Advanced Drug Delivery Reviews 95 (2015) 15–28

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Advanced Drug Delivery Reviews

journal homepage: www.elsevier.com/locate/addr

Integration of mesenchymal stem cells with nanobiomaterials for the

repair of myocardial infarction☆
Jin Han a,1, Jooyeon Park a,1, Byung-Soo Kim a,b,⁎
a
School of Chemical and Biological Engineering, Seoul National University, Seoul 151-744, Republic of Korea
b
Interdisciplinary Program of Bioengineering, Bio-MAX Institute, Institute of Chemical Processes, Seoul National University, Seoul 151-744, Republic of Korea

a r t i c l e i n f o a b s t r a c t

Article history: The integration of nanobiomaterials with stem cells represents a promising strategy for the treatment of myocar-
Received 1 June 2015 dial infarction. While stem cells and nanobiomaterials each demonstrated partial success in cardiac repair indi-
Received in revised form 27 August 2015 vidually, the therapeutic efficacy of the clinical settings for each of these has been low. Hence, a combination of
Accepted 10 September 2015
nanobiomaterials with stem cells is vigorously studied to create synergistic effects for treating myocardial infarc-
Available online 24 September 2015
tion. To date, various types of nanomaterials have been incorporated with stem cells to control cell fate, modulate
Keywords:
the therapeutic behavior of stem cells, and make them more suitable for cardiac repair. Here, we review the cur-
Stem cells rent stem cell therapies for cardiac repair and describe the combinatorial approaches of using nanobiomaterials
Biomaterials and stem cells to improve therapeutic efficacy for the treatment of myocardial infarction.
Myocardial infarction © 2015 Elsevier B.V. All rights reserved.
Nanomaterials
Regenerative medicine
Tissue engineering

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16
2. Mesenchymal stem cells (MSCs) for the treatment of myocardial infarction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16
2.1. Differentiation of MSCs into cardiac lineage cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16
2.2. Paracrine factor secretion of MSCs for cardiac repair . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
3. Nanobiomaterial-incorporated stem cell therapy for cardiac repair. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
3.1. Therapeutic molecule delivery systems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
3.2. Cell delivery vehicles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
3.3. Nanotopographical cues of nanobiomaterials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.4. Electrically conductive nanobiomaterials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
3.5. Intrinsic chemistry of nanobiomaterials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
4. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

Abbreviations: MI, myocardial infarction; MSC, mesenchymal stem cell; ESC, embryonic stem cell; iPSC, induced pluripotent stem cell; CSC, cardiac stem cell; PDGF-BB, platelet-derived
growth factor-BB; PlGF, placental growth factor; VEGF, vascular endothelial growth factor; SDF-1, stromal cell-derived factor 1; IGF-1, insulin growth factor 1; Cx43, connexin 43; MNC,
mononuclear cell; CMC, cardiomyocytes; ASC, adipose-derived stem cell; PLCL, poly(lactide-co-ε-caprolactone); PCL, polycaprolactone; PEI, polyethylenimine; PEG, polyethylene glycol;
BSA, bovine serum albumin; PVA, polyvinyl alcohol.
☆ This review is part of the Advanced Drug Delivery Reviews theme issue on "Multifunctional Nanodevices and Nanobots for Bioimaging, Cancer Diagnosis/Therapy, Stem Cell Therapy,
and Regenerative Medicine".
⁎ Corresponding author at: School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 151-744, Republic of Korea. Tel.: +82 2 880 1875;
fax: +82 2 2297 0838.
E-mail address: byungskim@snu.ac.kr (B.-S. Kim).
1
These authors contributed equally.

http://dx.doi.org/10.1016/j.addr.2015.09.002
0169-409X/© 2015 Elsevier B.V. All rights reserved.
16 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
1. Introduction
Cardiovascular diseases remain one of the leading causes of death
worldwide [1,2]. Over 80 million individuals in United States have one
or more types of cardiovascular diseases [2]. Myocardial infarction
(MI), in particular, serves as the major contributor to heart failure and
cardiac-associated disorders [1,3,4]. Previously, thrombolytic treat-
ments or surgical coronary artery bypass were introduced to treat MI;
however, these therapies merely promoted cardiac tissue regeneration
[5–7]. MI is characterized by reduced blood supply to the heart, which
results in cardiac cell death, tissue damage, ventricular malfunction,
and heart failure [4]. Hence, successful treatments for MI depend on sal-
vaging dying cells, recovering blood vessels, and regenerating cardiac
tissues [4,7]. In other words, cardiac tissue engineering approaches
that can prevent cell death and regenerate cardiac tissues are required
for effective treatment of MI.
Recently, significant advances have been made in the use of stem
cells for the treatment of MI [8–10] and a number of clinical trials
with stem cells showed encouraging results with cardiac function
restoration. Stem cells have regenerative potentials to replace the
dead cardiomyocytes, salvage dying cells, and induce angiogenesis
at the infarcted region [7]. A number of different cell types, including
mesenchymal stem cells (MSCs) [11–14], emb ryonic stem cells
(ESCs) [15–17], induced pluripotent stem cells (iPSCs) [18–20], and
cardiac stem cells (CSCs) [21], all showed salutary effects in repairing
injured myocardium. Among the various cell types, MSCs represent a
reliable source for cell therapy as they have shown safe and promis-
ing results in long-term clinical trials [12,14,22]. Upon implantation
into the infarcted heart, a fraction of MSCs differentiates into func-
tional cardiomyocytes and replaces the damaged cardiac tissues to
improve heart function [23–25]. Additionally, MSCs secrete various
therapeutic paracrine molecules for cardiac cell protection, angio-
genesis, and immune modulation for cardiac repair [1,26–28]. How-
ever, low cell survival, engraftment, and differentiation of stem cells
after implantation impede the therapeutic effect of the stem cells.
Hence, other treatment options are vigorously investigated for better
cardiac repair.
Nanobiomaterial-based treatments for MI recently drew significant
attention as new therapeutic approaches for cardiac repair [29,30].
Nanobiomaterials represent natural or synthetic biocompatible mate-
rials with nanoscale characteristics, which further provide exclusive
physicochemical properties such as nanoscale size, nanotopographical
cues, electrical conductivity, or distinct chemical features. Treatments
that utilize nanobiomaterials, including nanoparticles, nanofibrous
scaffolds, self-assembling peptides, and many others, showed signifi-
cant effects in repairing injured myocardium [29]. A number of
nanobiomaterials were previously used to deliver therapeutic mole-
cules including proteins, drugs, and genes for cardiac repair [31–35]. Ad-
ditionally, biomaterials utilized as cell carriers have improved the
therapeutic efficacy of cells for the treatment of MI [36,37]. More recent-
ly, nanobiomaterials that can modulate cell functions have provided a
new window for nanobiomaterial-incorporated cell therapy for MI,
and materials with unique topographical, electrical, or chemical charac-
teristics have been extensively studied to create synergistic effects with
cell therapy. Overall, the combination of conventional stem cell therapy
and nanobiomaterials propose a great therapeutic option for treating MI
[38–51].
Here, we review the current status of tissue regenerative approaches
for treating MI, particularly stem cell therapy, nanobiomaterial-based
biomolecule delivery, and nanobiomaterial-integrated stem cell thera-
py. We first review the mechanisms and recent advances in stem cell
therapy for MI. Next, we introduce nanobiomaterials with the potential
to efficiently deliver therapeutic molecules or cells for the treatment of
MI. Finally, we discuss biomaterials with unique topographical, electri-
cal, or chemical characteristics that can improve the therapeutic efficacy
of stem cells for cardiac repair.
2. Mesenchymal stem cells (MSCs) for the treatment of
myocardial infarction
A number of preclinical and clinical studies have been performed
using stem cells to repair damaged hearts [15,18,21,52,53]. Most of
these cells, including MSCs, ESCs, iPSCs, and CSCs, improved cardiac
function, suggesting that the implantation of stem cells is a promising
therapeutic approach for MI repair. ESCs and iPSCs represent ideal cell
sources for MI repair due to their pluripotency. ESCs can differentiate
into functional cardiomyocytes when implanted into the injured myo-
cardium [16,17]. Differentiation of ESCs at the lesion site further restores
contractile function of the heart and attenuates cardiac remodeling [15].
Similarly, the implantation of iPSCs into the injured myocardium results
in cell engraftment and differentiation into cardiomyocytes, which leads
to enhanced cardiac function [18–20]. Despite their therapeutic poten-
tial, ESCs pose ethical concerns and potential teratogenic risks while
the generation efficiency of iPSCs is still very low [54,55]. As an alterna-
tive, CSCs have attracted attention as a cell source with great cardiac dif-
ferentiation potential which can attribute to the repair of the damaged
heart. CSCs are c-kit-positive heart-resident stem cells that can differen-
tiate into cardiac lineage cells [55]. However, an extremely small
amount of CSCs within a heart and the difficulties in obtaining these
cells limit their clinical applications despite their high therapeutic effi-
cacy [55]. Meanwhile, MSCs have been extensively investigated as the
optimal cell source for MI. The cardiac differentiation potential of
MSCs is comparatively limited compared to ESCs, iPSCs, and CSCs. How-
ever, MSCs can easily be isolated from patients, expanded ex vivo, and
utilized in an autologous manner [7]. Additionally, the secretion of ther-
apeutic paracrine molecules from MSCs, including various types of
growth factors and cytokines, is known to repair the injured myocardi-
um [1,56–60]. The therapeutic efficacy of MSCs has been exhibited in a
number of clinical studies. The POSEIDON trial demonstrated that the
implantation of autologous or allogenic human MSCs can improve car-
diac function after MI [52]. Additionally, the APOLLO trial showed en-
hancements in cardiac function, perfusion, and neovasculogenesis
after the transplantation of adipose-derived stem cells [53]. Various
other trials, including the ANGEL, ATHENA, and MyStromal Cell, are cur-
rently undergoing clinical phase trials for chronic myocardial ischemia
[8]. In the following sections, we review the two major therapeutic
mechanisms of MSC therapy for MI, namely, (1) differentiation of
MSCs into cardiac lineage and (2) paracrine factor secretion of MSCs
(Fig. 1).
2.1. Differentiation of MSCs into cardiac lineage cells
Direct differentiation of the implanted MSCs into cardiac lineage
cells was originally proposed as the principal mechanism of stem cell
therapy for MI [7]. Yoon et al. showed that MSCs implanted to the in-
farcted myocardium differentiated into cardiomyocytes, endothelial
cells, and smooth muscle cells, and consequently improved cardiac
function [61]. Differentiation of adipose-derived and cortical bone-
derived stem cells into cardiomyocytes and vascular cells was observed
after in vivo implantation [56,62]. However, the fraction of MSCs that
differentiated into cardiomyocytes after in vivo implantation is relative-
ly low. Hence, many attempts were made to differentiate MSCs toward
cardiac lineages in vitro prior to in vivo implantation, with the help of
chemicals, small molecules, growth factors, and co-culture with cardiac
cells, to improve the therapeutic efficacy of MSCs [63]. To differentiate
MSCs into cardiomyocytes using chemicals, 5-azacytidine has been
widely utilized as the key chemical facilitator [64,65]. It was also report-
ed that the fluid shear stress in combination with 5-azacytidine treat-
ment further enhanced cardiomyogenic differentiation of MSCs [66].
In addition to 5-azacytidine, small molecules such as phorbol myristate
acetate can also promote cardiomyogenic differentiation of MSCs, and
stimulate electromechanical integration of MSCs with resident
cardiomyocytes [67]. Bartunek et al. further showed that pretreatment
 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
Fig. 1. Therapeutic mechanisms involved in stem cell therapy for the treatment of MI. Stem cells can either differentiate into cardiomyocytes or secrete therapeutic paracrine molecules to
salvage and regenerate heart tissue.
of MSCs with cardiomyogenic growth factors prior to implantation
improves cardiac lineage differentiation of MSCs in vivo and en-
hances the functional recovery of a chronically infarcted myocardi-
um [68]. The co-culture of MSCs with cardiac cells also promotes
cardiomyogenesis of MSCs, which can be further promoted by
pretreating the cardiac cells with iron oxide nanoparticles [51] or
by the alignment of the cells [69]. Once implanted into the damaged
heart, MSC-differentiated cardiomyocytes can reduce fibrosis, im-
prove angiogenesis, preserve gap junctions, and enhance cardiac
functions compared to the undifferentiated MSCs [51,67,68].
2.2. Paracrine factor secretion of MSCs for cardiac repair
Even though the cardiac function improvements after MSC implan-
tation were accompanied with the differentiation of MSCs, a previous
study addressed that the number of newly differentiated or generated
cardiomyocytes was too small to explain the benefits of MSC therapy
[70]. After MI, more than a billion cardiomyocytes undergo apoptosis
and necrosis; hence, the small fraction of MSC-differentiated
cardiomyocytes may not elicit any meaningful benefit for attenuating
cardiac remodeling post-MI. Growing evidence now suggests that
growth factors and cytokines secreted from the implanted MSCs play
a critical role in promoting cardiac repair [56,60]. MSCs secrete a variety
of growth factors and cytokines that exert protective effects on dam-
aged tissues and organs [57–59]. These paracrine molecules are in-
volved in cardiac tissue regeneration, neovascularization, cardiac
contractility, cardiac metabolism, myocardial protection, and even im-
mune responses for cardiac repair [71].
The therapeutic effects of paracrine molecules for cardiac repair
were demonstrated in the studies where the administration of condi-
tioned medium of stem cell cultures recapitulated the benefits of stem
cell implantation [27,70]. Gnecchi et al. demonstrated that the condi-
tioned medium from MSC cultures showed cytoprotective effects on
rat cardiomyocytes in vitro and significantly reduced cardiac fibrosis
in vivo [27]. Furthermore, paracrine factors secreted by the implanted
MSCs protected adjacent cardiomyocytes from undergoing cell death,
attenuated left ventricular remodeling in an ischemic heart, and in-
duced angiogenesis and vasculogenesis in vivo [28]. To enhance the
paracrine secretion of MSCs and elicit better tissue regeneration, previ-
ous studies introduced hypoxic culture conditions or MSC spheroids
generating mild hypoxia on in these cells [51,72–75]. It was also demon-
strated that hypoxia preconditioning of MSCs in vitro not only
17
stimulates their paracrine molecule secretion, but also results in their
enhanced survival in the infarcted myocardium [26].
These observations regarding the cardiac differentiation capability
and paracrine action of MSCs prompted the development of new thera-
peutic strategies that exert synergistic effects with these cells. Recently,
nanobiomaterials that improve the cardiac differentiation and harness
the paracrine action of MSCs have gained much attention. The advances
in nanobiomaterial-integrated stem cell therapy for cardiac repair are
reviewed in the following sections.
3. Nanobiomaterial-incorporated stem cell therapy for
cardiac repair
Mounting evidence for stem cell–derived paracrine action and stem
cells’ cardiac lineage differentiation for cardiac repair has introduced a
crucial step forward for the development of biomaterial-integrated
stem cell therapies. Recently, studies utilizing the nanoscale properties
of biomaterials to potentiate the therapeutic efficacy of stem cells
have made significant advances (Table 1). Biocompatible nanomaterials
can deliver therapeutic molecules for angiogenesis and stem cell re-
cruitment, both of which promote cardiac repair. Additionally, integra-
tion of nanobiomaterials with stem cells may exploit synergistic
effects for cardiac repair by overcoming the limitations of conventional
stem cell therapy. Nanoscale properties of nanobiomaterials, such as
nanotopography, electrical conductivity or unique chemical characteris-
tics, may modulate stem cells’ behaviors needed for effective MI repair,
including their structural alignment, cardiac differentiation, and para-
crine secretion. Additionally, nanoscale properties can also be used to
deliver therapeutic molecules or cells for better treatment of MI. In the
following sections, we provide an overview of the incorporation of
nanobiomaterials with stem cells for improved cardiac repair after MI
(Fig. 2).
3.1. Therapeutic molecule delivery systems
The delivery of therapeutic molecules, including genes and proteins,
can improve cardiac repair after MI by promoting angiogenesis [34,76]
or recruiting stem cells [77]. To maximize the therapeutic effect of
these biomolecules, sustained and controlled release of the drug is cru-
cial. Therefore, previous studies utilized nanobiomaterials to prolong
the expression of proteins for improved angiogenesis and recruitment
of stem cells [31,32,35,78]. Research showed that the delivery of
platelet-derived growth factor-BB (PDGF-BB) using self-assembling
18 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28

Table 1
Results of biomaterials and biomaterials-integrated stem cell therapy for MI in animals or in vitro studies.

Approach and materials Cells or Model Delivery Time Major observations (in vitro and in vivo)
biomolecules method point
(in vivo)

Protein or gene delivery materials

Chitosan-alginate nanoparticle [31] PlGF Rat Intramyocardial 8 weeks Heart function ↑, vascular density ↑, scar formation ↓,
injection inflammation ↓
Self-assembling peptide nanofiber [35] PDGF-BB Rat Intramyocardial 2 weeks CMC death ↓, systolic function ↑, infarct size ↓, contractility ↑
(AcN-RARADADARARADADA-CNH2) injection
Baculovirus nanocomplex [91] Angiopoeitin-1 Rat Intramyocardial 4 weeks Capillary density ↑, infarct size ↓, heart function ↑
with ASC injection

Cell delivery materials

Self-assembling peptide [37] CSC Mouse Intramyocardial 2 weeks Heart function ↑, fibrosis ↓, cardiac differentiation ↑
(AcN-RADARADARADARADA-CNH2) injection
Collagen patch [97] MSC Rat Epicardial 4 weeks MSC engraftment ↑, wall thickness ↑, heart function ↑
surface patch
PLCL patch [98] MSC Rat Epicardial 4 weeks Heart function ↑, MSC cardiac differentiation ↑, infarct size ↓,
surface patch
Self-assembling peptide nanofiber [101] MNC Pig Intramyocardial 4 weeks Heart function ↑, engraftment ↑, capillary density ↑
(AcN-RARADADARARADADA-NH2) injection
Self-assembling peptide nanofiber [103] MSC Rat Intramyocardial 4 weeks Heart function ↑, infarct size ↓, capillary density ↑,
(AcN-RARADADARARADADA-NH2) injection
Self-assembling peptide nanofiber [104] CSC Rat Intramyocardial 4 weeks Vascular enlargement ↑, heart function ↑, capillary density ↑
(AcN-RADARADARADARADA-CNH2) injection

Nanomaterials with topographical cues

Patterned PEG hydrogel [41] CSC Rat Epicardial 3 weeks Wall thickness ↑, organized collagen deposition ↑, surviving
surface patch muscle cells ↑,
Microelectrode arrays [69] (in vitro study) MSC and CMC N/A N/A N/A MSC conduction velocity ↑, MSC gap junction protein ↑
co-culture
PCL nanofiber [106] MSC Rat Epicardial 4 weeks Heart function ↑, fibrosis ↓, wall thickness ↑
surface patch

Nanomaterials with electrical conductivity

Carbon nanotube [47] (in vitro study) MSC N/A N/A N/A MSC cardiac-specific and gap junction protein ↑
Gold nanoparticle embedded BSA/PVA scaffold MSC with N/A N/A N/A MSC cardiac-specific and gap junction protein ↑
[111] (in vitro study) 5-azacytidine
Reduced graphene oxide [112] MSC spheroids Mouse Intramyocardial 2 weeks MSC gap junction protein ↑, heart function ↑
injection

Nanomaterials with chemical cues

Iron oxide nanoparticle [51] MSC and H9C2 Rat Intramyocardial 2 weeks MSC cardiac-specific and gap junction protein ↑, MSC paracrine ↑,
co-culture injection heart function ↑, capillary density ↑
Self-assembling peptide nanofiber [114] MSC-conditioned Mouse Intramyocardial 4 weeks Contractility ↑, vascularization ↑, heart function ↑
(Not specified) medium injection
Graphene [115] (in vitro study) MSC N/A N/A N/A MSC cardiac-specific and gap junction protein ↑
Graphene [121] (in vitro study) ESC N/A N/A N/A ESC cardiac differentiation ↑

Peptide sequence: R, arginine; A, alanine; D, aspartic acid.

peptide nanofibers [35] or placental growth factor (PlGF) using
chitosan-alginate nanoparticles [31] significantly attenuated cardiac re-
modeling and improved cardiac function after MI. PDGF-BB is a well-
known protein that promotes angiogenesis [79]; however, PDGF-BB
injected in vivo is rapidly cleared from the blood [80] and results in
low therapeutic efficacy. To overcome this problem and to prevent
rapid clearance of the growth factor in vivo, Hsieh et al. utilized self-
assembling peptides in protein delivery [35]. Self-assembling peptides
are composed of short peptide sequences linked to a fatty acid tail,
and can spontaneously assemble into nanofibers under various physio-
logical conditions. These peptides are biodegradable and suitable for
functional modifications, making them a good candidate for the treat-
ment of MI. The study showed that PDGF-BB delivered without self-
assembling peptides rapidly disappeared from the injected site after
24 h, and only a negligible amount of this protein was detected after
3 days. Conversely, self-assembling peptide nanofibers loaded with
PDGF-BB achieved a sustained release of PDGF-BB at the injection sites
for more than 14 days. This is mainly because PDGF-BB was tightly
bound to the self-assembling peptide nanofibers, which served as a res-
ervoir for the slow release of PDGF-BB at the infarcted region. Conse-
quently, the prolonged release of PDGF-BB from self-assembling
peptide nanofibers protected the cardiac cells in vivo and enhanced car-
diac function after MI. In contrast, the delivery of PDGF-BB without
peptide carriers showed no improvements in cardiac function. Similar
to the self-assembling peptide nanofibers that prolonged the release
of therapeutic molecules, chitosan-alginate nanoparticles were also
used to deliver angiogenic proteins in a sustainable manner [31]. PlGF
is another protein that stimulates angiogenesis [81]. Similar to PDGF-
BB, rapid clearance of PlGF after in vivo injection significantly impedes
the function of PlGF for therapeutic angiogenesis. Previously,
Binsalamah et al. developed chitosan-alginate nanoparticles that could
solve this problem, and showed over a 120-h period of sustained release
of PlGF in vitro using these nanoparticles. When injected in vivo, PlGF-
bound nanoparticles significantly improved cardiac function and pro-
moted vessel regeneration due to the sustained expression of PlGF at
the damaged region. Overall, these studies demonstrate that
nanobiomaterials, which provide growth factor protection from local
enzymatic degradation and sustained release of the protein at the injec-
tion site, can greatly enhance the therapeutic effects of drug delivery for
MI repair.
In addition to the direct delivery of therapeutic proteins to the MI re-
gion, nanobiomaterials have also been engaged in the delivery of genes.
Viral vector-based gene delivery, a commonly used method for gene
transfection may be immunogenic and oncogenic and thus its clinical
use is limited [82]. Therefore, non-viral nanomaterials have attracted
significant attention to replace viral vectors for gene delivery [83].
 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
Fig. 2. Integration of nanobiomaterials with stem cells for the treatment of MI. Nanobiomaterials can enhance the delivery efficacy of therapeutic molecules and cells for cardiac repair. In
addition, the nanotopographical, electrical, and chemical cues of nanobiomaterials can regulate stem cell functions for enhanced therapeutic efficacy for the treatment of MI.
Nanocarriers could prevent genes from endonuclease degradation, in-
hibit unwanted interactions with proteins, protect genes from immune
detection, and prolong gene circulation time for target delivery [83]. Che
et al. utilized polycaprolactone (PCL)/polyethylenimine (PEI) nanofi-
bers to deliver disulfide bond–cross-linked branched PEI nanoparticles
complexed with vascular endothelial growth factor (VEGF) genes
(ssPEI/VEGF nanoparticles) to cells cultured on nanofibers [32]. The
pores between PCL/PEI nanofibers enabled cells to grow in three dimen-
sions, and provided adequate mechanical properties for improved cell
adhesion and proliferation. Moreover, non-specific adsorption of
ssPEI/VEGF nanoparticles occurred uniformly over the surface of the
nanofibers, resulting in increased transgene uptake by the cultured
cells. The nanofibers had a high surface-to-volume ratio making it pos-
sible to immobilize a large quantity of ssPEI/VEGF nanoparticles. In ad-
dition, direct contact between the nanofiber-immobilized ssPEI/VEGF
nanoparticles and cells significantly reduced the amount of genes re-
quired for transfection compared to the bolus delivery of the nanoparti-
cles. Compared to branched PEI, a conventional polycation polymer
used for gene transfection [84], the use of ssPEI to form nanoparticles
with VEGF significantly reduced cytotoxicity and showed better trans-
fection efficiency of VEGF. Even though the utilization of PCL/PEI nano-
fibers has not been tested in vivo, this study showed that the
combination of nanofibers or nanoparticles with conventional gene de-
livery methods could suggest a promising gene therapy approach for
treating MI [32].
In addition to nanofibers, Zhang et al. introduced magnetic
nanobeads for enhanced gene delivery efficacy of VEGF-encoded adeno-
viral vectors (AdhVEGF) for the treatment of MI [78]. One of the major
drawbacks of the systemic infusion of vectors to treat MI is the low
targeting efficacy at the infarct region. Liver cells mostly take up the sys-
temically delivered vectors, and only a fraction of infused vectors are
targeted to the MI region. To overcome this limitation and promote ac-
tive targeting of adenoviral vectors to the infarcted region, magnetic
19
nanobeads (MNBs) were integrated with vectors. The conjugation of
MNBs with adenoviral vectors (MNBs/Ad) enabled higher gene trans-
duction efficiency in cultured cells under magnetic field stimulation
compared to Ad alone in vitro. Furthermore, the epicardial placement
of a magnet effectively attracted the MNBs/Ad complexes to the injured
myocardium when complexes were intravenously injected in vivo.
Overall, this gene delivery approach using MNBs significantly enhanced
the gene transfection efficiency in vivo, and reduced cardiac remodeling
for improved left ventricular function [78].
For stem cell homing and tissue repair, the delivery of stromal cell-
derived factor 1 (SDF-1) showed significant potential [85,86]. SDF-1 is
a chemokine involved in stem cell homing [87]. The expression of
SDF-1 is stimulated by the induction of MI, which in turn, recruits
stem cells to the injured myocardium [86]. The migration of stem cells
to the infarcted myocardium was further enhanced with SDF-1 over-
expression by SDF-1 gene delivery to the left ventricular wall using an
adenovirus vector [86]. However, locally delivered SDF-1 protein is vul-
nerable to rapid diffusion and cleavage. To overcome this problem,
Segers et al. designed SDF-1 that is resistant to cleavage, and tethered
this chemokine to self-assembling peptides (Fig. 3) [88]. The chemotac-
tic activity of SDF-1 was better preserved when SDF-1 was locally deliv-
ered with self-assembling peptides, and strong tethering of SDF-1 to
self-assembling peptide nanofibers showed controlled release of the
chemokine. SDF-1 protein was detectable for at least 7 days after injec-
tion only when it was delivered together with self-assembling peptide
nanofibers. Subsequently, delivery of nanofiber-integrated SDF-1 signif-
icantly enhanced stem cell homing, increased capillary density, and im-
proved cardiac function [88].
Nanobiomaterials can also be used to transfect cells in vitro prior to
implantation. For example, various types of nanobiomaterials effective-
ly transfected genes into cardiac progenitor cells [89], skeletal myo-
blasts [90], or adipose-derived stem cells [91]. Transfection of insulin
growth factor-1 (IGF-1) into cardiac progenitor cells using self-
20 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28

Fig. 3. Schematic illustration of nanobiomaterials utilized as vehicles for therapeutic molecule delivery for the treatment of MI. Incorporation of stem cell–recruiting SDF-1 fusion proteins
in self-assembling peptides. SDF-1–bound self-assembling peptides form nanofibers in vivo, retain chemotactic activity of SDF-1, and exhibit local release of SDF-1 at the infarcted region
for prolonged therapy for cardiac repair. Reproduced with permission from Ref. [88].

assembling peptide nanofibers prior to implantation enhanced cell sur-
vival in the infarcted heart and promoted heart repair [89]. When PEI
was used to transfect VEGF genes into skeletal myoblasts, implantation
of these transfected cells into the infarcted myocardium showed better
cell survival at the infarcted region and significantly improved cardiac
repair compared to naïve skeletal myoblasts [90]. In addition, transfec-
tion of the angiopoietin-1 gene into adipose-derived stem cells using
baculovirus nanocomplex prior to implantation promoted angiogenesis
in the infarcted heart with enhanced secretion of angiopoietin-1 [91].
Overall, these studies show that nanobiomaterials promote efficient
gene transfection of the cells, and propose a novel approach for cardiac
tissue engineering.
3.2. Cell delivery vehicles
Although stem cell therapy has emerged as a promising treatment
option for MI, its therapeutic benefit still remains limited due to poor
engraftment and survival of the implanted cells [92]. The efficient deliv-
ery and prolonged survival of stem cells have posed major challenges
for its application to treat MI [8]. Therefore, the development of new
therapeutic strategies that enhance the survival and extend the
reparative action of the implanted cells is critical to improve the out-
comes of stem cell therapy [93]. A number of nanobiomaterials have
been utilized as cell delivery systems, including cardiac patches and in-
jectable nanomaterials.
Nanostructured cardiac patches and injectable nanomaterials were
studied to improve stem cell retention after implantation in vivo [29,
94–96]. These cardiac patches are placed on the epicardial surface of
the infarcted heart and they require appropriate nanostructural and
chemical properties that are compatible with the native cardiac tissue.
For this reason, the fabrication of cardiac patches with collagen, the pre-
dominant extracellular matrix protein in the heart tissue, has attracted
significant attention due to its environmental compatibility [29]. Colla-
gen has a nanofibrillar structure and can bundle into a three-
dimensional scaffold such as cardiac patch, which mimics native cardiac
extracellular matrix. Simpson et al. previously demonstrated that colla-
gen patches seeded with MSCs improved MSC survival in vitro and re-
stored cardiac function after implantation (Fig. 4) [97]. Compared to
previous studies that showed less than 11% of cell engraftment after im-
plantation, MSCs delivered with collagen patches achieved a cell en-
graftment of 23% and resulted in improved cardiac function. After
injection, the survival and engraftment of MSCs is often limited by the

Fig. 4. Schematic illustration of nanobiomaterials utilized as vehicles for cell delivery for the treatment of MI. Stem cell–loaded collagen patch is placed on the epicardial surface of the
infarcted myocardial region and held with fibrin sealant. Stem cells implanted with collagen patch show significantly improved initial cell engraftment for better cardiac function improve-
ment compared to direct cell injection. Reproduced with permission from Ref. [97].
 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
inflammatory microenvironment at the infarct region and the leakage
of the implanted cells from the injection site. Collagen-based cardiac
patch provided cell-friendly extracellular matrix for better cell survival,
and uniformly fixed MSCs in proximity to the infarct area for better en-
graftment. Cardiac patches without MSCs showed no improvement in
the cardiac function, suggesting that the major therapeutic effect was
attributed to the implanted stem cells.
Along with the natural materials such as collagen, synthetic mate-
rials have also been investigated for cardiac patch development [29].
Among the various types of synthetic polymers explored, poly(lactide-
co-ε-caprolactone) (PLCL) was suggested as a suitable candidate for car-
diac patches due to its elasticity and biodegradability [98]. PLCL is a
highly elastic polymer, which can form nanofibrous scaffolds, and may
serve as an excellent matrix for cardiac tissue engineering applications.
When MSCs were seeded onto a PLCL-based cardiac patch and im-
planted onto an infarcted heart, both MSC survival and cardiac function
were significantly improved compared to the direct injection of MSCs
[98]. This is because the cardiac patch served as a mechanical extracel-
lular matrix for MSC survival and allowed MSCs to differentiate into
functional cardiomyocytes. The study further demonstrated that PLCL
cardiac patch alone could not restore cardiac function, while MSC
alone could only partially restore heart function and reduce fibrosis
[98]. However, contradictory results were also reported in other cardiac
patch studies, where implantation of cardiac patch alone improved car-
diac function [99,100]. Therefore, further studies are required to com-
prehensively investigate the efficacy of acellular patches and their
impact on stem cell therapy for cardiac repair.
Bio-inspired self-assembling peptide nanofibers can be utilized as a
cell-delivery vehicle for the treatment of MI [101–103]. Due to their bio-
compatibility and biodegradability, self-assembling peptides are uti-
lized not only in drug delivery but also in cell delivery for cardiac
repair. Previous studies exploiting self-assembling peptides as cell-
delivery vehicles showed great improvement in the survival of the im-
planted cells and facilitate myocardial regeneration after MI
[101–103]. This is primarily because self-assembling peptides may in-
crease cell retention and promote cell survival after in vivo delivery. To
further elucidate the function of self-assembling peptides, Cui et al.
demonstrated that these peptide nanofibers can enhance growth, sur-
vival, and differentiation of MSCs in vitro [103]. These peptides sponta-
neously assembled into nanofiber scaffolds that mimic the natural
extracellular matrix post-implantation. Consequently, MSCs mixed
with self-assembling peptides exhibited increased cell survival and re-
tention at the infarct size in vivo, resulting in improved cardiac function
[103]. Furthermore, Guo et al. integrated self-assembling peptides with
cardiac stem cells and demonstrated their therapeutic application in rat
MI model [104]. To enhance survival and cardiac differentiation of CSCs
in vivo, Guo et al. constructed self-assembling peptides tethered with
the cell-adhesion motif RGDSP. When these peptides were implanted
together with CSCs, they bundled into nanofiber scaffolds, providing a
suitable microenvironment for CSC adhesion and survival, and
protected CSCs from apoptosis and necrosis caused by anoxia. Increased
survival of the CSCs further enhanced their cardiac differentiation and
resulted in better cardiac repair. Compared to the groups treated with
either CSCs or self-assembling peptides alone, the combination of CSCs
and peptides significantly improved cardiac function and reduced colla-
gen deposition. Collectively, these studies suggest that self-assembling
peptides can serve as an efficient cell delivery vehicle increasing cell
survival, retention, and differentiation, and thereby promoting the ther-
apeutic efficacy of stem cell therapy for the treatment of MI.
3.3. Nanotopographical cues of nanobiomaterials
Nanobiomaterials with nanotopographical cues have been applied
in cell therapy to modulate cell behaviors such as cell differentiation,
gap junction protein expression, and paracrine molecule secretion for
cardiac tissue repair. For development of therapeutic nanobiomaterials
21
for the treatment of MI, researchers predominantly focused on the in-
nate anisotropic property of the heart because cardiac-mimetic struc-
tural nanotopography could pose significant effects on the cells [9,10,
29]. The heart is a constantly working, dynamic, and a complex organ.
For the functional activity of the heart, the cardiac cells that comprise
the heart muscles require a quick and efficient signal transfer. Therefore,
the alignment of the cardiac cells and proper gap junction protein distri-
bution are required for efficient cell signal transduction and synchro-
nous contraction of the heart [105]. To mimic the anisotropic property
of the heart, various nanotopography-associated approaches using
nanobiomaterials were introduced to modulate cell behaviors, includ-
ing cell alignment, differentiation and paracrine secretion, and then in-
tegrated with stem cells for cardiac repair.
A previous study demonstrated that contact guidance of the cells
by aligned nanotopography or electrical field stimulation is crucial in
the orientation and elongation of cardiomyocytes [38]. Interestingly,
this study showed that while the combination of nanotopography
and electrical stimulation can promote the contractile property of
cardiomyocytes, the topographical cue might serve as a stronger de-
terminant of cardiomyocyte alignment than electrical field stimula-
tion. Several studies previously demonstrated that aligned
nanofibers, which mimic the nanostructures of natural extracellular
matrices of the heart tissue [39], improve the alignment [40] and
beating of the cardiomyocytes [42]. Cardiomyocytes cultured on
aligned nanofibrous electrospun patches showed a higher beating
frequency and amplitude compared to those cultured on randomly
oriented fibrous patches [40]. Upon implantation in vivo, the
cardiomyocyte-seeded aligned cardiac patch improved cardiac func-
tion, while the cardiomyocytes-seeded randomly oriented patch re-
sulted in serious deterioration in the cardiac function [42]. This study
indicates that the anisotropic property of nanobiomaterials signifi-
cantly affects the therapeutic outcomes of cell therapy for the treat-
ment of MI, and is crucial for developing biomaterial-based therapy
for MI.
The anisotropic properties of nanobiomaterials further affected the
stem cell behavior for higher therapeutic efficacy. Kim et al. have
previously shown that nanotopographically defined polyethylene glycol
(PEG) hydrogel, which mimics the structural features of the native myo-
cardial matrix can augment adhesion, migration, and proliferation of
CSCs [41]. Implantation of stem cell–seeded anisotropic PEG scaffolds
showed successful integration with the host tissue due to the
cardiac extracellular matrix-mimicking nanostructures. Moreover,
nanopatterned PEG scaffold significantly improved retention and
growth of transplanted CSCs. Consequently, the implantation of CSC-
seeded aligned cardiac patch greatly reduced cardiac fibrosis and
improved cardiac function [41]. The anisotropic properties of
nanobiomaterials also improved the paracrine action of stem cells.
Kang et al. recently demonstrated that MSCs cultured on aligned
fibronectin-immobilized PCL nanofibers exhibited enhanced expression
of angiogenic and cardioprotective genes [106]. This study showed that
MSCs cultured on anisotropic fibronectin-immobilized PCL nanofibers
exhibited aligned cellular phenotypes found in native cardiac tissue.
Furthermore, fibronectin immobilization significantly elevated the ex-
pression of genes involved in stem cell homing, anti-apoptosis, anti-
inflammation, and angiogenesis in MSCs. Subsequently, implantation
of MSC-seeded anisotropic scaffolds significantly improved the cardiac
function in rat MI model (Fig. 5) [106]. The mechanisms for the
fibronectin-immobilized PCL nanofiber-mediated cellular signaling
and paracrine molecule expression were not clarified in this study.
Thus, further studies differentiating the effects of fibronectin immobili-
zation from nanofiber alignment are required. Overall, even though the
above-mentioned studies by Kim et al. [41] or Kang et al. [106] did not
elucidate the mechanisms involved in cell behavior modulation on
nanotopographically designed polymer scaffold, they certainly show
that developing nanobiomaterials mimicking the nanostructures of car-
diac extracellular matrix is required for improved cardiac repair.
22 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28

Fig. 5. Schematic representation of nanotopography of nanobiomaterials utilized for cardiac tissue engineering. Poly(glycidyl methacrylate) (pGMA)-coated aligned polycaprolactone
(PCL) nanofibers were prepared, and fibronectins were immobilized onto the pGMA-PCL nanofibers. MSCs seeded on the nanofibers exhibit aligned cellular phenotypes found in natural
extracellular matrix of the heart. The MSCs-seeded fibronectin-immobilized aligned nanofibrous cardiac patches implanted onto a rat MI model show better tissue compatibility and sig-
nificantly improved cardiac function. Reproduced with permission from Ref. [106].

To further analyze the effect of cell alignment on the intercellular in-
tegration and electrical conduction of stem cells, a previous study intro-
duced cellular alignments during MSC-cardiomyocyte co-culture [69].
Pijnappels et al. showed that a forced alignment of MSCs on a
directionally patterned substrate significantly improved cardiac differ-
entiation of MSCs and their functional integration with cardiomyocytes
in the MSC-cardiomyocyte co-culture [69]. This study further demon-
strated the alignment-dependent increase in MSCs’ conduction velocity
and expression of the gap junction protein, connexin 43 (Cx43). This
study shows that nanopatterns not only regulate the behaviors of the
cells cultured on scaffolds but also affect the synchronous integration
of MSCs with the surrounding cardiac environment. Taken together,
the aforementioned studies suggest that cell alignment contributes to
the therapeutic outcomes of cell therapy, and the development of
nanobiomaterials with proper anisotropic property is desirable for suc-
cessful cardiac repair.
3.4. Electrically conductive nanobiomaterials
Along with the topographical property, electrical conductivity is an-
other unique feature of the heart that is critical in developing therapeu-
tic biomaterials for cardiac repair. Unlike cells in other organs, cardiac
cells need gap junction–based intercellular coupling and electrical con-
duction for natural heart function. Thus, electrochemically unsuitable
therapies may pose electrophysiological discordance with native cardi-
ac tissues, and may cause arrhythmias upon application [105]. There-
fore, it has been speculated that electrical conductivity may modulate
the behaviors of stem cells and affect their electrochemical signaling
pathways that contribute to cardiac tissue integration. To mimic and
preserve the electrical conductivity of the heart, previous studies fo-
cused on the development of conductive nanobiomaterial-integrated
therapeutics that are electrically compatible with native cardiac
tissues. Various nanobiomaterials, including carbon- and gold-based
nanomaterials, showed electrical conductivity [107–109], and
researchers have incorporated these nanomaterials with cell therapy
for treating MI.
A number of previous studies showed that integration of carbon
nanofibers or carbon nanotubes with polymer scaffolds promotes the
adhesion and proliferation of cardiomyocytes [43] and cardiac progeni-
tor cells [44]. In addition, these conductive nanomaterials significantly
promoted the expression of Cx43 in cardiac cells and stimulated the
electrical coupling between the cells. Cx43 expression is particularly im-
portant for MI repair because it is essential for intercellular coupling, ar-
rhythmic risk reduction, and functional cardiomyogenic differentiation
of the cells [67,73,105,110]. Although the mechanisms for Cx43 upregu-
lation have not been clearly elucidated, the overexpression of Cx43 in
cells cultured on conductive scaffold suggests a potential use of conduc-
tive nanobiomaterials for cardiac repair. To better understand the ther-
apeutic potential of conductive materials and analyze the beating
behavior of the cells on a conductive scaffold, Martines et al. and
Kharaziha et al. investigated the electrophysiological function of
cardiomyocytes seeded on a chitosan matrix incorporated with carbon
nanofibers [48] or poly(glycerol sebacate):gelatin scaffolds embedded
with carbon nanotubes [50], respectively. Cardiac cells seeded on a con-
ductive nanobiomaterial-integrated scaffold exhibited a synchronous
beating behavior and an increased expression of cardiac-specific genes
involved in cardiac muscle contraction and electrical coupling. Taken to-
gether, the aforementioned studies demonstrate that conductive
nanobiomaterials not only enhance the electrical coupling between
the cells with Cx43 upregulation, but also induce synchronous beating
of the cardiac tissue. The in vivo application of carbon nanomaterial–in-
tegrated scaffolds and the mechanisms involved in how the conductive
materials affect cell behaviors require further studies.
Gold nanomaterials were also utilized to improve the therapeutic ef-
fects of cell therapy. Cardiomyocytes cultured in hydrogel scaffolds inte-
grated with gold nanoparticles [45] or gold nanowires [46] showed an
increased Cx43 expression compared to cardiomyocytes cultured in
hydrogels without the integration of gold nanomaterials. Dvir et al.
 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
Fig. 6. Schematic representation of electrically conductive nanobiomaterials utilized for cardiac tissue engineering. Incorporation of electrically conductive RGO flakes in MSC spheroids
greatly improves the expression of the gap junction protein Cx43, which is critical in native cardiac tissue compatibility and function. RGO-incorporated MSC spheroids are directly injected
into the infarcted region of a rat heart for greater cardiac function improvements compared to MSC spheroids incorporated with insulating graphene oxide flakes. Reproduced with per-
mission from Ref. [112].
incorporated gold nanowires into porous alginate scaffolds to develop
three-dimensional conductive nanocomposites [46]. Non-conducting
alginate pore walls bridged with conductive gold nanowires demon-
strated enhanced electrical signal propagation, and enabled electrical
communication between the cardiomyocytes seeded on the scaffolds.
Consequently, cardiomyocytes grown on the conductive nanocompos-
ite showed a better aligned tissue structure, and exhibited higher levels
of the proteins involved in muscle contraction. The cells demonstrated
synchronous contraction when the conductive nanocomposites were
electrically stimulated [46]. Similar to the studies with carbon-based
nanomaterials, this study suggests that the integration of gold
nanomaterials with polymeric scaffolds may improve the therapeutic
efficacy of cell therapy.
To determine whether conductive nanomaterials can provide
cardiomimetic cues to MSCs for cardiac differentiation, a number of
studies investigated the MSCs’ phenotype changes upon treatment
with conductive materials. Mooney et al. demonstrated that electri-
cal stimulation of carbon nanotube–embedded scaffolds significantly
enhanced the cardiomyogenic differentiation of MSCs seeded on the
scaffolds [47]. Electrical stimulation reoriented the cells perpendicu-
lar to the direction of the current and significantly augmented the
expression of cardiac-specific biomarkers including Cx43. Without
the carbon nanotubes, however, electrical stimulation alone did not
promote cardiac protein expression in MSCs. To further differentiate
the effect of electrical stimulation from the conductive materials,
Crowder et al. further investigated carbon nanotube-containing syn-
thetic polymer scaffolds without electrical stimulation on the cardiac
differentiation of MSCs [49]. They observed that the electrically con-
ductive nanobiomaterials alone promoted cardiac differentiation of
the stem cells and their Cx43 expression without any electrical stim-
ulation. These studies, however, did not clarify the mechanisms in-
volved in cardiac differentiation and Cx43 enhancement of MSCs
cultured on conductive materials. Further studies elucidating the ef-
fects of conductive materials on cell behavior modulation and differ-
entiation are therefore required for the therapeutic application of
conductive materials for MI.
23
Conductive nanobiomaterials have further been utilized together
with conventional cardiac differentiation methods to stimulate car-
diac differentiation of MSCs. Ravichandran et al. demonstrated that
5-azacytidine treatment on MSCs seeded onto gold nanoparticle–
embedded conductive Bovine serum albumin (BSA)/Polyvinyl alco-
hol (PVA) nanofibrous scaffolds greatly stimulated the MSC differen-
tiation towards cardiomyogenic lineage and enhanced Cx43
expression [111]. This study indicates that the electrical conductivity
of nanobiomaterials can exert synergistic effects with conventional
cardiac differentiation methods and that the combined approach
can be a superior therapy for MI. Park et al. further demonstrated
the therapeutic effect of conductive nanomaterials in vivo, and re-
cently showed that the conductive and intrinsic chemical properties
of reduced graphene oxide (RGO) can have synergistic effects to
modulate the MSC function for the treatment of MI (Fig. 6) [112].
RGO is composed mostly of carbon atoms and it has unique mechan-
ical, chemical, and electrical properties [113]. The study showed that
the incorporation of RGO flakes in MSC spheroids significantly en-
hanced the Cx43 expression of MSCs. In contrast, graphene oxide
flakes, which are insulating due to the existence of functional groups
that hinder the transfer of π-electrons, significantly reduced the
Cx43 expression in MSCs. Based on this data, the authors postulated
that the enhanced expression of Cx43 in MSCs is attributed to the
electrical conductivity and cell-extracellular matrix interactions
generated by RGO flakes. Ultimately, the implantation of MSC spher-
oids integrated with RGO flakes significantly reduced tissue fibrosis
and improved cardiac function.
Conductive nanobiomaterials have actively participated in intercel-
lular coupling and cardiac differentiation, showing great potential in
therapeutic applications for the treatment of MI. However, the mecha-
nisms behind their effects on stem cell behaviors still remain elusive.
Moreover, conductive nanobiomaterials need to properly mimic the
electrical conductivity of the native heart to reduce the arrhythmic
risks of in vivo application [105]. Future studies elucidating the
mechanisms involved in MSC behavior modulation by conductive
nanomaterials, and the development of in vivo mimetic conductive
24 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28
materials would provide a better therapeutic window for the applica-
tion of conductive nanobiomaterials for cardiac repair.
3.5. Intrinsic chemistry of nanobiomaterials
Nanobiomaterials with distinct chemical properties can also be ap-
plied to stem cell therapy for MI. Previous studies speculated that nano-
scale properties of nanobiomaterials might pose unique chemical
characteristics that can affect adhesion, growth, differentiation and
paracrine secretion of stem cells. Nanobiomaterials with functional
chemical properties can capture growth factors secreted by stem cells
and be used for acellular therapies for MI [114], stimulate stem cells to
secrete more paracrine molecules for enhanced therapeutic efficacy
[112], induce differentiation of stem cells toward cardiac lineage [115],
and control intracellular signaling pathways for cell function
modulation [51].
Self-assembling peptide nanofibers that capture paracrine mole-
cules have been utilized for MI repair [114]. Webber et al. demon-
strated that heparin-presenting self-assembling peptides can
bundle into a matrix consisting of fibrillar nanostructures similar to
the natural cardiac extracellular matrix [114]. The presentation of
heparin on the surface of self-assembling peptide nanofibers further
enabled the binding of growth factors secreted from the stem cells in
cell cultures via heparin-binding domains [114]. Using recombinant
proteins of VEGF and basic fibroblast growth factor (bFGF) loaded
in the nanofibers, the effects of the nanofiber-captured growth fac-
tors from the stem cell–conditioned medium were recapitulated.
When heparin-presenting self-assembling peptides were incubated
with hypoxic stem cell conditioned medium, they captured more
than 58% of bFGF present in the medium. When implanted into the
damaged heart, the growth factor–bound nanofibers successfully re-
stored the hemodynamic function of the heart by modulating the
cardiac metabolism [114]. This study shows the therapeutic poten-
tial of synthetic nanomaterials in combination with stem cell para-
crine factors for the treatment of MI.
Cell-sheet engineering also serves as a treatment option for the
treatment of MI. Implantation of stem cell sheets can significantly
enhance the stem cells’ capability to repair the injured myocardium
compared to the implantation of dissociated stem cells [116].
Okura et al. have previously demonstrated successful treatment of
MI using MSCs cultured on thermo-responsive dishes [117]. The
study demonstrated that the implantation of cell sheets composed
of MSC-derived cardiomyoblast-like cells resulted in functional dif-
ferentiation of the cells towards cardiomyocytes in vivo and en-
hanced cardiac function [117]. With the unique thermo-responsive
properties of poly(N-isopropylacrylamide) (PIPAAm), PIPAAm-
grafted surface provides a useful tool to form cell sheets as it allows
cells to adhere and detach depending on the temperature [116]. Fur-
ther studies demonstrated that nanoscale thickness of PIPAAm coat-
ing on the surface plays a crucial role in the formation and
detachment of the cell-sheets [118,119]. Akiyama et al. showed
that cell adhesion on PIPAAm-grafted surface is inhibited when the
coating thickness of PIPAAm increases from 15.5 ± 7.2 nm to
29.5 ± 8.4 nm, because PIPAAM grafting inhibits adsorption of cell
adhesion proteins like fibronectin to the surface [118].
Iron oxide nanoparticles have also been utilized for cell function
modulation and cardiac repair due to their innate chemistry. Widely
used for in vivo imaging, iron oxide nanoparticles were recently ap-
plied to modify the cell functions of H9C2 cells, a cardiomyoblast
cell line, in an MSC-H9C2 co-culture [51]. The co-culture of MSCs
with cardiac cells previously demonstrated priming of MSCs toward
cardiac lineage and the implantation of these cardiac primed-MSCs
into the infarcted heart improved cardiac function [67]. For co-
culture, the rat cardiomyoblast cell line H9C2 has attracted much
attention due to its easy accessibility compared to primary
cardiomyocytes. However, the low level of Cx43 expression in
H9C2 cells [120] limits the effectiveness of MSC cardiac priming by
MSC-H9C2 co-culture [51]. Han et al. recently showed that the inter-
nalization of iron oxide nanoparticles can augment the Cx43 expres-
sion in H9C2 cells by delivering metal ions and modulating the
intracellular c-Jun N-terminal kinase signaling [51]. Changes in in-
tracellular ion levels can induce significant alteration in cell signal-
ing, and may cause cell cytotoxicity. Hence, Han et al. speculated
that a change in the concentration of intracellular iron ion level
might change cell signaling pathways. Conforming to previous stud-
ies, exogenous delivery of metal ions caused significant toxicity to
the cells. However, using iron oxide nanoparticles, Han et al. success-
fully increased metal ion levels within the cells without cytotoxicity,
and initiated cell signaling alteration for Cx43 upregulation. The en-
hanced Cx43 expression in the H9C2 cells further stimulated the in-
tercellular crosstalk between the H9C2 cells and MSCs. Hence, MSCs
co-cultured with iron oxide nanoparticle–internalized H9C2 cells
showed improved cardiac differentiation and paracrine molecule se-
cretion. Consequently, the implantation of co-cultured MSCs re-
duced fibrosis, enhanced angiogenesis, and improved cardiac
function (Fig. 7A) [51].
In addition to iron oxide nanoparticles, carbon-based
nanomaterials have also shown significant potential in stimulating
cell signaling and cardiac differentiation of stem cells. Park et al. re-
cently showed that graphene, a single layer of carbon atoms stimu-
lated the MSC differentiation process toward cardiomyogenic
lineage [115]. When MSCs were cultured on graphene, its unique
chemical nanostructure stimulated the expression of the extracellu-
lar matrix and signaling molecules related to cardiomyogenic differ-
entiation, and in turn, enhanced cardiomyogenic gene expression in
MSCs (Fig. 7B) [115]. Lee et al. also demonstrated that graphene pro-
motes the cardiac differentiation of ESCs [121]. ESCs cultured on
graphene exhibited significant changes in the extracellular signal-
regulated kinase and the focal adhesion kinase signaling compared
to ESCs cultured on glass or matrigel. Consequently, ESCs cultured
on graphene showed great increase in the expression of genes asso-
ciated with cardiac transcription, early cardiomyocytes, late
cardiomyocytes, and gap junctions [121].
The intrinsic chemistry of nanobiomaterials also contributed to the
enhanced paracrine secretion and Cx43 expression of MSCs cultured
in spheroid forms (Fig. 6) [112]. MSCs in spheroid forms have very lim-
ited cell-extracellular matrix interactions, which are necessary for para-
crine secretion and cell survival. Recently, Park et al. demonstrated that
RGO efficiently adsorbs extracellular matrix proteins via π-π stacking
and hydrophobic interactions [122], providing better cell-extracellular
matrix interactions for MSCs in spheroids [112]. When RGO flakes
were cultured in serum-containing cell culture medium, significant ad-
sorption of extracellular matrix protein fibronectin was observed on the
surface of RGO flakes. Furthermore, incorporation of these RGO flakes in
MSC spheroids increased the expression of cell signaling molecules
associated with cell-extracellular matrix interactions. The enhanced
cell-extracellular matrix interactions then stimulated paracrine mole-
cule secretion and Cx43 expression. The implantation of these RGO
flake-incorporated MSC spheroids significantly improved the cardiac
function in mouse MI models compared to MSC spheroids [112]. Collec-
tively, previous studies showed that the integration of nanobiomaterials
with stem cells improves the therapeutic outcomes of conventional
stem cell therapy for the treatment of MI. Hence, defining the intrinsic
chemical properties of the nanomaterials that can influence the stem
cell function would provide a basis for the development of better thera-
peutic options for future MI therapies.
4. Conclusions
Stem cell therapy has become one of the most encouraging treat-
ment options for MI. The simple delivery of stem cells has been applied
in a number of preclinical and clinical studies; however, the therapeutic
 J. Han et al. / Advanced Drug Delivery Reviews 95 (2015) 15–28 25

Fig. 7. Effects of intrinsic chemical properties of nanobiomaterials on cell function and cardiac repair. (A) Mechanisms involved in iron oxide nanoparticle–induced Cx43 expression en-
hancement in cardiomyoblast H9C2 cell line, and in vivo therapeutic efficacy of the co-cultured MSCs. *p b 0.05. Internalization of iron oxide nanoparticles affects intracellular JNK signaling
pathway for Cx43 upregulation, which is crucial in intercellular crosstalk between MSCs and H9C2. MSCs co-cultured with iron oxide nanoparticle–internalized H9C2 (cMSC) show better
cardiac lineage development and in vivo therapeutic efficacy. (B) Schematic diagram of the signal transduction pathways associated with cardiomyogenic differentiation and cell survival,
and Western blot analyses regarding the molecules involved in the signal transduction pathways of MSCs cultured on graphene or coverslips for 3 weeks. Graphene stimulates the expres-
sion of extracellular matrix and signaling molecules involved in cardiomyogenic differentiation of MSCs. Quantification of the relative protein expressions of the cell signaling molecules.
*p b 0.05 compared to MSCs cultured on coverslips. Reproduced with permission from Refs. [51,115].

efficacy of stem cells still needs improvement and the efficient clinical
translation of stem cell therapy remains elusive. To improve the thera-
peutic efficacy of stem cells, nanobiomaterials have been utilized in a
number of different forms. Nanobiomaterial-integrated strategies for
the treatment of MI include the delivery of therapeutic molecules or
cells, the development of cardiac niche-mimicking biomaterials with
electrical conductivity or anisotropic nanotopography, and the exploita-
tion of the chemical properties of nanomaterials. Nanoparticles and self-
assembling peptides have been utilized in drug or cell delivery systems
to improve the delivery efficiency of the therapeutic molecules and or
the cells at the infarcted myocardium. Additionally, nanotopographical
cues and electrical conductivity of nanomaterials have been incorporat-
ed in nanobiomaterial designs to mimic the native properties of the
heart and promote cardiomyocyte-like functions in stem cells. Finally,
the interactions between the inherent chemical properties of
nanobiomaterials with the stem cells were shown to affect the thera-
peutic behaviors of these stem cells. Taken together, the integration of
nanobiomaterials with stem cells showed a synergistic effect for the
treatment of MI. The development and utilization of functional
nanobiomaterials that can achieve cooperative therapy with stem cells
would provide a promising strategy for cardiac repair after MI in the
future.
Acknowledgments
This research was supported by the National Research Foundation of
Korea (2014073757) and the Korea Health Industry Development Insti-
tute (KHIDI), Ministry of Health and Welfare (HI14C3270, HI14C1550
and HI15C0498), Republic of Korea.
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