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Recent Contributions of Elastin-Like Recombinamers to Biomedicine and Nanotechnology

28 Feb 2014-Current Topics in Medicinal Chemistry (Curr Top Med Chem)-Vol. 14, Iss: 6, pp 819-836

TL;DR: An updated review of the recent challenges overcome by using Elastin-like Recombinamers in the fields of nano- and biomedicine, ranging from nanoscale applications in surface modifications and self-assembled nanostructures to drug delivery and regenerative medicine.
Abstract: The emergence of the new scientific field known as nanomedicine is being catalyzed by multiple improvements in nanoscience techniques and significant progress in materials science, especially as regards the testing of novel and sophisticated biomaterials. This conjuncture has furthered the development of promising instruments in terms of detection, bioanalysis, therapy, diagnostics and imaging. Some of the most innovative new biomaterials are protein-inspired biomimetic materials in which modern biotechnology and genetic-engineering techniques complement the huge amount of information afforded by natural protein evolution to create advanced and tailor-made multifunctional molecules. Amongst these protein-based biomaterials, Elastin-like Recombinamers (ELRs) have demonstrated their enormous potential in the fields of biomedicine and nanoscience in the last few years. This broad applicability derives from their unmatched properties, particularly their recombinant and tailor-made nature, the intrinsic characteristics derived from their elastin-based origin (mainly their mechanical properties and ability to self-assemble as a result of their stimuli-responsive behavior), their proven biocompatibility and biodegradability, as well as their versatility as regards incorporating advanced chemical or recombinant modifications into the original structure that open up an almost unlimited number of multifunctional possibilities in this developing field. This article provides an updated review of the recent challenges overcome by using these recombinant biomaterials in the fields of nano- and biomedicine, ranging from nanoscale applications in surface modifications and self-assembled nanostructures to drug delivery and regenerative medicine.

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Recent Contributions of Elastin-Like Recombinamers to Biomedicine and
Nanotechnology
F. Javier Arias
*
, Mercedes Santos, Alicia Fernández-Colino, Guillermo Pinedo and
Alessandra Girotti
BIOFORGE Research Group, University of Valladolid, CIBER-BBN, Edificio I+D, Paseo de Belen 11, 47011,
Valladolid, Spain
Abstract: The emergence of the new scientific field known as nanomedicine is being catalyzed by multiple improvements
in nanoscience techniques and significant progress in materials science, especially as regards the testing of novel and so-
phisticated biomaterials. This conjuncture has furthered the development of promising instruments in terms of detection,
bioanalysis, therapy, diagnostics and imaging. Some of the most innovative new biomaterials are protein-inspired
biomimetic materials in which modern biotechnology and genetic-engineering techniques complement the huge amount of
information afforded by natural protein evolution to create advanced and tailor-made multifunctional molecules. Amongst
these protein-based biomaterials, Elastin-like Recombinamers (ELRs) have demonstrated their enormous potential in the
fields of biomedicine and nanoscience in the last few years. This broad applicability derives from their unmatched proper-
ties, particularly their recombinant and tailor-made nature, the intrinsic characteristics derived from their elastin-based
origin (mainly their mechanical properties and ability to self-assemble as a result of their stimuli-responsive behavior),
their proven biocompatibility and biodegradability, as well as their versatility as regards incorporating advanced chemical
or recombinant modifications into the original structure that open up an almost unlimited number of multifunctional pos-
sibilities in this developing field. This article provides an updated review of the recent challenges overcome by using these
recombinant biomaterials in the fields of nano- and biomedicine, ranging from nanoscale applications in surface modifica-
tions and self-assembled nanostructures to drug delivery and regenerative medicine.
Keywords: Elastin-like recombinamers, self-assembly, stimuli-responsive, tissue engineering, surface modification, drug de-
livery, nanotechnology.
INTRODUCTION
Protein based-materials have attracted the attention of
numerous researchers in recent years as promising advanced
biomaterials for use in the field of biomedicine, especially as
a result of recent improvements in recombinant DNA tech-
nology, which allow us to design and manufacture materials
by exploiting the abilities of natural proteins [1]. Some of the
most widely studied protein-derived materials are the so-
called elastin like recombinamers (ELRs) [2-4], taking into
account its recombinant nature this new nomenclature was
proposed [5] in replacement of the more conventional termi-
nology elastin- like polymers (ELPs), which include those
first chemically synthesized materials.
Elastin is an elastic, insoluble protein that is present in
many tissues such as skin or lung where elasticity is a key
requirement. Elastin possesses several extraordinary charac-
teristics, such as an ability to undergo high deformation
without breaking and subsequent recovery of the original
conformation once the stress disappears. The origin of this
ability resides in the structure of the recurrent sequences
(VPGVG, VPGG, VGVAPG) found in the soluble elastin
*Address correspondence to this author at the BIOFORGE Research Group,
University of Valladolid, CIBER-BBN, Edificio I+D, Paseo de Belen 11,
47011, Valladolid, Spain; Tel: ??????????; Fax: ??????????;
Email: arias@bioforge.uva.es
precursor tropoelastin [6]. It is also worth noting that this
elastic behavior is an energy-conserving process, thus allow-
ing the resulting elastic fibers to undergo billions of relaxa-
tion-stretching cycles [7].
ELRs are smart, genetically engineered biomaterials in-
spired by natural elastin and based on the very same recur-
ring amino acid sequences as found in tropoelastin. Some of
the most relevant characteristics of ELRs, which are derived
from those of the natural protein, serve as an example of how
elastin’s mechanical properties are retained by way of cross-
linked ELR matrices [8]. These mechanical properties be-
come more interesting in conjunction with other properties
such as biocompatibility, stimuli-responsive behavior and
the ability to self-assemble [9-10]. The most widely studied
ELRs are based on the recurring pentapeptide sequence Val-
Pro-Gly-Xaa-Gly, where Xaa is any natural amino acid ex-
cept proline. All functional ELRs present temperature sensi-
tiveness in aqueous solution. Thus, below a characteristic
temperature, known as the transition temperature (Tt), the
polymer chains remain disordered and relatively extended
with a random coil conformation. They are also fully hy-
drated, mainly by a hydrophobic hydration characterized by
the presence of ordered water clathrate-like structures around
the apolar moieties in the polymer [11]. Once this Tt is ex-
ceeded, this disordered structure is “broken” and the “re-
leased” polymer chains adopt an ordered structure known as

2 Current Topics in Medicinal Chemistry, 2014, Vo l. 14, No. 7 Arias et al.
a -spiral, which folds hydrophobically to generate a phase
separation. The initial step in this process is the formation of
triply stranded -spiral filaments, which go on to form
nanoparticles several hundreds of nanometers in length as a
result of continued growth of these filaments. Finally, the
filaments come together to form a visibly phase-separated
state. The process is completely reversible upon lowering the
temperature below the Tt [10]. However, temperature is not
the only stimuli to which ELRs are responsiveness: other
external stimuli, such as pH [12], UV-vis light [13] or ion
concentration [14] also have a marked effect on the behavior
of ELRs.
Some disagreement still standing about the -spiral sig-
nificance, some authors have demonstrated the existence of
other conformations, such as polyproline II in elastin [15],
proposing an alternative model where -spiral are accompa-
nied by isolated -turns [16] and extended structures. [15,
17] The presence of high amount of water in the system,
increasing the dynamics of the recombinamer chains would
explain the apparent contradictory results obtained by differ-
ent experimental techniques.
ELRs are obtained by recombinant DNA technologies,
which allow an extremely precise control over the amino
acid composition. This technology opens up the possibility
of designing ELRs with the specific properties required for a
certain application [18, 19]. This design versatility, together
with their high biocompatibility, bioactivity and self-
assembling behavior, makes ELRs unmatchable materials for
studies in the biomedical field [18]. As a result, ELRs have
been used in wide variety of applications, such as tissue en-
gineering [20] or drug delivery [21]. Similarly, they have
also been applied in the field of nanotechnology due to the
range of nanostructures, such as nanofibers [22] or nanopar-
ticles [23], that can be formed from them.
The aim of this review is to try to shed some light on the
state-of-the-art of these and other applications of ELRs in the
fields of biomedicine and nanotechnology.
ELRs IN TISSUE ENGINEERING
The potential of ELRs in biomedical engineering has
been widely explored over the last decade, particularly in the
field of tissue engineering for the construction of bioartificial
tissues. Indeed, these recombinant protein polymers can even
be suitably designed and molded to restore the structural and
biological functions of the extracellular matrix (ECM) [24].
The ability to biosynthesize ELRs guarantees their repro-
ducibility, thereby avoiding the potential variability and risk
of using animal-derived structural proteins. Moreover, their
characteristics make them biomimetic and re-absorbable
materials with a defined mechanical behavior [25, 26] that
allows them to be cast with a specific topography [27, 28]
whilst retaining their tunable temperature responsiveness
[29] and controlled structural features. ELR sequences based
on the repetition of certain modular ECM structural protein
motifs allow the physicochemical [10], mechanical [30], and
biological [31] properties of the natural proteins from which
they are derived to be reproduced [32]. These properties
have made ELRs one of the materials of choice when inves-
tigating new biomaterials for biomedical applications.
It has been widely demonstrated that ELRs are able to be
formed into plastic or viscoelastic scaffolds for tissue engi-
neering [33]. Due to the possibility of partially substituting
their amino acid sequence to modulate potential cross-
linking or their ability to self-assemble, ELRs that are able to
form physical or chemical gels have been synthesized.
Moreover, some of them are injectable and can form a stable
continuous implant in situ, a property that is particularly in
demand as regards minimizing surgical invasiveness [28, 34,
35]. Such injectable 3D ELR scaffolds have been shown to
be suitable for supporting in vitro cell culture and the differ-
entiation of adipose-derived stem cells into chondrocytes
without the need to add external supplements [36], and
Chaikof’s group has confirmed their stealth properties when
implanted in vivo, where they show long-term stability in
different tissues without inducing an inflammatory response
[37]. Although these ELR-based hydrogels confirmed their
effectiveness as tissue-engineering scaffolds with excellent
mechanical properties and biocompatibility, the absence of a
cell-anchorage sequence limits both cell adhesion and subse-
quent colonization of the artificial scaffold, even if the im-
plant is retained in the host’s body for more than a year [37].
In a recent publication from the same group, this deficiency
was overcome by both coating their surface with the struc-
tural ECM protein fibronectin or integrating this protein into
the ELR gel by absorption or cross-linking, which resulted in
modulated cell adhesion, viability, proliferation, and migra-
tion of both endothelial and mesenchymal stem cells [38].
It is well known that binding between cell-surface recep-
tors and ECM proteins activates the intracellular signaling
pathways that deeply influence a cell’s behavior and fate
[39]. This activation is mostly due to interaction between
specific motifs on the ECM proteins and cell receptors. The
short amino acid sequences that form protein motifs can eas-
ily be incorporated into the sequences of recombinant pro-
tein-based polymers (or recombinamer) using standard ge-
netic-engineering techniques, thus endowing these recombi-
namers with functionality, for example to increase cell-
transfection efficiency [40], promote angiogenesis [41], in-
hibit endothelial cell migration [42] or to nucleate bone min-
eralization [43]. Similarly, the inclusion of integrin-mediated
cell-adhesion motifs (short sequences present in several
structural ECM proteins) into some advanced scaffolds has
been reported to be one of the factors that prevent contrac-
tion and scar formation in skin and nerve-tissue regeneration.
In this respect, the well-known Arg-Gly-Asp (RGD) guides
the colonization of these artificial ECMs by recruiting the
host’s own cells, thereby avoiding cell-cell binding, limiting
wound contraction and aiding tissue regeneration [44]. The
inclusion of integrin-binding sites into the ELR backbone
also favors material-cell interactions [45].
Several examples of RGD-containing bio-functionalized
ELRs have been reported to present higher rates of cell at-
tachment with respect to controls [46, 47]. This enrichment
is preserved when the ELR-RGD is coated onto other bio-
logically inert substrates, or is cast in 3D cell supports [48,
49]. Two recent studies in this field described the results
obtained by coating scaffolds made of conventional biomate-
rials as poly(lactic) acid (PLA) or poly(lactide-co-glycolide)
(PLGA) with ELR-RGD. In both cases, the ELR layer cover-
ing the primary component conferred a stronger cell-

Recent Contributions of Elastin-Like Recombinamers Current Topics in Medicinal Chemistry, 2014, Vol. 14, No. 7 3
adhesion ability. In the first work [50], the authors described
an alternative and more efficient bio-functionalization of
model PLA surfaces achieved using ELR-RGD instead of
conventional short peptides containing an RGD motif. PLA
scaffolds were either absorbed or covalently grafted with
protein material in the form of peptides, ELRs or bovine
serum albumin (BSA) and their bio-functionalization effi-
ciency evaluated. All possible combinations were compared
in early and late cell-surface interaction studies and three
different factors found to influence the process. The first of
these is the presence of RGD, which always significant im-
proved early cell adhesion and cell proliferation, and the
second is the type of molecule presenting the RGD motif,
with ELR-RGD being a significant enhancer with respect to
the short peptides-RGD. The final factor is the covalent liga-
tion of molecules to PLA. This resulted in larger cell areas
on the ELR-grafted surfaces, cells which acquired a well
spread morphology with strong stress fibers, possibly due to
better transmission of the mechanical stimuli. The resulting
hybrid ELR-PLA scaffolds restricted nonspecific protein
adsorption, enhanced cell adhesion and anchorages, and in-
creased the proliferation ratio, thus demonstrating that the
use of ELRs is a more efficient strategy for guiding cellular
activity than the use of short peptides [50].
In the second example, scaffolds for neural tissue engi-
neering were produced by coating microporous PLGA with
various concentrations of ELR-RGD via a simple tempera-
ture-dependent sol-gel transition. Subsequent in vitro assays
showed that the ELRs improved neural progenitor cell adhe-
sion and proliferation in a concentration-dependent manner.
Moreover, when the cell culture on ELR-RGD is grown in
combination with retinoic acid, differentiation of the pro-
genitor cells into neuronal and astroglial lineages is induced
[51].
The use of ELRs ensures complete composition control
and allows the influence of different parameters to be studied
as independent components. In this regard, a preliminary
study of the influence of cell ligand density and scaffold ri-
gidity on neurite growth has been carried out to improve the
performance of a 3D hydrogel formed for neural tissue re-
generation. Hydrogel stiffness was modulated by using in-
creasing cross-linker/ELR stoichiometric ratios, whereas the
cell adhesion motif density was controlled by homogenously
mixing RGD and its scrambled non-adhesive negative con-
trol. The resulting data confirmed that the softer and RGD-
richer 3D hydrogel is the best scaffold for stimulating neurite
growth [52].
A similar ELR-RGD scaffold has been used recently to
cultivate pancreatic beta-TC6 cells in which the formation of
islet-like structures was observed. In this study, the surfaces
were easily coated by making use of one of the most charac-
teristic ELR properties, namely the temperature-dependent
phase transition that determines the sol-gel transition. In con-
trast to conventional substrates, the beta cells seeded on the
ELR-RGD surfaces showed high cell viability, self-
organisation into multicellular spheroids with an islet-like
architecture, and clustering, presumably caused by the pres-
ence of RGD, which triggers a signaling cascade, in the ELR
backbone. Moreover, the pancreatic cells grown on ELR-
RGD improved insulin expression and secretion stimulated
by glucose, increased the expression of cell-to-cell adhesion
molecules and of ECM fibrous proteins. The overall results
suggested that ELR-RGD promotes pseudoislet formation
and therefore that it is a suitable scaffold for manufacturing
pancreatic islet cells for transplantation in vitro. [53]
The possibility of employing ELR-RGD materials to im-
prove the regeneration, or induce the differentiation, of endo-
thelium [41], cartilage [36], bone [48, 54], muscle [55], pan-
creas [53], or neuronal tissue [51] has been described. Inter-
esting results have also been obtained in the case of epithe-
lial oral tissue, for which conventional artificial 3D scaffolds
are unable to sustain the potential proliferative capacity of
oral epithelial cells during extended culture times. In order to
improve cell adhesion on the natural structural protein colla-
gen, an ELR-RGD/collagen mixture was electrospun and
cross-linked to fabricate a suitable highly porous epithelial
3D scaffold [56]. The nanofiber 3D ELR scaffold is initially
cultivated with human fibroblasts that are able to proliferate,
infiltrate the network and neo-synthesize their own ECM,
finally forming a lamina propria equivalent. Co-culture with
oral epithelial cells seeded over this lamina equivalent gen-
erates a non-keratinized, multilayered oral epithelial mucosal
equivalent. The 3D hybrid scaffold is able to sustain in vitro
culture for six weeks. Moreover, it was found to express the
specific oral epithelial marker and is histologically very
similar to the native version. In contrast, the collagen-only
electrospun 3D scaffold used as negative control was poorly
colonized by the cells and its resulting epithelium was thin-
ner [57]. Other types of epithelial cells have also been found
to benefit if cultured on ELR-RGD as a substrate for subse-
quent subretinal transplantation. Indeed, ELR-RGD supports
are able to sustain the growth and maintain the phenotype,
and functional characteristics of retinal pigment epithelial
cells [58, 59].
RGD is not the only cell adhesion motif engineered into
an ELR back-bone. Thus, Tirrell and coworkers have com-
pared the in vitro cell-adhesion properties of bidimensional
ELR scaffolds containing the human fibronectin motifs RGD
or Arg-Glu-Asp-Val (REDV) [47, 60, 61]. Although both
these motifs were found to enhance the adhesion of HUVEC
endothelial cells, the former was more efficient than the lat-
ter, which, in contrast, specifically interacts with the endo-
thelium [62, 63].
The porosity of the artificial scaffold has been reported to
be one of the parameters that induce regeneration for pore
diameters ranging from 40 to 140 μm [64]. As well as in
regenerative medicine, another important property of a tem-
porary scaffold is that it must degrade and be absorbed once
the host can neo-synthesize its own ECM. In light of this, a
number of trigger-biodegradable biomaterials have been bio-
synthesized [65, 66]. Both these features can be achieved in
3D scaffolds obtained by chemical cross-linking of an enzy-
matically biodegradable ELR-REDV. Thus, HUVECs cul-
tured in highly interconnected porous 3D hydrogels are able
to infiltrate the porous network and acquire a well spread
morphology [67]. The same ELR-REDV and collagen have
also been enzymatically cross-linked using variable ratios of
both proteins to form 3D scaffolds. The resulting collagen-
ELR scaffolds improved the elasticity and bioactivity in
terms of adhesion selectivity and proteolytic sensitivity with

4 Current Topics in Medicinal Chemistry, 2014, Vo l. 14, No. 7 Arias et al.
respect to those formed from collagen alone. A comparison
of HUVEC and fibroblast cultures grown on the hybrid scaf-
folds confirmed that REDV specificity (in terms of both
morphology and proliferation rate) was ratio-dependent. This
study therefore suggests that the collagen-ELR ratio can be
varied to fit the characteristics of different tissues, thereby
preventing hypertrophy of the scar and modulating angio-
genesis [63].
A further bioactive sequence identified in a secreted
heparin-binding protein (CCN1) that is expressed at injury-
repair sites was recently incorporated into a triblock copoly-
mer to enhance cell adhesion. This 20-mer peptide, namely
V2, specifically binds to integrin
v
3
, membrane receptors
present in HUVECs. As a result, the HUVECs cultured on
the ELR-V2 adhered, spread, migrated and also showed a
quiescent phenotype in the presence of an integrin-mediated
stimulus [68].
A series of tailored-made ELRs have been synthesized in
order to obtain adult cardiomyocytes. In this case, due to
their poor proliferation, instead of inserting a cell-adhesion
motif into the ELR sequence, the authors preferred to include
the receptor-binding domain of insulin-like growth factor
binding protein 4 (IGFBP4), which promotes the differentia-
tion of embryonic stem cells into cardiomyocytes. The ELR-
IGFBP4 sol-gel transition allowed stable coating of the cul-
ture substrate and the possibility of cell differentiation was
subsequently evaluated. A high IGFBP4 concentration is
required to induce this differentiation in a standard culture as
it is very unstable. A comparison of the results obtained in
this study showed that absorbed ELR-IGFBP4 is more effi-
cient than absorbed IGFBP4 alone or the standard method of
cardiomyocyte differentiation involving continuous addition
of soluble IGFBP4 to the medium. As such, the functional-
ized ELR-IGFBP4 improves IGFBP4 stability, thereby re-
sulting in a stronger and constant effect on the culture [69].
The studies highlighted above indicate that the use of
ELRs to construct 2/3D scaffolds or to improve the bioactiv-
ity of conventional materials has significantly increased the
performance of conventional scaffolds. Moreover, their
modular nature means that the function of biomaterials can
be enhanced by redesigning their amino acid composition
and adding bioactive motifs that may convert the scaffold
from being a mere support to a vector that performs the fine
and delicate role of cell-material communicator during re-
constitution of the new host tissue.
APPLICATION OF ELRs IN SURFACE ENGINEER-
ING
Surface engineering is an area in the field of biomaterial
science devoted to the control and modification of surfaces
and interfaces in order to study the molecular mechanisms
underlying protein adsorption and cell-extracellular interac-
tions. A large number of biological reactions occur either on
surfaces or at interfaces, and cell behavior can be condi-
tioned by a given well-defined topography and bioactivity of
these surfaces. Moreover, in the field of implantable devices,
surface contacts with the organism dictate cellular behavior
and biointegration. In light of the above, the development of
appropriate systems that mimic in vivo cellular environments
in order to enable in vitro studies of cell-matrix interactions
is a key requirement for future progress in several areas of
bioscience and biotechnology. [70]
The ability to manufacture surface coatings that are bio-
compatible and stable under physiological conditions is a
valuable objective in surface engineering in order to design
devices with an improved biological performance. In this
sense, ELRs are excellent candidates for the development of
smart surfaces as, together with their extensive potential to
self-assemble, their sequence, functionality and bioactivity
can be closely controlled using recombinant technologies.
Modification of Surface Topography by ELRs
The design of systems with a well-defined topography
and controlled chemical and mechanical properties is a hot
research topic due to their potential biomedical applications,
especially for improving cell-material interactions [71, 72].
Modification with stimuli-responsive ELRs allows surfaces
to vary their physical and chemical properties in response to
external stimuli, which is often essential for endowing ad-
vanced materials and devices with desirable features.
In this sense, Bandiera [73] has described the behavior of
different cell lines cultured on surfaces coated with an ELR
that adopts a defined pattern of concentric circles. The good
adhesiveness displayed by all the cell lines studied is note-
worthy, with the specific behavior depending on the cell type
concerned. The difference in cell response appears to reside
in the ability of cells to somehow discriminate the physical
structure of the substrate. Thus, human endothelial and
epithelial cell lines align along the grooves, probably in the
direction in which they encounter the least resistance, as al-
ready described for other biological systems. [74]
To study the influence of different topographies on cell
behavior, Garcia-Arevalo et al. carried out a comparative
analysis of the precise contribution and importance of two
different surface topographies, namely nanofibers and films,
on cell behavior [45]. Inert surfaces were covered with a film
or an electrospun layer of an aqueous solution of ELRs bear-
ing a specific adhesion sequence such as RGD. Surfaces with
different topographies showed different properties in terms
of wettability, roughness and surface free-energy. The mor-
phological and proliferative responses of the seeded human
fibroblasts were analyzed, and the authors concluded that
higher proliferation rates were obtained for those surfaces
coated with films which presented a higher surface energy
and were more hydrophilic. The lower proliferative values
for surfaces covered with fibers was due to the greater ad-
sorption of non-specific proteins from the culture medium,
thereby hindering cell-surface interactions. The surface ad-
hesion and proliferative values for both these topographies
were higher for those surfaces coated with the RGD-
containing ELR than for the control surfaces covered with an
ELR lacking the RGD domain [45].
Similarly, Martin et al. have fabricated 3D structures
made of ELR-based hydrogels by means of replica molding.
These structures have a fully controlled surface micro-
topography in addition to tunable mechanical properties and
a smart nature. These surfaces exhibited grooves or pillars
with different dimensions and distances. The controlled to-
pography and bioactivity resulting from the RGD domain are
noteworthy factors to study and, eventually, to regulate cell

Recent Contributions of Elastin-Like Recombinamers Current Topics in Medicinal Chemistry, 2014, Vol. 14, No. 7 5
behavior [27]. The hydrogels were obtained by chemical
cross-linking on top of PDMS stamps; their swelling and
mechanical properties could be modulated by varying the
polymer:cross-linker ratio. The thermoresponsive behavior
of the starting ELRs was maintained in the hydrogel, which
exhibited different dimensions above and below its transition
temperature, without topography modification [27]. Mi-
cropatterned gels were subsequently obtained from elastin-
like amphiphilic multiblock copolymers with reversible
thermogelling properties under mild, physiological condi-
tions. In this case, the one-step physical gelation process
comprises heating the ELR solution above its gelation tem-
perature on top of a PDMS mould (see Fig. 1). The mechani-
cal properties of the hydrogel can be tuned by varying the
polymer solution concentration [28]. Micropatterned mem-
branes have been obtained from ELRs containing an RGD
domain that incorporate both biomolecular and physical sig-
naling by Tejeda-Montes et al. [75] These membranes are
obtained by using hexamethylene diisocyanate (HDMI) as
chemical cross-linker. Rat mesenchymal stem cells (rMSCs)
attach to membranes both with and without RGD but only
exhibit well-developed focal adhesion mediated by integrin
binding for those containing an RGD domain. In membranes
with surface topographies, cells show morphological varia-
tions and contact guidance depending on the surface topog-
raphy, exhibiting an aligned and thin morphology on chan-
nels and growing within and around the posts, with actin
cytoskeletons engulfing them. This response of the cells to
the membrane’s biomolecular and physical features suggests
that both elements could be tailored to synergistically alter
specific cell behavior. These microstructured substrates
simulate the microscale structure of tissues and could be
used as implantable platforms or as a substrate for studying
spatially controlled cell behavior.
In order to study the influence of surface chemistry and
topography on bone marrow mesenchymal stem cell prolif-
eration and differentiation, micropatterned PIPAAm films
were prepared and modified by adsorption of an elastin-like
polymer containing an adhesion domain (ELR-RGD). Tak-
ing advantage of the thermal responsiveness of the ELR-
RGD, rhythmic temperature changes were applied to flex
and contract the 2D scaffold in an attempt to mimic me-
chanical stress on the cells and to provide dynamic culture
conditions that are expected to improve bone formation. The
presence of the recombinamer proved crucial for maintaining
cell attachment under dynamic culture conditions, as cor-
roborated using control tissue culture polystyrene surfaces
(TCPS), for which the number of attached cells decreased
with time under the same conditions [48].
Biofunctionality Control Using ELRs
Many physicochemical methodologies have been pro-
posed to modify the chemical and topographical features of
substrates. Initial research mainly focused on the adsorption
of natural proteins and short peptides. However, the use of
engineered proteins can provide the advantages of both
strategies and avoid the main drawbacks concerning the im-
munogenic problems presented by some proteins or the loss
of effectiveness of oversimplified peptides. [20] In this
Fig. (1). Micropatterned gels with different features obtained by ELR replica molding. Adapted with permission from Soft Matter [28].

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37 citations


Journal ArticleDOI
TL;DR: It is shown that it is possible to replicate different patterns on the hydrogel surface, thus allowing the use of this type of hydrogels to improve applications that require cell guidance or even differentiation depending on the surface topography.
Abstract: Elastin-like recombinamer click gels (ELR-CGs) for biomedical applications, such as drug delivery or tissue engineering, have been developed by taking advantage of the click reaction (CuAAC) in the absence of traditional crosslinking agents ELRs are functionalized with alkyne and azide groups using conventional chemical techniques to introduce the reactivity required to carry out the 1,3-dipolar cycloaddition under mild biocompatible conditions, with no toxic by-products and in short reaction times Hydrogels with moduli in the range 1,000–10,000 Pa have been synthesized, characterized, and tested in vitro against several cell types The cells embedded into ELR-CGs possessed high viability and proliferation rate The mechanical properties, porosity and swelling of the resulting ELR-CGs can easily be tuned by adjusting the ELR concentration We also show that it is possible to replicate different patterns on the hydrogel surface, thus allowing the use of this type of hydrogel to improve applications that require cell guidance or even differentiation depending on the surface topography

30 citations


Cites background from "Recent Contributions of Elastin-Lik..."

  • ...cell adhesion and proliferation or sequences sensitive to enzymes [11]....

    [...]


Journal ArticleDOI
TL;DR: The strategy developed in this work about using ELR as polymeric vector and aptamers as supplier of specificity to deliver therapeutic material into MUC1-positive breast cancer cells shows promising potential and continues paving the way for ELRs in the biomedical field.
Abstract: The search for new and biocompatible materials with high potential for improvement is a challenge in gene delivery applications. A cell type specific vector made of elastin-like recombinamer (ELR) and aptamers has been specifically designed for the intracellular delivery of therapeutic material for breast cancer therapy. A lysine-enriched ELR was constructed and complexed with plasmid DNA to give positively charged and stable polyplexes. Physical characterization of these polyplexes showed a particle size of around 140 nm and a zeta potential of approximately +40 mV. The incorporation of MUC1-specific aptamers into the polyplexes resulted in a slight decrease in zeta potential but increased cell transfection specificity for MCF-7 breast cancer cells with respect to a MUC1-negative tumor line. After showing the transfection ability of this aptamer-ELR vector which is facilitated mainly by macropinocytosis uptake, we demonstrated its application for suicide gene therapy using a plasmid containing the gene of the toxin PAP-S. The strategy developed in this work about using ELR as polymeric vector and aptamers as supplier of specificity to deliver therapeutic material into MUC1-positive breast cancer cells shows promising potential and continues paving the way for ELRs in the biomedical field.

27 citations


Journal ArticleDOI
Abstract: Vascular disease is a leading cause of death worldwide, but surgical options are restricted by the limited availability of autologous vessels, and the suboptimal performance of prosthetic vascular grafts. This is especially evident for coronary artery by-pass grafts, whose small caliber is associated with a high occlusion propensity. Despite the potential of tissue-engineered grafts, compliance mismatch, dilatation, thrombus formation, and the lack of functional elastin are still major limitations leading to graft failure. This calls for advanced materials and fabrication schemes to achieve improved control on the grafts' properties and performance. Here, bioinspired materials and technical textile components are combined to create biohybrid cell-free implants for endogenous tissue regeneration. Clickable elastin-like recombinamers are processed to form an open macroporous 3D architecture to favor cell ingrowth, while being endowed with the non-thrombogenicity and the elastic behavior of the native elastin. The textile components (i.e., warp-knitted and electrospun meshes) are designed to confer suture retention, long-term structural stability, burst strength, and compliance. Notably, by controlling the electrospun layer's thickness, the compliance can be modulated over a wide range of values encompassing those of native vessels. The grafts support cell ingrowth, extracellular matrix deposition and endothelium development in vitro. Overall, the fabrication strategy results in promising off-the-shelf hemocompatible vascular implants for in situ tissue engineering by addressing the known limitations of bioartificial vessel substitutes.

26 citations


Cites background from "Recent Contributions of Elastin-Lik..."

  • ...They are positioned midway between natural products and synthetic polymers, as they are engineered in a controlled and highly reproducible way while still maintaining inherent properties of the natural elastin such as elastic mechanical behavior, hemocompatibility, and bioactivity (Arias et al., 2014)....

    [...]


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Xiaoming Sun1, Zhuang Liu1, Kevin Welsher1, Joshua T. Robinson1  +3 moreInstitutions (1)
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Posted Content
Xiaoming Sun1, Zhuang Liu1, Kevin Welsher1, Joshua T. Robinson1  +3 moreInstitutions (1)
TL;DR: It is found that simple physisorption via π-stacking can be used for loading doxorubicin, a widely used cancer drug onto NGO functionalized with antibody for selective killing of cancer cells in vitro.
Abstract: Two-dimensional graphene offers interesting electronic, thermal and mechanical properties that are currently explored for advanced electronics, membranes and composites. Here we synthesize and explore the biological application of nano-graphene oxide NGO, single-layer graphene oxide sheets down to a few nanometers in lateral width. We develop functionalization chemistry to impart solubility and compatibility of NGO in biological environments. We obtain size separated pegylated NGO sheets that are soluble in buffers and serum without agglomeration. The NGO sheets are found to be photoluminescent in the visible and infrared regions. The intrinsic photoluminescence of NGO is used for live cell imaging in the near-infrared with little background. We found that simple physisorption via pi-stacking can be used for loading doxorubicin, a widely used cancer drug onto NGO functionalized with antibody for selective cancer cell killing in vitro. Owing to the small size, intrinsic optical properties, large specific surface area,low cost, and useful non-covalent interactions with aromatic drug molecules, NGO is a promising new material for biological and medical applications.

2,633 citations


Journal ArticleDOI
26 Oct 1999-Chemical Reviews
TL;DR: Kevin Shakesheff investigates new methods of engineering polymer surfaces and the application of these engineered materials in drug delivery and tissue engineering.
Abstract: s, and 360 patents, and edited 12 books. He has also received over 80 major awards including the Gairdner Foundation International Award, Lemelson-MIT prize, ACS’s Applied Polymer Science and Polymer Chemistry Awards, AICHE’s Professional Progress, Bioengineering, Walker and Stine Materials Science and Engineering Awards. In 1989, Dr. Langer was elected to the Institute of Medicine of the National Academy of Sciences, and in 1992 he was elected to both the National Academy of Engineering and the National Academy of Sciences. He is the only active member of all three National Academies. Kevin Shakesheff was born in Ashington, Northumberland, U.K., in 1969. He received his Bacheclor of Pharmacy degree from the University of Nottingham in 1991 and a Ph.D. from the same institution in 1995. In 1996 he became a NATO Postdoctoral Fellow at MIT, Department of Chemical Engineering. He is currently an EPSRC Advanced Fellow at the School of Pharmaceutical Sciences, The University of Nottingham. His research group investigates new methods of engineering polymer surfaces and the application of these engineered materials in drug delivery and tissue engineering. 3182 Chemical Reviews, 1999, Vol. 99, No. 11 Uhrich et al.

2,411 citations


Performance
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No. of citations received by the Paper in previous years
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20203
20195
20182
20172
20162
20156