Mater. Res., Part A 2015, 103 Partial Resurfacing of the Knee with the BioPoly Implant: Interim
(7), 2460−2481. Report at 2 Years. JBJS Open Access 2017, 2 (2), No. e0011.

4334 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

(146) Pina, S.; Ribeiro, V.; Oliveira, J. M.; Reis, R. L. Pre-clinical and (162) Mellor, L. F.; Huebner, P.; Cai, S.; Mohiti-Asli, M.; Taylor, M.
Clinical Management of Osteochondral Lesions. In Regenerative A.; Spang, J.; Shirwaiker, R. A.; Loboa, E. G. Fabrication and
Strategies for the Treatment of Knee Joint Disabilities; Oliveira, J. M., Evaluation of Electrospun, 3D-Bioplotted, and Combination of
Reis, R. L., Eds.; Springer International Publishing: Cham, 2017; pp Electrospun/3D-Bioplotted Scaffolds for Tissue Engineering Appli-
147−161. DOI: 10.1007/978-3-319-44785-8_8. cations. BioMed Res. Int. 2017, 2017, 6956794.
(147) Jeuken, M. R.; Roth, K. A.; Peters, J. R. W. R.; Van Donkelaar, (163) Rajasekharan, A. K.; Bordes, R.; Sandström, C.; Ekh, M.;
C. C.; Thies, C. J.; Van Rhijn, W. L.; Emans, J. P. Polymers in Andersson, M. Hierarchical and Heterogeneous Bioinspired Compo-
Cartilage Defect Repair of the Knee: Current Status and Future sitesMerging Molecular Self-Assembly with Additive Manufactur-
Prospects. Polymers 2016, 8 (6), 219. ing. Small 2017, 13 (28), 1700550.
(148) Kon, E.; Filardo, G.; Perdisa, F.; Venieri, G.; Marcacci, M. (164) Su, C.; Su, Y.; Li, Z.; Haq, M. A.; Zhou, Y.; Wang, D. In situ
Clinical results of multilayered biomaterials for osteochondral synthesis of bilayered gradient poly(vinyl alcohol)/hydroxyapatite
regeneration. Journal of Experimental Orthopaedics 2014, 1, 10. composite hydrogel by directional freezing-thawing and electro-
(149) Slivka, M. A.; Leatherbury, N. C.; Kieswetter, K.; Niederauer, phoresis method. Mater. Sci. Eng., C 2017, 77, 76−83.
G. G. Porous, Resorbable, Fiber-Reinforced Scaffolds Tailored for (165) Yao, H.; Kang, J.; Li, W.; Liu, J.; Xie, R.; Wang, Y.; Liu, S.;
Articular Cartilage Repair. Tissue Eng. 2001, 7 (6), 767−780. Wang, D.-A.; Ren, L. Novel β-TCP/PVA bilayered hydrogels with
(150) Pförringer, D.; Harrasser, N.; Mühlhofer, H.; Kiokekli, M.; considerable physical and bio-functional properties for osteochondral
Stemberger, A.; van Griensven, M.; Lucke, M.; Burgkart, R.; repair. Biomedical Materials 2018, 13 (1), 015012.
Obermeier, A. Osteoinduction and -conduction through absorbable (166) Bittner, S. M.; Smith, B. T.; Diaz-Gomez, L.; Hudgins, C. D.;
bone substitute materials based on calcium sulfate: in vivo biological Melchiorri, A. J.; Scott, D. W.; Fisher, J. P.; Mikos, A. G. Fabrication
behavior in a rabbit model. J. Mater. Sci.: Mater. Med. 2018, 29 (2), and mechanical characterization of 3D printed vertical uniform and
17. gradient scaffolds for bone and osteochondral tissue engineering. Acta
(151) Thomas, M. V.; Puleo, D. A. Calcium sulfate: Properties and Biomater. 2019, 90, 37−48.
clinical applications. J. Biomed. Mater. Res., Part B 2009, 88B (2), (167) Deliormanlı, A. M.; Atmaca, H. Biological Response of
597−610. Osteoblastic and Chondrogenic Cells to Graphene-Containing PCL/
(152) Dell’Osso, G.; Bottai, V.; Bugelli, G.; Manisco, T.; Cazzella, Bioactive Glass Bilayered Scaffolds for Osteochondral Tissue
N.; Celli, F.; Guido, G.; Giannotti, S. The biphasic bioresorbable Engineering Applications. Appl. Biochem. Biotechnol. 2018, 186 (4),
scaffold (Trufit((R))) in the osteochondral knee lesions: long-term 972−989.
clinical and MRI assessment in 30 patients. Musculoskelet Surgery (168) Filardo, G.; Perdisa, F.; Gelinsky, M.; Despang, F.; Fini, M.;
2016, 100 (2), 93−6. Marcacci, M.; Parrilli, A. P.; Roffi, A.; Salamanna, F.; Sartori, M.;
(153) Dhollander, A. A.; Liekens, K.; Almqvist, K. F.; Verdonk, R.; Schütz, K.; Kon, E. Novel alginate biphasic scaffold for osteochondral
Lambrecht, S.; Elewaut, D.; Verbruggen, G.; Verdonk, P. C. A pilot regeneration: an in vivo evaluation in rabbit and sheep models. J.
study of the use of an osteochondral scaffold plug for cartilage repair Mater. Sci.: Mater. Med. 2018, 29 (6), 74.
in the knee and how to deal with early clinical failures. Arthroscopy (169) Kang, H.; Zeng, Y.; Varghese, S. Functionally graded
2012, 28 (2), 225−33. multilayer scaffolds for in vivo osteochondral tissue engineering.
(154) Joshi, N.; Reverte-Vinaixa, M.; Díaz-Ferreiro, E. W.; Acta Biomater. 2018, 78, 365−377.
Domínguez-Oronoz, R. Synthetic Resorbable Scaffolds for the (170) Erickson, A. E.; Sun, J.; Lan Levengood, S. K.; Swanson, S.;
Treatment of Isolated Patellofemoral Cartilage Defects in Young Chang, F.-C.; Tsao, C. T.; Zhang, M. Chitosan-based composite
Patients:Magnetic Resonance Imaging and Clinical Evaluation. bilayer scaffold as an in vitro osteochondral defect regeneration
American Journal of Sports Medicine 2012, 40 (6), 1289−1295. model. Biomed. Microdevices 2019, 21 (2), 34.
(155) Verhaegen, J.; Clockaerts, S.; Van Osch, G.; Somville, J.; (171) Lin, T.-H.; Wang, H.-C.; Cheng, W.-H.; Hsu, H.-C.; Yeh, M.-
Verdonk, P.; Mertens, P. TruFit Plug for Repair of Osteochondral L. Osteochondral Tissue Regeneration Using a Tyramine-Modified
DefectsWhere Is the Evidence? Systematic Review of Literature. Bilayered PLGA Scaffold Combined with Articular Chondrocytes in a
Cartilage 2015, 6 (1), 12−19. Porcine Model. Int. J. Mol. Sci. 2019, 20 (2), 326.
(156) Neto, A. S.; Ferreira, J. M. F. Synthetic and Marine-Derived (172) Maciulaitis, J.; Miskiniene, M.; Rekštytė, S.; Bratchikov, M.;
Porous Scaffolds for Bone Tissue Engineering. Materials 2018, 11 (9), Darinskas, A.; Simbelyte, A.; Daunoras, G.; Laurinaviciene, A.;
1702. Laurinavicius, A.; Gudas, R.; Malinauskas, M.; Maciulaitis, R.
(157) Matta, C.; Szű cs-Somogyi, C.; Kon, E.; Robinson, D.; Neufeld, Osteochondral Repair and Electromechanical Evaluation of Custom
T.; Altschuler, N.; Berta, A.; Hangody, L.; Veréb, Z.; Zákány, R. 3D Scaffold Microstructured by Direct Laser Writing Lithography.
Osteogenic differentiation of human bone marrow-derived mesen- Cartilage 2019, 0 (00), 1−11.
chymal stem cells is enhanced by an aragonite scaffold. Differentiation (173) Mahapatra, C.; Kim, J.-J.; Lee, J.-H.; Jin, G.-Z.; Knowles, J. C.;
2019, 107, 24−34. Kim, H.-W. Differential chondro- and osteo-stimulation in three-
(158) Kon, E.; Filardo, G.; Shani, J.; Altschuler, N.; Levy, A.; Zaslav, dimensional porous scaffolds with different topological surfaces
K.; Eisman, J. E.; Robinson, D. Osteochondral regeneration with a provides a design strategy for biphasic osteochondral engineering. J.
novel aragonite-hyaluronate biphasic scaffold: up to 12-month follow- Tissue Eng. 2019, 10, 204173141982643.
up study in a goat model. Journal of Orthopaedic Surgery and Research (174) Wu, X. Biphasic osteochondral scaffold fabrication using
2015, 10, 81−81. multi-material mask projection stereolithography. Rapid Prototyping
(159) CartiHeal Announces FDA IDE Approval of its Agili-C Implant Journal 2019, 25 (2), 277−288.
for the Treatment of Joint Surface Lesions. https://www.cartiheal.com/ (175) Bhardwaj, N.; Kundu, S. C. Electrospinning: A fascinating
news/cartiheal-announces-fda-ide-approval-of-its-agili-c-implant-for- fiber fabrication technique. Biotechnol. Adv. 2010, 28 (3), 325−347.
the-treatment-of-joint-surface-lesions/ (accessed October 31, 2019). (176) Casanova, M. R.; Reis, R. L.; Martins, A.; Neves, N. M. The
(160) Du, Y.; Liu, H.; Yang, Q.; Wang, S.; Wang, J.; Ma, J.; Noh, I.; Use of Electrospinning Technique on Osteochondral Tissue
Mikos, A. G.; Zhang, S. Selective laser sintering scaffold with Engineering. In Osteochondral Tissue Engineering: Nanotechnology,
hierarchical architecture and gradient composition for osteochondral Scaffolding-Related Developments and Translation; Oliveira, J. M., Pina,
repair in rabbits. Biomaterials 2017, 137, 37−48. S., Reis, R. L., San Roman, J., Eds.; Springer International Publishing:
(161) Liao, J.; Tian, T.; Shi, S.; Xie, X.; Ma, Q.; Li, G.; Lin, Y. The Cham, 2018; pp 247−263. DOI: 10.1007/978-3-319-76711-6_11.
fabrication of biomimetic biphasic CAN-PAC hydrogel with a (177) Liu, Y.; Zhang, X.; Xia, Y.; Yang, H. Magnetic-Field-Assisted
seamless interfacial layer applied in osteochondral defect repair. Electrospinning of Aligned Straight and Wavy Polymeric Nanofibers.
Bone Res. 2017, 5, 17018. Adv. Mater. 2010, 22 (22), 2454−2457.

4335 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336
ACS Biomaterials Science & Engineering pubs.acs.org/journal/abseba Perspective

(178) Tindell, K. R.; Holloway, J.; Ronken, S. In Controlling fiber defect using a microfluidic printing head. Biofabrication 2019, 11 (4),
alignment using magnetically-assisted electrospinning for interfacial 044101.
tissue repair, Society for Biomaterials Annual Meeting and Exposition (196) Skoog, S. A.; Goering, P. L.; Narayan, R. J. Stereolithography
2019: The Pinnacle of Biomaterials Innovation and Excellence - in tissue engineering. J. Mater. Sci.: Mater. Med. 2014, 25 (3), 845−
Transactions of the 42nd Annual Meeting, Seattle, WA, Transactions of 856.
the Annual Meeting of the Society for Biomaterials and the Annual (197) Wicker, R. B.; MacDonald, E. W. Multi-material, multi-
International Biomaterials Symposium: Seattle, WA, 2019. technology stereolithography. Virtual and Physical Prototyping 2012, 7
(179) Balu, R.; Singaravelu, S.; Nagiah, N. Bioceramic Nanofibres by (3), 181−194.
Electrospinning. Fibers 2014, 2 (3), 221−239. (198) Raimondi, M. T.; Eaton, S. M.; Nava, M. M.; Laganà, M.;
(180) Zhang, Y.; Liu, X.; Zeng, L.; Zhang, J.; Zuo, J.; Zou, J.; Ding, Cerullo, G.; Osellame, R. Two-Photon Laser Polymerization: From
J.; Chen, X. Polymer Fiber Scaffolds for Bone and Cartilage Tissue Fundamentals to Biomedical Application in Tissue Engineering and
Engineering. Adv. Funct. Mater. 2019, 29 (36), 1903279. Regenerative Medicine. J. Appl. Biomater. Funct. Mater. 2012, 10 (1),
(181) Yunos, D. M.; Ahmad, Z.; Salih, V.; Boccaccini, A. R. Stratified 56−66.
scaffolds for osteochondral tissue engineering applications: Electro- (199) Ç akmak, S.; Ç akmak, A. S.; Kaplan, D. L.; Gümüsḑ erelioğlu,
spun PDLLA nanofibre coated Bioglass®-derived foams. J. Biomater. M. A Silk Fibroin and Peptide Amphiphile-Based Co-Culture Model
Appl. 2013, 27 (5), 537−551. for Osteochondral Tissue Engineering. Macromol. Biosci. 2016, 16 (8),
(182) Gacasan, E. G.; Sehnert, R. M.; Ehrhardt, D. A.; Grunlan, M. 1212−1226.
A. Templated, Macroporous PEG-DA Hydrogels and Their Potential (200) Amadori, S.; Torricelli, P.; Panzavolta, S.; Parrilli, A.; Fini, M.;
Utility as Tissue Engineering Scaffolds. Macromol. Mater. Eng. 2017, Bigi, A. Multi-Layered Scaffolds for Osteochondral Tissue Engineer-
ing: In Vitro Response of Co-Cultured Human Mesenchymal Stem
302, 1600512.
Cells. Macromol. Biosci. 2015, 15 (11), 1535−1545.
(183) Woodard, L. N.; Kmetz, K. T.; Roth, A. A.; Page, V. M.;
(201) Chen, K.; Ng, K. S.; Ravi, S.; Goh, J. C. H.; Toh, S. L. In vitro
Grunlan, M. A. Porous Poly(ε-caprolactone)-Poly(l-lactic acid) Semi-
generation of whole osteochondral constructs using rabbit bone
Interpenetrating Networks as Superior, Defect-Specific Scaffolds with
marrow stromal cells, employing a two-chambered co-culture well
Potential for Cranial Bone Defect Repair. Biomacromolecules 2017, 18 design. J. Tissue Eng. Regener. Med. 2016, 10 (4), 294−304.
(12), 4075−4083. (202) Yang, J.; Zhang, Y. S.; Yue, K.; Khademhosseini, A. Cell-laden
(184) Scaffaro, R.; Lopresti, F.; Botta, L.; Rigogliuso, S.; Ghersi, G. hydrogels for osteochondral and cartilage tissue engineering. Acta
Integration of PCL and PLA in a monolithic porous scaffold for Biomater. 2017, 57, 1−25.
interface tissue engineering. Journal of the Mechanical Behavior of (203) Chang, C.-H.; Lin, C.-C.; Chou, C.-H.; Lin, F.-H.; Liu, H.-C.
Biomedical Materials 2016, 63, 303−313. Novel bioreactors for osteochondral tissue engineering. Biomed. Eng.
(185) Tang, G.; Zhang, H.; Zhao, Y.; Zhang, Y.; Li, X.; Yuan, X. 2005, 17 (01), 38−43.
Preparation of PLGA Scaffolds with Graded Pores by Using a Gelatin- (204) Hansmann, J.; Groeber, F.; Kahlig, A.; Kleinhans, C.; Walles,
Microsphere Template as Porogen. J. Biomater. Sci., Polym. Ed. 2012, H. Bioreactors in tissue engineeringprinciples, applications and
23 (17), 2241−2257. commercial constraints. Biotechnol. J. 2013, 8 (3), 298−307.
(186) Deville, S.; Saiz, E.; Nalla, R. K.; Tomsia, A. P. Freezing as a (205) Li, K.; Zhang, C.; Qiu, L.; Gao, L.; Zhang, X. Advances in
Path to Build Complex Composites. Science 2006, 311 (5760), 515. Application of Mechanical Stimuli in Bioreactors for Cartilage Tissue
(187) Grenier, J.; Duval, H.; Barou, F.; Lv, P.; David, B.; Letourneur, Engineering. Tissue Eng., Part B 2017, 23 (4), 399−411.
D. Mechanisms of Pore Formation in Hydrogel Scaffolds Textured by (206) Martin, Y.; Eldardiri, M.; Lawrence-Watt, D. J.; Sharpe, J. R.
Freeze-Drying. Acta Biomater. 2019, 94, 195. Microcarriers and Their Potential in Tissue Regeneration. Tissue Eng.,
(188) Annabi, N.; Nichol, J. W.; Zhong, X.; Ji, C.; Koshy, S.; Part B 2011, 17 (1), 71−80.
Khademhosseini, A.; Dehghani, F. Controlling the Porosity and (207) Zhou, A.; Ye, Z.; Zhou, Y.; Tan, W.-s. Bioactive poly(ε-
Microarchitecture of Hydrogels for Tissue Engineering. Tissue Eng., caprolactone) microspheres with tunable open pores as microcarriers
Part B 2010, 16 (4), 371−383. for tissue regeneration. J. Biomater. Appl. 2019, 33 (9), 1242−1251.
(189) Haugh, M. G.; Murphy, C. M.; O’Brien, F. J. Novel Freeze- (208) Carvalho, M. R.; Reis, R. L.; Oliveira, J. M. Mimicking the 3D
Drying Methods to Produce a Range of Collagen−Glycosaminoglycan biology of osteochondral tissue with microfluidic-based solutions:
Scaffolds with Tailored Mean Pore Sizes. Tissue Eng., Part C 2010, 16 breakthroughs towards boosting drug testing and discovery. Drug
(5), 887−894. Discovery Today 2018, 23 (3), 711−718.
(190) Pawelec, K. M.; Husmann, A.; Best, S. M.; Cameron, R. E. (209) Beddoes, C. M.; Whitehouse, M. R.; Briscoe, W. H.; Su, B.
Understanding anisotropy and architecture in ice-templated biopol- Hydrogels as a Replacement Material for Damaged Articular Hyaline
ymer scaffolds. Mater. Sci. Eng., C 2014, 37, 141−147. Cartilage. Materials 2016, 9 (6), 443.
(191) An, J.; Teoh, J. E. M.; Suntornnond, R.; Chua, C. K. Design (210) Gong, J. P.; Katsuyama, Y.; Kurokawa, T.; Osada, Y. Double-
and 3D Printing of Scaffolds and Tissues. Engineering 2015, 1 (2), Network Hydrogels with Extremely High Mechanical Strength. Adv.
261−268. Mater. 2003, 15 (14), 1155−1158.
(192) Jammalamadaka, U.; Tappa, K. Recent Advances in (211) Arnold, M. P.; Daniels, A. U.; Ronken, S.; García, H. A.;
Biomaterials for 3D Printing and Tissue Engineering. J. Funct. Friederich, N. F.; Kurokawa, T.; Gong, J. P.; Wirz, D. Acrylamide
Biomater. 2018, 9 (1), 22. Polymer Double-Network Hydrogels: Candidate Cartilage Repair
(193) Daly, A. C.; Freeman, F. E.; Gonzalez-Fernandez, T.; Materials with Cartilage-Like Dynamic Stiffness and Attractive
Critchley, S. E.; Nulty, J.; Kelly, D. J. 3D Bioprinting for Cartilage Surgery-Related Attachment Mechanics. Cartilage 2011, 2 (4),
and Osteochondral Tissue Engineering. Adv. Healthcare Mater. 2017, 374−383.
6 (22), 1700298. (212) Means, A. K.; Ehrhardt, D. A.; Whitney, L. V.; Grunlan, M. A.
(194) Radhakrishnan, J.; Subramanian, A.; Krishnan, U. M.; Thermoresponsive Double Network Hydrogels with Exceptional
Sethuraman, S. Injectable and 3D Bioprinted Polysaccharide Hydro- Compressive Mechanical Properties. Macromol. Rapid Commun.
gels: From Cartilage to Osteochondral Tissue Engineering. Bio- 2017, 38 (20), 1700351.
(213) Means, A. K.; Shrode, C. S.; Whitney, L. V.; Ehrhardt, D. A.;
macromolecules 2017, 18 (1), 1−26.
Grunlan, M. A. Double Network Hydrogels that Mimic the Modulus,
(195) Idaszek, J.; Costantini, M.; Karlsen, T. A.; Jaroszewicz, J.;
Strength, and Lubricity of Cartilage. Biomacromolecules 2019, 20 (5),
Colosi, C.; Testa, S.; Fornetti, E.; Bernardini, S.; Seta, M.; Kasarełło,
2034−2042.
K.; Wrzesień, R.; Cannata, S.; Barbetta, A.; Gargioli, C.; Brinchman, J.
E.; Świȩszkowski, W. 3D bioprinting of hydrogel constructs with cell
and material gradients for the regeneration of full-thickness chondral

4336 https://dx.doi.org/10.1021/acsbiomaterials.0c00753
ACS Biomater. Sci. Eng. 2020, 6, 4324−4336