石墨烯生物材料到仿生学

www.advmat.
de
www.MaterialsViews.com
Graphite Oxide to Graphene. Biomaterials to Bionics
REVIEW
Brianna C. Thompson,* Eoin Murray, and Gordon G. Wallace*
synthetic scaffolds to make tissue engi-

The advent of implantable biomaterials has revolutionized medical treatment, neered structures is shown in Figure 1.
allowing the development of the fields of tissue engineering and medical The engineering of different kinds
bionic devices (e.g., cochlea implants to restore hearing, vagus nerve stimu- of tissues has specific challenges and
requirements, from the form factor of the
lators to control Parkinson’s disease, and cardiac pace makers). Similarly,
implant (e.g., blocks, tubes, membranes,
future materials developments are likely to continue to drive development sponges etc.) and the physical properties
in treatment of disease and disability, or even enhancing human potential. of the materials (e.g., stiffness, softness,
The material requirements for implantable devices are stringent. In all cases porosity, degradability) to the desired bio-
they must be nontoxic and provide appropriate mechanical integrity for the logical outcomes (e.g., promoting cell pro-
liferation, differentiation, cell invasion,
application at hand. In the case of scaffolds for tissue regeneration, biodeg-
cell repulsion). All tissue engineering tar-
radability in an appropriate time frame may be required, and for medical gets require different materials and fabri-
bionics electronic conductivity is essential. The emergence of graphene and cation strategies, and so the fields of tissue
graphene-family composites has resulted in materials and structures highly engineering and biomaterials develop-
relevant to the expansion of the biomaterials inventory available for implant- ment require a diverse array of expertise.
able medical devices. The rich chemistries available are able to ensure The form factors of implants can
vary tremendously between applica-
properties uncovered in the nanodomain are conveyed into the world of
tions – a variety of which are illustrated
macroscopic devices. Here, the inherent properties of graphene, along with in Figure 2. Some applications require
how graphene or structures containing it interface with living cells and the the replacement of large blocks of tissue
effect of electrical stimulation on nerves and cells, are reviewed. (such as muscle or fat), and in these
applications, it is often necessary to
devise a strategy to allow for transport
of nutrients and waste into/out of the implant. Other applica-
1. Tissue Engineering tions need a membrane (to replace skin in wound healing or
to replace internal membranes), or fibrous structures (in the
The goal of a tissue engineer is to fix or replace tissues or engineering of nerve or muscle fibers), or something much
organs that have been damaged or destroyed by accident or dis- more complicated and specific (in the engineering the compli-
ease. In order to achieve this, one approach is to combine a fab- cated structure of an artificial kidney or heart). As fabrication
ricated scaffold composed of one or more materials having the technologies become more advanced, more tools to enable the
desired properties with cells (either sourced and differentiated production of these more complicated structures such as 3D
externally or internally to be populated with endogenous cells in printing, electrospinning, wet-spinning, extrusion, or litho-
situ), chemical cues (drugs, growth factors or other endogenous graphy become available. Each fabrication strategy imparts its
signals), and physical cues to encourage cells to develop into own requirements on the material used, so the ultimate mate-
functional tissues. The general process of combining cells and rial properties must be balanced between the requirements for
the tissue development needed, processability, and being ame-
nable to fabrication protocols that enable the ideal form factor
Dr. B. C. Thompson to be realized.
School of Mechanical and Aerospace Engineering However, despite the need for tailoring material and implant
Nanyang Technological University
639798, Singapore properties for different parts of the body, one property is
E-mail: bthompson@ntu.edu.sg required in all tissue engineering – biocompatibility. The con-
Dr. E. Murray cept of biocompatibility is often poorly defined, but is gener-
Institute for Sports Research ally taken to mean that the material or implant lacks toxicity
Nanyang Technological University and immunogenicity and will have no adverse effect on the
639798, Singapore
biological system into which it is being introduced. Cells from
Prof. G. G. Wallace
the patient are used to seed cell scaffolds in order to reduce the
Intelligent Polymer Research Institute
ARC Center of Excellence for Electromaterials Science potential for an immune¡ reaction against transplants of tissues
University of Wollongong or stem cells from donors, or xenotransplantation, reducing the
2500, Australia risk of implant/organ rejection. The aim is to allow a “normal”
E-mail: gwallace@uow.edu.au tissue to develop that has the same genetic and epigenetic cues
DOI: 10.1002/adma.201500411 as the other tissues in the patient.
Adv. Mater. 2015, 27, 7563–7582 © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim wileyonlinelibrary.com 7563
15214095, 2015, 46, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/adma.201500411 by Rwth Aachen Hochschulbibliothek, Wiley Online Library on [14/02/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
www.advmat.de
REVIEW
While the concept of biocompatibility is loosely defined

in the literature, the regulatory requirements to test the bio- Brianna Thompson is a
compatibility of devices for implantation are not. The Food Research Fellow working on
and Drugs Administration (USFDA), the US-based consumer developing novel ophthal-
protection and regulatory body, set out a comprehensive set of mological prostheses within
requirements and tests for the biocompatibility of a device, as the School of Aerospace
summarized in Table 1 and documented in full in ref. [2]. Any Engineering, Nanyang
device to be implanted into humans must pass all of these tests Technological University.
before it can be released to market in the USA. The guidelines She has previously worked
provided by the USFDA are based on ref. [3], however, with on neural interfacing and
2009–2010 revisions to the ISO10993 guidelines (particularly regeneration, bionic inter-
ISO10993–1, dealing with “evaluation and testing in the risk facing, and drug delivery at
management process”), the USFDA released a draft revised University of Wollongong,
version in April 2013 (UCM348890), which while similar to Australia, and biological applications of ionic liquids
#G95–1, gives more detailed guidelines for the expected test at Monash University, Melbourne, Australia. Her main
outcomes, follow-up tests that researchers should consider research interests lie in finding uses for novel materials
in special cases, increased detail on how previously approved and fabrication process in biological systems.
materials had been treated, and updated definitions for more
novel materials, including nanostructured materials such as Eoin Murray is a Senior
nanoparticles, carbon nanotubes, or graphene, or materials that Research Fellow/R+D Project
are polymerized in situ.[4] At the time of writing this article, Manager at the Institute for
these guidelines remained in draft form, retrievable from the Sports Research at Nanyang
USFDA website.[5] Most other large markets around the world Technological University,
base their product testing requirements on either IS10993 or Singapore. He has previously
more directly from the guidelines laid down by the USFDA. worked on the development
of materials for bionics as an
Australian Research Council
2. Bionics Superscience Fellow at the
University of Wollongong,
One often-overlooked but significant influence over the behavior Australia, nanoparticle
of cells, tissue, or biological systems is electromagnetic signalling. synthesis and nano-structure formation at the Leibniz-
A wide range of cell processes are influenced by naturally occur- Institut für Neue Materialien, Germany, and nanoparticles
ring electrical signals that modify biological behaviors, in the form and conducting materials as an IRCSET Embark scholar at
of electromagnetic fields, ionic electrical charges, or even direct Dublin City University. His main research interests lie in
electrical charges (Figure 3).[6] Medical bionics involves the inter- the development and real-world application of multifunc-
facing of biological and electronic systems, in order to externally tional materials.
replicate or influence these electronic signals. Controlling the
electrical field or providing voltages, currents, or charge to stimu- Gordon Wallace is currently
late cells via electrodes can be used to communicate information the Executive Research
to cells (stimulation, discussed in Section 2.2.1), while measuring Director at the ARC
ionic fluxes associated with action potentials or muscle contrac- Center of Excellence for
tion allows communication from tissue to electronic systems Electromaterials Science and
(recording systems are discussed in more detail in Section 2.2.2). Director of the Intelligent
Polymer Research Institute.
He previously held an ARC
2.1. Excitable Cells Federation Fellowship
and currently holds an
Cellular systems that generate electrical signals, such as nerves ARC Laureate Fellowship.
and muscle, obviously respond to electrical signals provided to Professor Wallace’s research
them. Other cellular systems are also known to respond to elec- interests include organic conductors, nanomaterials and
trical stimulation. electrochemical probe methods of analysis, and the use of
these in the development of Intelligent Polymer Systems. A
current focus involves the use of these tools and materials
2.1.1. Nerves in developing bio-communications from the molecular to
skeletal domains in order to improve human performance
In the nervous system, electrical signals are communicated via medical Bionics.
around the body in the form of controlled membrane potential
(as shown in Figure 3C) changes resulting in charge gradients
7564 wileyonlinelibrary.com © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Adv. Mater. 2015, 27, 7563–7582
www.advmat.de
REVIEW
Figure 1. The tissue engineering process. 1) Cells are isolated from the patient or derived from established stem cell cultures. 2) A scaffold is con-
structed with the correct physical, mechanical, chemical and structural cues to encourage the development of cells into tissues. 3) Cells are seeded onto
the scaffold in vitro and transferred into a bioreactor. The scaffold encourages cell proliferation and/or differentiation, the construct is incubated until
the nascent tissue is ready for implantation. 4) The tissue-engineered scaffold with cells is re-implanted into the patient to develop into an integrated
tissue. Adapted with permission.[1] Copyright 2007, Elsevier.
across cell membranes and action potentials along nerves, Additionally, electrical fields or direct electrical stimulation
where the membrane potential may fluctuate from −70 mV to can influence nerves at the molecular level, influencing prolif-
+35 mV briefly before returning to the −70 mV resting poten- eration, migration, axonal regeneration, and function of nerve
tial (a schematic of the voltage changes in membrane poten- cells.[9,10] Migration, axonal regeneration, and function are of
tial during action potentials is shown in Figure 4). Electrical the greatest interest from a tissue-engineering perspective,
fields or electrical stimulation may interact with the distribu- where axons and glial cells need to be encouraged to regrow
tion of the gated ion channels or may modulate the function of along or repopulate damaged peripheral or spinal nerve gaps.
voltage-influenced channels to modulate the firing of nerves at Electrical fields and electrical stimulation have been used to
this physiological level. enhance or direct axonal growth in vitro and in vivo and to
Figure 2. Examples of potential form factors for tissue-engineered structures for different applications. Many form factors require different fabrication
techniques, which can impart their own restrictions and requirements on material properties.
www.advmat.de
REVIEW
Table 1. Biocompatibility testing requirements outlined by USFDA by electrical fields or electrical stimulation. Bones and dental
#G95–1(Use of International Standard ISO-10993, “Biological Evalua- tissues are influenced by the application of small electrical
tion of Medical Devices Part 1: Evaluation and Testing”,1995)[2] and 2013
draft amendments[4] for approval of implanted medical devices (and
fields,[17] meaning that bionic interference with these systems
materials) with a lifespan of greater than 30 days in humans for implan- may be applied to increase injury repair, or increase implant
tation into bone/tissue, or into areas in direct contact with the circula- integration with the body. Recently, application of electric
tory system (blood). vfields to the dental field has generated much media attention
with the promise to remove the need for dental fillings after
Test Blooda) Bone/Tissuea) a recent highly publicized discovery by King’s College London
Cytotoxicity 䊉䊉 researchers claiming to increase remineralization of dental
Sensitization 䊉䊉 caries with application of a small electric current to accelerate
䊉䊉
the movement of calcium and phosphate into the area, allowing
Genotoxicity
the body to regenerate its own tissues in a technique they have
Implantation 䊉䊉
called electrically accelerated and enhanced remineralization.[18]
Irritation or intracutaneous reactivity 䊉 + In addition to these hard-tissue orthopaedic applications,
Systemic toxicity (acute) 䊉 + electrical signals have been shown to greatly influence and
Subchronic toxicity 䊉 +
direct soft-tissue wound healing (for more details on electrical
influences on wound healing, readers are directed to the excel-
Chronic toxicity 䊊䊊
lently titled reviews written by McCaig and co-workers).[8,19]
Carcinogenicity 䊊䊊
a)
䊉 – required for initial evaluation; 䊊 – may be required for further evaluation;
+ – are included in the Draft proposal for future FDA guidelines (UCM348890)
2.2. Electrical Stimulation and Recording
(April 2013).
The interfacing of biology and electronics provides two-way
control the migration of neuronal, astrocyte, and Schwann communication – the ability to record from cellular systems
cells.[9,11] and the ability to stimulate them.
2.1.2. Muscle 2.2.1. Stimulation Mechanisms
As with nerves, muscles have the ability to undergo depolariza- Whilst it is known that electrical fields or electrical stimula-
tion and repolarization in an action potential (Figure 4). How- tion are important in wound healing, nerve and muscle devel-
ever, the end result of an action potential in muscle is not the opment and function, and cell migration,[8,19] there are many
transmission of information (as in nerves), but the contraction unknowns about the electrical parameters required for elec-
of the muscle cell. Muscles also typically have a more-negative trical control of behavior and about the mechanisms of cell
membrane potential than neurons, generally sitting at closer to response to electromagnetic cues. Molecular and cell level
−90 mV than the approximately −50 to −70 mV across neural mechanisms for electromagnetic influence on cell behavior
membranes. have been speculated upon, however no definitive answers have
The electrical effects on muscle development have received been given. In fact, it is likely that there are combinations of
less attention than neural cells and tissues. However, several the many proposed mechanisms, including direct effects on
papers have shown interesting changes in cultured muscle cells specific voltage-gated receptors,[20] electrophoretic asymmetric
that have received electrical stimulation,[12] and on a whole- distributions of membrane receptors,[8] membrane lipid redis-
body level, electrical stimulation is applied to muscle tissue tribution, changes in fluidity of the plasma membrane, and
in the form of neuromuscular electric stimulation (NMES) the influence on lipid rafts, or modification of cell signalling
for training and rehabilitation purposes in sports science.[13] cascades.[21] McCaig et al. proposed a model in which the elec-
Chronic low-frequency stimulation (1–10 Hz) of in vitro and trical and chemical gradients that drive neural migration and
in vivo muscle has been used to induce contractions in order axonal regeneration may be combined, wherein the receptors
to study the impact of muscle use on its development. These for neurotrophins may be electrophoretically concentrated on
studies have found that electrical stimulation that results in one side of the cell or another, leading to asymmetric recep-
contraction of muscle cells induces changes in the transcrip- tion of the chemical cues.[8] However, this alone would not
tion of genes and the metabolism of the cells, with the pulse account for cell responses to AC electrical stimulation, which
frequency determining the muscle phenotype.[12,14–16] has been shown to have positive effects on axonal regeneration.
With incremental advances in imaging techniques, molecular
markers, and spectroscopic techniques, the tools to study these
2.1.3. Other Tissues influences should allow more-detailed characterization of cell
response in the near future.
In addition to the excitable tissues above, which generate their In addition, as traditional metal electrodes have always been
own electrical signals (and are therefore an obvious target for separated from the tissue undergoing stimulation, the differ-
electrical stimulation to change behavior), several other tis- ences between electric-field stimulation, ionic currents, and the
sues and processes have been shown to be greatly influenced provision of current/voltage directly at the membrane of the
www.advmat.de
REVIEW
Figure 3. Naturally occurring electrical influences in biological systems. From A) Galvani's frog leg experiments with touching (1), electrically stimu-
lating (2) or coupling (3) sciatic nerves to stimulate muscle contraction, to B) Borgens et al.'s work with transected lamprey axons,[7] there is a long
history of electrical stimulation of, and electrical recordings from, biological tissues. At the cellular level, membrane potentials (Vm) can be established
across intact plasma membranes (C) or transepithelial potentials (VTEP) across epithelial layers (D) due to the actions of ion-selective membrane
channels, with cells having a net negative charge compared with their environment. Disruption of VTEP in wound injuries leads to a flow of current,
as positive ions can now flow into the cell (E) or across the epithelium (F), demonstrating an electrical signal that mediates a biological response in
non-excitable tissues. Reproduced with permission.[8] Copyright 2005, American Physiological Society.
cell remain unstudied and are completely unknown. However, around the body. Understanding how that information is coded,
with the advent of processable, potentially soluble conductors, passed on, and integrated could be of great use to many fields
such as chemically polymerized conducting polymer nanoparti- of medicine, and to our understanding of the human brain, in
cles, the work of Martin and co-workers in growing conducting addition to having use in future control of artificial prosthetics.
polymers around living cells,[22,23] or some of the graphene Electrical recording of these action potentials with electrodes
hydrogels discussed below, there is the opportunity to separate has allowed some information on neural networking to be gath-
these modes of electrical stimulation and to study differences ered both in vitro and in vivo, but materials available for the
in the responses of cells. production of recording electrodes have, until recently, limited
how much information can be recorded from the complex and
information-rich circuitry of the mammalian nervous system.
2.2.2. Recording from Tissues Improvements in the flexibility, compliance, and electrode den-
sity available have been needed to get sufficient information to
As mentioned in Section 2.1.1 and 2.1.2, nerve and muscle tis- understand the systems.
sues generate their own electrical signals, in the form of action For recording action potentials from single excitable cells,
potentials (illustrated Figure 4). The nervous system uses these intracellular recordings or patch-clamping are typically used.
electrical signals to pass and process large amounts information Microelectrodes are either inserted into the cell (in intracellular
www.advmat.de
REVIEW
Figure 4. Action potentials in neural, skeletal, and cardiac muscles,

showing typical membrane potentials, action potential lengths and refrac-
tory periods for the depolarization events in different cells.
recordings) or sealed onto a small area of the membrane of

the cell (typically via a micrometer-sized glass pipette), and
variations in the membrane potential of the cell are monitored.
These labor-intensive, specialized measurements have pro-
duced useful information about cell physiology and changes
in cell behavior in response to applied stimuli, but, however,
can only be applied to in vitro situations, and the limita-
tion to recording a single cell at once limits the use of these
techniques.
Most recordings for multiple cells (named neural works
when the cells being measured are neural in origin) or tissues
are done through microelectrode array (MEA) systems, with
the most popular in vivo arrays being the Michigan and Utah
arrays shown in Figure 5. These metallic probes are inserted
into neural tissues, where the uncoated tips (of micrometer
dimension) record the neural-network activity. The size and
impedance of the electrodes are important to allow recording
with sufficiently high signal to noise ratio, to record with the
high sensitivity required for the monitoring of the numerous
and fast signals transferred in a neural network.
2.2.3. Requirements for Electrical Stimulation and Recording

Electrodes
The requirements for electrodes are the same as those for any
implanted materials or device – they must be biocompatible Figure 5. Microelectrode arrays that have been used for in vivo record-
and non-immunogenic, and have the ability to transmit high- ings of neural activity. a) Silicon 100 electrode array with 400 µm spacing
fidelity electronic information across the electrode–cellular between the electrodes, developed at the University of Utah. b) Poly-
interface.[25] The composition and structure of this interfacial amide “bioactive electrode” array. c) Michigan thin film 256-shank array
of 1024 multiplexed sites with mounted signal processing electronics
region is determined by the cascade of molecular and cellular (arrow). Adapted with permission.[24] Copyright 2002, Nature Publishing
events initiated by the trauma of implantation, with both stim- Group.
ulating and recording electrodes often showing a decreased
effectiveness over time, due to the formation of a fibrotic scar
around the electrode. One major reason for this scar forma- avoid toxicity or immunogenicity issues, but the mechanical
tion is the difference in compliance between the electrode and properties should be flexible or soft enough to match those of
tissue, with the hard electrode often undergoing microscale the tissue that the electrode is being implanted into in order
movement within the soft tissue as the body moves. Not only to avoid this shear-induced inflammation. As well as the flex-
does the chemistry of the electrodes need to be optimized to ibility to move with the body, the electrode also needs to have
www.advmat.de
REVIEW
sufficient toughness to hold together under surgical and physi- mechanical and electrical properties often quoted in discus-
ological conditions. sions regarding graphene are generally referring to this gra-
Depending on the application, the electrode may need to phene type; however, the production and further processing of
degrade away to make room for re-developing tissues, or may this form of graphene is difficult, especially on a large scale,
need the durability to last and function in the body for many and, thus, presently, has limited application in bionics. “Few-
years. The degradation or durability of the electrode needs to layer graphene” consists of a number of stacked sheets of
be considered for each proposed application, and the materials pristine graphene (usually less than 10) and is a by-product
used as a conductor chosen accordingly. There are very few of the production of monolayer graphene. The sheet number
physiologically degradable materials that will have sufficient influences the material properties resulting in harder, stiffer
electrical properties to be used as effective electrodes. materials with slight changes in electrical properties (with
A final, vital property of an electrode is the electrical proper- the degree of stacking influencing the bandgap of the mate-
ties. The impedance of the electrode must be sufficiently low to rials) that are more robust for use in materials processing.
enable safe stimulation or efficient recording in physiological Graphene oxide (GO) is a highly oxidized form of graphene
conditions. This means that the electrode must have sufficient produced by the oxidation of graphite, opening of carbon
conductivity in a slightly salty, aqueous, and 37 °C environment nanotubes, or by bottom-up synthesis from glucose resulting
to allow fine control over current, voltage, or electric field, or in the exfoliation of single monolayer sheets in solution. The
the recording of tiny fluctuations in any of these parameters. production of graphene oxide can be relatively inexpensively
For the recording of electrical signals, field potentials (elec- scaled-up to industrial levels. The resulting graphene oxide
troencephalograms and electrocortiograms) are low frequency sheets are charge-stabilized in solution by carboxylic groups
(1–200 Hz) and low current (5–300 µV for electroencephalo- on the periphery and have multiple hydroxyl and epoxide sites
grams measured on top of the skull, or 0.01–5 mV for electro- and non-aromatic sp3 carbons in addition to the graphitic sp2
cortiograms measured on the surface of the brain, or can be carbons on the basal plane. These defect sites allow modifi-
measured locally with sharp, low-surface-area penetrating elec- cation of the graphene sheets and easy processing through
trodes (<1 mV, <200 Hz).[26] Action potential measurements are solution methods, making it ideal for materials development.
also low potential (500 µV) but require much higher frequency However, despite retaining most of the mechanical strength,
recording (0.1–7 kHz).[26] the reduction of aromaticity results in a loss of the electrical
properties, with GO being, at best, a semi-conductor.
In order to restore the aromaticity and hence the conduc-
3. Graphene, Graphene Oxide, and Reduced tivity, GO can be converted to reduced graphene oxide (rGO)
by a range of chemical or thermal treatments leading to a
Graphene Oxide
source of large-scale electrically conducting graphene. How-
“Graphene” can be used to refer to a broad range of carbon- ever, rGO is not pristine graphene, as oxidation and reduction
based materials, with widely varying properties due to large of graphite leads to residual defects on the basal plane or at the
differences on a molecular level from synthesis or function- periphery, which, at best, results in conductivity in the 10s or
alization. Similarly, nomenclature varies widely between 100s of S cm−1. While many orders of magnitude higher than
research groups and fields, and this review will use the GO, this is an order of magnitude less than that attainable with
nomenclature proposed by Sanchez et al. (Figure 6).[27] pristine graphene. Nevertheless, this could be an ideal material
Briefly, “pristine” or “monolayer” graphene is a single two- for bionic applications as the reduced conductivities are in the
dimensional sheet of defect-free, polycyclic, hexagonally required range for tissue stimulation and the defects on rGO
arranged, sp2-bonded aromatic carbon, and is generally pro- sheets have also been shown to charge-stabilize the graphene in
duced either by mechanical exfoliation of graphite, epitaxial a number of solutions, allowing easy processing and composite
growth, or chemical vapor deposition. The unique, exemplary production.[28]
Figure 6. Several forms of graphene that have been used for tissue engineering and bionics – pristine graphene (left), few layer graphene (middle),
and graphene oxide (right). Reproduced with permission.[27] Copyright 2012, American Chemical Society.
www.advmat.de
cellular uptake and excretion in vivo.[30–32] A systematic study

REVIEW
This review will primarily cover the most recent progress

in the use of pristine graphene, graphene oxide, and reduced of the size-dependence of graphene uptake has not yet been
graphene oxide in scaffolds in the field of tissue engineering carried out, but some generalizations can be derived from the
and bionics, including composites and hybrid materials formed current literature. Nano-graphene (lateral dimension <100 nm)
with inorganic and organic polymeric substrates and matrices. has been shown to have low cytotoxity and high cellular uptake,
and can be excreted renally and fecally.[31,33] Above this size
range, graphene has been shown to have a size-dependent rela-
3.1. Properties of Graphene Relevant to Use for Tissue tionship of uptake and excretion. Larger graphene sheets can
Engineering and Bionics be internalized by phagocytosis or endocytosis, but are prefer-
entially excluded according to increasing size.[32–34] However, it
Cellular compatibility is complex and very sensitive to many has recently been shown that large GO sheets are susceptible
physical, electrical, and chemical cues. Due to the many to oxidation and holing by peroxidase, which could mean that
methods of production and the large numbers of physical in vivo degradation is possible, leading to a change in their size
and chemical forms available, graphene has a broad range profile and, hence, cellular clearance.[35]
of physical properties that must be taken into account when
considering its biocompatibility. Here we discuss many of the
properties relevant to the biological applications of graphene 3.1.2. Surface Area and Surface Chemistry
materials, which are compared with those of carbon nanotubes
in Table 2. The aspect ratio of graphene also results in surface areas
many orders of magnitude greater than those of most other
nanomaterials, including ultrasmall nanoparticles and carbon
3.1.1. Physical Dimension nanotubes. The theoretical maximum of over 2600 m2 g−1 for
pristine graphene, while usually reduced by wrinkling, stacking
As it consists of large monolayers or few-atomic layers, the and defects, has significant scope for functionalization or inter-
three-dimensional aspect ratio of graphene is unique. While actions with biological entities.
the thickness is not more than that of a few-atoms, excluding Depending on the surface chemistry, biologically relevant
wrinkling, the lateral dimensions of a single sheet can vary moieties or polymeric materials can be adsorbed or covalently
from 10 nm to over 10 µm. This can have a huge effect on linked to the surface to promote biocompatibility or encourage
Table 2. Comparison of properties of graphene and carbon nanotubes relevant to biological applications. Reproduced with permission.[29] Copyright
2013, American Chemical Society.
www.advmat.de
REVIEW
cell growth. However, the surface of graphene is complex and of materials for implantation, with such composites retaining
has substantial variation across the range of graphenic mate- the processability and biocompatibility of the matrix, but
rials. Chemically, pristine and few-layer graphene are aromatic, with huge improvements in both strength and conductivity.
hydrophobic, and difficult to functionalize; physically, they are Indeed, it has been shown that single or few graphene layers
generally flat (possibly wrinkled). Graphene oxide, on the other do not affect the surface mechanical properties experienced
hand, is dependent on the material synthesis – the type and by the cells in soft thermoplastics, while still imparting
extent of oxidative defects on the surface depends on the extent conductivity.
of oxidation but generally results in good hydrostatic stabiliza-
tion in aqueous solutions; moreover, the multiple basal groups
allow easy chemical attachment and hydrogen bonding of func- 3.1.4. Electronic Properties
tional biomolecules or even cell adhesion. However, the long
term stability of GO, due to its reactive nature, means that the Electrical stimulation has been shown to improve the differen-
shelf life and thus the potential lifetime of materials that are tiation and directionality of electro-responsive cells and is the
based on GO, could be limited. Reduced graphene oxide is an basis of the field of bionics. The conductivity of pristine gra-
intermediate material, and its surface is again dependent on phene is theorized to be up to 20 000 S cm−1, and while GO
the extent of reduction during synthesis. Extensive reduction, is an insulator, processable rGO has been shown to restore
while restoring the sp2 hybridization and increasing the con- the conductivity to 100s of S cm−1. This very high conductivity
ductivity, reduces the number of edge and basal defects, which combined with graphene's biocompatibility makes it an ideal
results in the reduction of solubility and the capacity for func- material, either as a thin layer or as part of a composite in
tionalization. Controlled reduction leads to a balance between bionics.[38,42–45]
biofunctional versatility and electronic properties that can be
tuned for specific purposes.[28,36–38]
4. Graphene for Tissue Engineering
3.1.3. Mechanical Properties As outlined in Section 3.1, pristine graphene, GO, and rGO
have many properties that need to be considered for their
Cellular biosystems require very specific and varied mechan- application in tissue engineering. The biocompatibility is the
ical properties from their support structures,[39] with the first and foremost consideration for most applications. Carbon
elastic moduli of tissues ranging from 50–100 Pa for neural, nanotube research has demonstrated that free nano-dimen-
muscle, and fat cells to 2–4 GPa for articular cartilage and sional carbon-based particles can interfere with the metabo-
osteo tissues (see Figure 7). The stiffness of graphene is lism and membrane integrity of cells, leading to potential toxic
also variable, albeit on a lesser scale, and is dependent on a effects.[46] In addition, if there is degradation of the material,
number of factors, including the layer number and the extent or any mechanism by which nanomaterials can enter the blood
of oxidation, with the elastic moduli of GO less than half stream, the effects of the particles on tissues, such as lungs,
that of pristine graphene.[29,40] Graphene/polymer and gra- liver, and kidneys, and the ability of the body to excrete the
phene/ceramic composites are an ideal route to production materials needs to be considered.
Figure 7. Variation in elastic moduli of tissues within the human body. Reproduced with permission.[41] Copyright 2009, Nature Publishing Group.
www.advmat.de
REVIEW
Table 3. Summary of scientific literature describing pristine graphene, GO, and rGO materials for biocompatibility and tissue engineering.
In vivo/In vitro Graphene type Biological model tested Comments References

In vivo GO i.v. mouse Toxic above 25 mg kg−1, granuloma formation [47]
−1
i.v. mouse >10 mg kg significant toxic effects, lung effects [48]
i.v. mouse Size and dose of GO affected distribution in tissue. Smaller [40]
GO was less toxic.
i.p. mouse 20 mg kg−1 no adverse effects at 7 days [49]
In vitro Pristine Graphene Primary mouse hippocampal cells Graphene slightly improved cell growth over polystyrene [50]
Primary rat cortical neurons No effect on neural cell growth or survival [51]
Human neuroblastoma cell line SH-SY5Y No effect on neural cell growth or survival [44]
Rat PC-12 cells Graphene didn’t affect the morphology of differentiated neu- [42]
rons, electrical stimulation enhanced neurite outgrowth
Primary retinal ganglion neurons Graphene coating increased cell survival compared to [52]
uncoated glass
Mouse hippocampal neural stem cells Cells more proliferative on graphene “foam”, cells could be [53]
electrically stimulated
In vitro GO Human hepatoma cells GO less toxic than CNTs [54]
Mouse C2C12 myoblast cell line Cells more “muscular” on GO-coated glass [55]
Monkey kidney epithelial cells (Vero) GO sheets caused ROS generation and apoptosis at [56]
100 µg mL−1. Functionalized sheets didn't cause toxicity.
Human MG-63 osteosarcoma Cells grew well on and into GO hydrogel [57]
Nil GO hydrogels dehydrated to aerogels promising for scaffolds [58]
In vitro rGO Mouse C2C12 myoblast cell line Cells better on rGO than glass, GO better than rGO [55]
Rat hippocampal neurons rGO showed same cell viability as glass [59]
Nil rGO hydrogel promising for tissue engineering [60]
In addition to doing no harm to the tissues of the body, 4.1. Graphene-Only Structures
graphene must support the desired function of the tissue-
engineering implant. This may involve maturation of undif- 4.1.1. Pristine Graphene
ferentiated cells into a functioning structure, or proliferation
of cells in vivo to populate a synthetic scaffold, with differ- “Pristine” graphene sheets (CVD grown, unfunctionalized gra-
entiation and proliferation often requiring opposing envi- phene) possess remarkable properties – notably phenomenal
ronmental niches. The required function may be to promote theoretical mechanical properties and ballistic conductivity.
migration of cells from the implanted scaffold or material, or However, pristine, single-layer graphene alone is unlikely to
may be to promote invasion of the engineered structure with prove viable for tissue engineering, due to the restrictions in
endogenous cells, or encouragement of angiogenesis (the large-scale fabrication, and difficulties in the processing of the
development of new blood vessels to supply nutrients and material, even on a lab scale. This being said, several research
remove waste from developing tissues). The function of the groups have been able to transfer pristine graphene onto a
structure will vary between applications, and the properties variety of surfaces[42,44,50–52] (summarized in Table 3), allowing
of the material to be used for the tissue engineered structure their properties as surface coatings to be explored. Commonly
must be tailored accordingly. A simple example of this lies in used cell-culture surfaces, such as tissue-culture polystyrene,[50]
the mechanical properties of the structure – the mechanical Permanox,[51] and glass,[44] in addition to silicon/silicon dioxide
properties of the implanted material should approximately surfaces[44] and biodegradable polymers commonly used for
match the mechanical properties of the tissue in which they tissue engineering[42] have been coated with CVD-grown
will be implanted, with bone-engineering materials needing graphene.
to be hard, while neural-engineering structures need to be Li et al. found that for primary murine hippocampal cells,
soft, flexible, and gel-like. The ability to tune the properties a layer of pristine graphene on polystyrene did not change the
of the graphene family of materials make them excellent cell morphology, and that graphene increased GAP-43 expres-
candidates for a wide range of tissue targets, either in gra- sion as well as neurite number and length after 2–7 days in
phene/GO/rGO-only applications (Section 4.1. and Table 3), culture.[50] Sahni et al. cultured rat hippocampal cells on Per-
as composites with inorganic materials (Section 4.2.) or as manox substrates coated in graphene and found the viability,
composites with organic materials – polymers and hydrogels toxicity, morphology, and metabolism of cells grown on the
(Section 4.3.) graphene were unaffected.[51] Using neural differentiation
www.advmat.de
REVIEW
model cell lines, both Park et al. (using human neuroblastoma aminated glass surfaces with both GO and rGO (reduced in
SH-SY5Y cells[44]) and Sherrell et al. (using rat PC-12 cells[42] situ) and grew a muscle cell line, C2C12 cells, on the surfaces
also found no change in viability or morphology on silicon/ to look at myoblast growth and differentiation into multinu-
silicon dioxide or biodegradable poly(lactic-co-glycolic acid) cleated myotubes.[55] They demonstrated that the muscle cells
(PLGA), respectively. Bendali et al. found a slight positive developed into longer myotubes on GO surfaces, and these cells
change when growing primary adult and neonatal retinal gan- expressed more genes typical of a myogenic phenotype. This
glion neurons on glass/sapphire coated with graphene with no is likely to be due to the increased surface roughness of GO,
additional peptide or glial layers to support neural growth, con- as well as the increased absorption of serum proteins on the
cluding that the graphene coating promoted neural survival.[52] oxygen- and hydroxyl-functionalized GO surfaces. These find-
Overall, pristine graphene was demonstrated to have a neutral ings suggest that further investigations into the use of GO for
to slightly positive effect on the survival and metabolism of skeletal tissue engineering is justified; however, both surface
neural cells, with no adverse effects on neural cell culture pub- roughness and increased serum protein binding are unlikely
lished to date. to be important drivers of cell compatibility in a 3D structure.
More importantly, as will be discussed in Section 5, these Studies on neural targets have also been performed, with Kim
studies have all explored the use of graphene-coated substrates et al. finding that rGO modification of glass surfaces did not
for electrode applications, where graphene shows good promise affect hippocampal cell growth over 30 days,[59] a finding with
as a conducting, biocompatible material for providing electrical important implications for modifying or creating more-efficient
signals to, or recording electrical signals from, neural cells. It neural recording or stimulating electrodes.
has been reported that pristine graphene retains sufficient con- While studies on soluble or surface-confined GO or rGO
ductivity after transfer to a flexible substrate to electrically stim- are important for assessing cell and tissue responses, these
ulate the differentiation of neural cells, demonstrating that this do not represent 3D structures for tissue engineering. Several
promise is not unfounded,[42] despite the difficulties in making papers have produced 3D structures from GO or rGO mate-
large-scale pristine graphene coatings. rials, which would be suitable for tissue-engineering purposes.
Hydrogels have been produced by self-assembly of GO under
high temperature for long periods, resulting in gels with over
4.1.2. GO and rGO Structures 97% water.[63] Lim et al. used these materials as a substrate for
growth of an osteosarcoma cell line, demonstrating that the
GO and rGO structures, due to the possibility of large-scale cells adhered to the surface of the hydrogels, and that the cells
synthesis, improved processability, and chemical function- did proliferate, albeit at a lower rate than cells on a tissue-cul-
ality, hold more promise than the single or few-sheet graphene ture-optimized substrate, and with a decrease in cell metabolic
materials above for developing robust implantable structures. state after several days in culture.[57] Another GO hydrogel has
The development of GO and rGO has resulted in an increased been produced by Sui et al., using vitamin C to reduce the GO
ability to explore the compatibility and effects of graphene into a hydrogel, rather than the thermal reduction mentioned
materials with various cells and tissues as coatings and disper- above;[60] however, the biocompatibility of these materials was
sions[27,33,61,62] and even as 3D structures created out of these untested. Another graphene structure that could be used for
materials alone.[63] tissue engineering is the graphene aerogels described by Hu
There have been several in vivo studies investigating the et al. formed by lyophilizing GO hydrogels and exposing them
effects of exposing mice to soluble graphene intravenously, or by to subsequent microwave irradiation.[58] The mechanical prop-
inhalation, intraperitoneal injection, or ingestion[31,40,64,65,47,48] erties of the resulting 3D scaffolds make them very interesting
(summarized in Table 3). A number of groups have observed for tissue-engineering applications where a block of com-
insignificant or no toxicity of GO on animals.[47,65] How- pressible material is required (such as skeletal muscle or fat
ever, others have found negative effects at higher doses, tissues). Finally, the graphene “foam” recently described by Li
(e.g., 10 mg kg−1) including accumulation of GO in the lungs, et al. as a platform for neural-stem-cell differentiation has
cell apoptosis, inflammatory responses, and adverse effects on great potential for a neural stimulation or recording elec-
the liver, spleen, and kidney, resulting in sickness or death of trode.[53] Their CVD-synthesized multilayer graphene foam
the animals.[47,66] As well as the amount of GO or rGO the ani- showed sufficient electrical conductivity to electrically stimu-
mals are exposed to, the state of aggregation of GO in the blood late a differentiated nerve cell growing on its surface, and the
has been found to be the major determining factor on how GO 3D structure had sufficient mechanical strength to be a free-
will accumulate in or be cleared by the body.[40] However, other standing 3D scaffold.
research has pointed out that impurities in GO dispersions are
likely to cause inconsistency in studies at animal levels and cell
levels, with a “purified” GO showing no in vitro or in vivo tox- 4.2. Graphene/Inorganic Composites
icity.[49] There have also been many studies that have looked at
exposure of cultured cells to soluble GO and rGO in vitro, and Hybrid materials, formed from traditional biocompatible
for reviews dealing with in vitro toxicity studies of graphene organic and inorganic materials with graphene materials as a
in greater detail: we recommend Zhang et al.[67] or Sanchez filler, have shown enhanced properties in tissue-engineering
et al.[27] as starting points. systems. For example, hydroxyapatite (HA), a naturally occur-
The modification of surfaces with GO and rGO has been ring mineral form of calcium apatite, makes up approximately
performed for several tissue targets. Ku and Park modified 50% of the weight of bone. Unsurprisingly, as it has excellent
www.advmat.de
the haemolytic activity of GO to almost zero.[79] More impor-

REVIEW
osteoconductive and osteoinductive capabilities, it is com-

monly used as a filler in bone replacement, in osteo-tissue tantly, biocompatible polymers have been used to improve
engineering,[68] or more frequently as a coating on implants to direct graphene–cell interactions by the reduction of surface-
promote osseointegration.[69] However, brittleness and fracture active oxygen species.[56] Reactive oxygen species cause oxida-
toughness render the material inappropriate for load-bearing tive stress in cells, inducing cell toxicity, and reducing these
applications.[70,71] Reduced graphene oxide has been shown to active species can help control apoptosis, leading to improved
be an ideal filler for these applications, as it enhances the frac- cell uptake.[80] Polymers such as poly(ethylene glycol), dextran,
ture toughness of hydroxyapatite without compromising the and poloxamer copolymers have been shown to reduce these
biocompatibility.[70,72,73] These organic/inorganic hybrids can be hydrophobic interactions of graphene with cells,[31,62,78,81]
made by simply mixing rGO and HA solutions at the required and composites made from these materials could be easily
concentrations,[70] by in situ biomimetic mineralization of cleared from the body in mouse in vivo studies. Furthermore,
calcium in the presence of GO to form GO–Ca-bound vaterite biopolymer functionalization of graphene can also have the
microspheres and subsequent reduction to rGO[72] or reduction effect of increasing the effectiveness of graphene implants
of GO without any harsh reducing chemicals by the oxidative by, for example, increasing the osteoinduction of implants for
polymerization of dopamine and subsequent biomimetic min- bone-tissue therapy by coating graphene oxide with fibrin.[82]
eralization in simulated body fluid.[73] HA–rGO composites Synthetic and naturally occurring polymers, such as poly-
produced by mixing HA and rGO prior to spark plasma sin- caprolactone (PCl),[83,84] poly(lactic acid)/poly(glycolic acid)
tering showed higher fracture toughness than pure HA, and, (PLA, PLGA),[85,86] chitosan,[85,87] collagen,[83,88] and agarose,[89]
at 1% (w/w) loading of rGO, greater proliferation and osteo- have frequently been used as substrate materials for tissue-
blast differentiation were observed.[70] Samples produced by engineering scaffolds due to their excellent biocompatibility
in situ mineralization were found to have a high in vitro bone and processability.[90] However, tissue engineering requires a
activity, and GO/rGO–CaCO3 materials showed good HA for- wide range of physical properties that these polymers do not
mation under physiological conditions whilst supporting osteo- necessarily possess. Composite materials using the polymer as
blast attachment.[72] Use of polydopamine to reduce GO to the matrix and carbon-based fillers have the potential to tailor
rGO in situ led to increased mineralization of composites over the properties of these substrates to match those required for
14 days, and materials were found to exhibit no cytotoxicity up the repair of multiple tissue types. Ideal filler materials for the
to 20 µg mL−1.[73] production of these composites would enhance the mechanical
In order to improve the processability and elasticity while and/or electrical properties while retaining the biocompatibility,
retaining their strength and osseointegration potential, gra- processability, and biodegradability of the original polymer.
phene/hydroxyapatite materials have recently been combined Graphene, especially graphene oxide, with its flexibility, poten-
with polymeric matrices. Bone cement formed from HA tial for chemical functionalization, biocompatibility, and
and PMMA with small amounts of GO showed far superior mechanical strength,[91] is a very promising filler material for
mechanical properties due to the high surface area and chem- these composite substrates.
ical functionalization of GO, which led to good interaction Tissue-engineering scaffold performance relies heavily on
between the elements.[74] Poly(lactic acid)/GO/HA composites the mechanical properties of the substrate material. Shape
also showed excellent hardness, elasticity, and load transfer retention and the structural integrity are dependent on the
with GO loadings of 0.1–1% (w/w),[75] and could be electro- tensile strength, while cell proliferation and gene expression
spun into fibers with much greater surface area for improved are affected by the elasticity of the underlying material.[92]
cell adhesion and proliferation.[76] This processability widens The mechanical properties can be altered by changing the
the field of potential applications for graphene–inorganic com- polymer crosslinking density, or using other polymer addi-
posite materials greatly. tives[93] or more easily by addition of fillers such as graphene.
In addition to HA composites, titanium dioxide (TiO2) has There is ample evidence that only very low graphene concen-
been used for bone and dental tissue engineering, and shows trations (<0.05%) are required to induce very large changes
some promise due to the lower rate of corrosion under physi- in the mechanical strength of synthetic biopolymers[38,94]
ological conditions. The compatibility of TiO2/rGO composites without affecting other mechanical properties affecting cell
was tested with several bacterial and human cell lines, and no growth, meaning that cell proliferation and differentiation are
toxic effects were seen for any graphene loading used, with preserved or even enhanced in the hybrid materials in most
more graphene in the composite leading to increased osteo- cases.
calcin expression (taken as an indicator of bone formation) in Hydrogels (water-swellable crosslinked polymers) have been
human osteoblast cells over 21 days.[77] used extensively for tissue-engineering applications. Due to
their high water content, biocompatibility, and adaptable struc-
tures, hydrogels resemble natural soft tissue more than other
4.3. Graphene/Polymer composites biomaterials.[95] However, single-component hydrogels some-
times lack the structural strength and elasticity required as
Despite many studies demonstrating the cytocompatibility scaffold materials. Chitosan, a commonly used hydrogel due to
of graphene and graphene oxide,[47,54,78] the compatibility its versatile chemistry and structural similarity to extracellular
can be improved by surface-chemistry functionalization of matrix materials,[27,96] shows greatly improved mechanical prop-
the nanosheets. Using naturally occurring polymers like chi- erties when blended with graphene oxide.[97] Furthermore, the
tosan to bind to GO nanosheets has been shown to reduce addition of GO or rGO, either as a blend or covalently linked
www.advmat.de
REVIEW
via the amine groups of the polymer, resulted in enhanced con- tissue-engineering scaffold development for real-world appli-
trol over swelling, degradation, shape retention, and cellular cations, in particular in nerve guidance or linear growth of
adhesion.[27,37,98,99] Similarly, using GO as a filler in hydrogel skeletal muscle. Conducting graphene/polymer composites
composites with poly(vinyl alcohol) (PVA),[100] Fmoc (N-flu- that retain the processability of the polymer are ideal mate-
orenyl-9-methoxycarbonyl),[101] and poly(methyl methacrylate) rials for these studies, as they can be processed into highly
(PMMA)[102] showed promisingly large improvements in struc- structured geometries (such as those shown in Figure 2)
tural strength and elasticity. along which cell-growth can be controlled. In addition, the
Due to the load-bearing conditions required for allogenous directionality of cell growth and proliferation has been shown
bone-grafts, reinforcement of biocompatible polymeric mate- to be influenced by stimulation along patterned conducting
rials for osteointegration is even more critical. Carbon nano- networks, including those made with graphene, as shown in
tubes have previously been shown to improve the mechanical Figure 8.[16,52] Reduced graphene oxide/polyester compos-
strength and stiffness of biodegradable polymers to allow ites have been shown to retain the thermal properties of the
sufficient load transfer,[103] but lack the functional groups polymer,[38] allowing composites with enhanced properties to
to form good interfaces with the polymer matrix. Due to the be processed into detailed three-dimensional scaffolds using
two-dimensional nature and surface chemistry of GO, com- additive fabrication techniques. Model neural cells then pref-
posites of graphene oxide in a polymer matrix (in this case erentially adhere to and differentiate along the extruded fibers.
polypropylene fumarate) exhibited noticeably higher stiffness This ability to easily control and modify the mechanical and
and mechanical strength than similar materials produced conducting properties of scaffolds processed from GO and
with multi- and single-wall nanotubes.[104] Following further rGO composites make these materials very promising for
processing using electrospinning, graphene oxide–polymer- tissue-engineering applications.
nanofiber composites had good mechanical-property enhance-
ment at low graphene levels and excellent mouse osteoblast
cell attachment and proliferation.[105] Similar materials were 5. Graphene for Bionics
also shown to modulate osteoblast functions and cell-substrate
interactions.[106] However, there is a limit to graphene loading, To date, all clinically approved electrode materials for medical
as lower concentrations of graphene (<0.5%) in the polymer bionics are metallic. Materials such as platinum or titanium,
matrix yield more appropriate tensile strengths and frac- and alloys containing them, provide a wide, stable electrochem-
ture toughness for osseointegration and, importantly, greatly ical window, wherein unwanted electrochemical reactions can
improved survival of osteoblasts.[107] be avoided.
The dispersion of chemically reduced graphene oxide is a It is recognized, however, that such electrodes do not meet
very useful recent development for composite development.[28] all of the desirable criteria for current and envisaged med-
It re-introduces conductivity to GO which, while less con- ical bionic implants. The advent of organic conductors and
ducting than pristine graphene, has the advantage of residual the building of a knowledge base associated with them has
chemical functionalities, which can be used to form dispersions sparked the interest of biomedical engineers in more recent
and composites. Recently, dispersions of rGO have been used times.[109] For example, organic conducting polymers can be
to form graphene/polymer blends and composites with expo- rendered highly bioactive and the mechanical properties can
nentially increased strength and conductivity using common be tuned over a wide range, and conformable, even stretch-
biocompatible polymers. For example, chemically converted able formulations are available.[22,110] However, these poly-
rGO has been blended with chitosan, a linear polysaccharide, to mers may lack the stability needed for some applications. The
produce swellable hydrogels with conductivities much greater electrochemical stability, potential for high conductivity, and
than the pristine polymer in both the wet and dry states.[37,97,99] tuneable mechanical properties of graphene have been used
Similar composites have also recently been produced without to create a variety of electrodes for stimulation and recording
the need for extra reduction steps or harsh, toxic reducing in biological systems. While the field is still in its infancy (and
agents by the in situ reduction of graphene oxide in biocom- the long-term chemical stability of pristine graphene and rGO
patible polymer matrices using either biocompatible reducing need to be proven for long-term stimulation of tissues under
agents such as ascorbic acid or bovine serum albumin,[53] physiological conditions), there have been several reports of
simultaneous reduction/polymerization,[108] or simply the electrical communication between graphene materials and
reducing characteristics of the solubilizing polymer at mildly biological systems.
elevated temperatures.[70,99,108] While the efficiencies of reduc-
tion of these more biologically compatible processes have not
yet achieved the high degree of reduction achieved with the 5.1. Stimulation
harsher reduction methods, this field is progressing rapidly,
and holds great promise for producing high-conductivity, low- Stimulation of both muscle and nerves has been achieved with
defect, and processable rGO materials. Using these methods, graphene-based materials. Ahadian et al. grew a myoblast cell
well-dispersed nanocomposites are formed in the absence of line (C2C12 cells) on glass slides coated with either GO or
the possibly harmful chemical residues associated with reduced thermally reduced GO, finding that cells preferred to grow on
graphene oxide. reduced graphene oxide, with gene-expression studies con-
Controlling the placement and direction of growth of cells firming increased differentiation on the rGO substrate.[15] They
along three-dimensional structures is an important facet of allowed myoblast differentiation for 8 days before applying
www.advmat.de
REVIEW
Figure 8. Preferential growth of neonatal retinal ganglion cells on graphene tracks on sapphire/graphene substrates (sapphire looks lighter and 10 µm
graphene tracks darker in differential interference contrast images) (a,b,c). Fluorescence (d,e) and SEM images (f) show that while cell bodies may
settle on either sapphire or graphene, neurites prefer to grow on and along on the CVD-synthesized pristine graphene. Reproduced with permission.[52]
Copright 2013, Wiley-VCH.
electrical stimulation for 2 days (8 V at 10 Hz and 10 ms), and fluxes) evoked by electrical stimulation via graphene. Park et al.
found there to be a significant effect on the extent of myoblast demonstrated that human neural stem cells grown directly on
differentiation. pristine graphene transferred onto glass grew successfully with
Most neural studies to date have used cell lines to investigate enhanced neuronal differentiation, and further that a calcium
the effect of electrical stimulation via graphene materials. Sev- flux could be initiated within the cultured cells with electrical
eral publications by Heo and co-workers have considered the stimulation.[45] Li et al. described the differentiation of neural
cell–cell coupling and mobility under electric fields established stem cells in 3D graphene foams (CVD-synthesized graphene
with transparent few-layer-graphene-coated tissue-culture sub- grown on nickel foams, with subsequent removal of the metal
strates.[43,111] The initial study found that low non-contact elec- to leave a freestanding graphene structure).[53] Cells growing
tric fields (down to 4.5 mV mm−1 with charge-balanced biphasic on the surfaces demonstrated increased proliferation in 3D
10 s trains of 0.25 s pulses at 1 Hz) generated by graphene– compared to 2D, with upregulation of Ki67, and were pushed
poly(ethylene terephthalate) for relatively low stimulation toward a more-astrocytic and neuronal lineage. The researchers
periods (as low as 32 min) led to increased cell–cell coupling demonstrated the ability of graphene to act as a conduit to
in SHSY5Y neural cells, with some protein-expression studies directly stimulate (monophasic cathodic pulses of 0.5–1 µA) a
demonstrating increases in cytoskeletal protein expression with calcium flux in individual differentiated cells. In further work
stimulation.[111] Subsequent studies showed increased mobility by the same group, NSCs were differentiated on 2D graphene
in cells cultured alongside transparent, graphene electrodes into a network, and the networks were further stimulated with
at a specific electric field. Cells exposed to an electric field graphene (Figure 9). They found that more neuronal cells
of 45 mV mm−1 for 4 h were significantly more mobile than were developed on the graphene surface than on tissue-culture
unstimulated cells.[43] The same group found that SHSY5Y cell plastic, and that more spontaneous post-synaptic action poten-
layers in which “wounds” were made healed faster with 2 h non- tials were generated with cells grown on graphene.[113] A study
contact electric stimulation provided by graphene electrodes, by Zhang et al. also showed that a cell-line model of neural dif-
with significant effects still present in neural cells treated with ferentiation (PC-12 cells) could be stimulated to undergo cal-
F-actin polymerization inhibitor cytochalasin D, suggesting that cium fluxes with charge injection by rGO rather than pristine
electrical stimulation provided with graphene may have applica- graphene (functionalized with methoxy poly(ethylene glycol)),
tion in controlling neural cells recovering from injury.[112] with the functionalized, amphiphilic rGO leading to better
Live imaging of calcium fluxes within cells has been used by charge-injection properties, and correspondingly higher cal-
several groups to study action potentials (or transient calcium cium fluxes/action potentials in the cultured cells.[114]
www.advmat.de
REVIEW
Figure 9. Comparison of stimulation of calcium fluxes in neural stem cells evoked by electrical stimulation through pristine graphene (A,B) and potas-
sium (C,D), demonstrating the ability of graphene to act as a stimulating electrode to evoke an action potential in cultured neural cells. (A) and (C) show
the Fluo-4-stained cells and the fluorescence intensity profile over distance before and after induction of cells, and (B) and (D) show the difference in
fluorescence intensity over time. Reproduced with permission.[113] Copyright 2013, Elsevier.
In addition to these acute neural stimulation studies, a flexible SU-8 photoresist probe that had been coated with
enhancement of nerve growth or differentiation through more- CVD-synthesized graphene.[115] They found that the introduc-
chronic, long-term stimulation via graphene has been demon- tion of hydroxyl functionalities on pristine graphene by steam
strated. In a multi-parametric study completed by Liu et al., plasma achieved after fabrication of the probe was needed to
NGF release from functionalized rGO-containing structures decrease the hydrophobicity of the probe (from 91.1° ± 5.6° to
combined with stimulation provided by indium tin oxide (ITO) 41° ± 5°) This, is turn, reduced the graphene–bio interfa-
subtrates enhanced the neurite length in PC-12 cells (a cell cial impedance (decreasing from 7216 to 5242 Ω mm−2 and
line that provides a model of neural differentiation). Using the increased the capacitance from 0.7 to 1.4 µF cm−2), providing
same neural-differentiation model, Sherrell et al. showed that an increased fidelity of the recording (Figure 11). Using the
chronic, high-frequency electrical stimulation (250 Hz, biphasic hydrophilic graphene, they were able to record and separate sig-
1 mA cm−2 pulses of 100 µs duration for 8 h per day) supplied nals with signal-to-noise ratio of 27.8 ± 4.0 dB.
to PC-12 cells that were growing directly on uncoated, CVD-syn- Other groups have used graphene in field-effect transistor
thesized bilayer graphene transferred onto flexible PLGA films (FET) setups as bioelectronics interfaces for measurement of
led to a significant increase in the neurite length (Figure 10).[42] action potentials from cultured cardiomyocytes. Cohen-Karni
This, combined with their finding that the cells responded et al. reported an average signal-to-noise ratio of 4 with single-
more strongly to the characteristics of the underlying polymers layer exfoliated graphene-FETs that demonstrated both p- and
than the graphene coating, deomstrated the potential for gra- n- type recording around the Dirac point of the material.[116]
phene to be used for chronic stimulation in electrically driven Hess et al. developed the work further by reporting record-
nerve-repair applications. ings from cultured cardiomyocyte-like HL-1 cells grown on
arrays of graphene-FETs, allowing the tracking of contractions
across a 2D surface, and could average five spikes to increase
5.2. Recording the signal-to-noise ratio to 70.[117] This excellent electronic per-
formance and the electrochemical stability of graphene led
Electrical recordings of cultured or primary-derived tissues the authors to conclude that use of graphene in such arrays
have been achieved using graphene as an electrode material. could lead to large advances in the field of bioelectronics, with
Chen et al. recorded signals from both crayfish axon action graphene-solution gated FETs outperforming current state-of-
potentials and electrocardiograms of zebrafish hearts using the-art devices.
www.advmat.de
REVIEW
Figure 10. Chronic electrical stimulation with graphene leads to enhanced differentiation of a neural model of differentiation. PC-12 cells grown on a
pristine graphene film with electrical stimulation (b) demonstrate longer neurites (c,d) than those grown without electrical stimulation (a). Reproduced
with permission.[42] Copyright 2014, Wiley-VCH.
6. Future of Graphene for Tissue Engineering and of graphene platforms to control the development of neural
Bionics stem cells toward astrocytic and neuronal lineage has been
demonstrated. The degree of reduction of graphene oxide has
Graphene has burst onto the world stage, and is heralded as been used to control the attachment and differentiation of
a star performer by many in materials science. Of particular myoblasts, with the latter promoted using electrical stimula-
relevance here, those in search of an increased biomaterials tion. The usefulness of graphene as a biomaterial capable of
inventory have been captivated by the performance reported to acting as a conduit for electrical stimulation has been clearly
date. The evaluation of this performance in the literature can demonstrated. Some studies have shown that graphene can be
be difficult because of the inconsistent nomenclature (herein cleared from in vivo experiments (although clearance has been
we use the terms “pristine graphene”, “GO”, and “rGO”) used shown to be dependent on the use of a smaller sheet size).
to describe the wide array of “graphene” structures evalu- Others have shown enzymatic degradation of graphene in
ated. However, graphene (in its myriad forms) does provide vivo.[118]
the ability to provide a direct interface to living cells. These Now, much work must be done if practical devices are to be
graphene structures can be created using chemical vapor realized. The myriad graphene chemistries available should
deposition to build graphene from the bottom up, or through allow manipulation that ensures processability and materials
chemical processing from graphite to GO or rGO, to provide amenable to fabrication to create diverse structures. Exquisite
formulations that can be sprayed or printed and used to create control over the processing of graphene to produce a product
practically useful structures. Inevitably the chemistry of the gra- with well-defined composition and structure is required. This
phene structure must be such that it is compatible with other will require the on-going development of characterization tools
materials, most notably for the applications of interest here, to probe the graphene structure in dispersions as well as the
biopolymers. impact of variations on the rheological properties. While the
It has been demonstrated that graphene composition can use of chemical vapor deposition can provide exquisite control
be tuned to facilitate cell adhesion and proliferation. Encour- over graphene formation, this approach is limited in the form
aging results have been shown in studies with osteoblasts, in of devices that can be created.
terms of the use of graphene to adjust the mechanical proper- Chemical processing provides greater flexibility, enabling
ties of the scaffold and neuronal cells, where graphene con- the formation of stable, neat graphene dispersions amenable
ductivity is used to facilitate electrical stimulation. The use to fabrication via 2D or 3D printing or fiber spinning. Such
www.advmat.de
REVIEW
or organic in nature, we will help realize the development of
novel routes to fabrication, including 3D printing and fiber
spinning.
Given the performance characteristics relevant to use of
graphene in bionics that have been demonstrated to date, the
challenges confronting all of us to ensure effective and efficient
implementation are worthy of consideration and need to be
addressed.
Acknowledgements
The authors wish to acknowledge the financial support of the Australian
Research Council through the Australia Laureate Fellowship (for G.W.
and B.T.), and the Superscience Fellowship scheme (for E.M.). B.T. and
E.M. contributed equally to this work.
Received: January 26, 2015

Revised: March 4, 2015
Published online: April 27, 2015
[1] L. E. Freed, F. Guilak, in Principles of Tissue Engineering (3rd ed.),

(Eds: R. Lanza, R. Langer, J. Vacanti), Academic Press, Oxford, UK
2007, p. 137.
[2] USFDA Center for Devices and Radiological Health, 1995 Memo-
randum #G95–1 “Use of International Standard ISO-10993, ‘Bio-
logical Evaluation of Medical Devices Part 1: Evaluation and Testing’”,
US Food and Drug Administration, Rockville, MD, USA 1995.
[3] International Standard ISO10993 “Biological Evaluation of Medical
Devices”, International Standards Organisation 1997.
[4] USFDA Center for Devices and Radiological Health, Use of Interna-
Figure 11. Comparison of electrocardiographic recordings with silver tional Standard ISO-10993, “Biological Evaluation of Medical Devices
wire, an untreated graphene probe, and a “hydrophilic” graphene Part 1: Evaluation and Testing” - Draft Guidance for Industry and
probe. (A) shows the experimental setup, wherein a graphene probe is Food and Drug Administration Staff, US Food and Drug Administra-
located near the ventricle of a zebrafish heart in vitro, and (B) shows the tion, Silver Springs, MD, USA 2013.
increased fidelity of recording obtained with the hydrophilic graphene, [5] USFDA website, http://www.fda.gov/, accessed: March 2015 .
with all waves typical of an electrocardiogram (PQRST waves) observable. [6] Z. M. Summers, J. A. Gralnick, D. R. Bond, mBio 2013,
Reproduced with permission.[115] Copyright 2013, Elsevier. 4, 1.
[7] R. B. Borgens, L. F. Jaffe, M. J. Cohen, Proc. Natl. Acad. Sci. USA
1980, 77, 1209.
dispersions can also be readily formulated into composite [8] C. D. McCaig, A. M. Rajnicek, B. Song, M. Zhao, Physiol. Rev. 2005,
structures with other biomaterials to provide an appropriate 85, 943.
mechanical environment and the addition of bioactivity. How- [9] a) E. Beaumont, E. Guevara, S. Dubeau, F. Lesage, M. Nagai,
M. Popovic, J. Spinal Cord Med. 2014, 37, 93; b) Y.-J. Chang,
ever, accurate control of many parameters in numerous chem-
C.-M. Hsu, C.-H. Lin, M. S.-C. Lu, L. Chen, Biochim. Biophys.
ical steps is needed to provide a reliable material. In addition,
Acta, Gen. Subj. 2013, 1830, 4130; c) J. Huang, Y. Zhang, L. Lu,
while much progress has been made in a short span of time X. Hu, Z. Luo, Eur. J. Neurosci. 2013, 38, 3691; d) A. N. Koppes,
in the development of biocompatible reduction methods for the A. L. Nordberg, G. M. Paolillo, N. M. Goodsell, H. A. Darwish,
conversion of GO to rGO, the efficiency of the conversion using L. Zhang, D. M. Thompson, Tissue Eng., Part A 2014, 20, 494;
cell-compatible techniques his yet to match the effectiveness of e) Y. Liu, R. M. Grumbles, C. K. Thomas, J. Neurophysiol. 2014,
the harsher conventional reduction methods. 112, 660; f) B. Luo, J. Huang, L. Lu, X. Hu, Z. Luo, M. Li, J. Neu-
Liquid-crystalline dispersions of graphene oxide, using gra- rosci. Res. 2014, 92, 893; g) G. Zhang, X. Huo, A. Wang, C. Wu,
phene sheets with a sufficient lateral dimension such that C. Zhang, J. Bai, Neural Regener. Res. 2013, 8, 2531.
they form ordered liquid-crystalline forms under the right [10] a) T. Goroszeniuk, D. Pang, Curr. Pain Headache Rep. 2014, 18,
conditions, have characteristics that present other opportuni- 12; b) M. Knikou, Plos One 2014, 9, e102313; c) J. J. Lemaire,
A. Sontheimer, H. Nezzar, B. Pontier, J. Luaute, B. Roche,
ties. These dispersions can be wet-spun into long lengths of
T. Gillart, J. Gabrillargues, S. Rosenberg, C. Sarret, F. Feschet,
micrometer-sized fiber, or printed using ink-jet, reel-to-reel, or F. Vassal, D. Fontaine, J. Coste, Ann. Fr. Anesth. Reanim. 2014,
even 3D printing techniques. The fabricated structures can then 33, 88; d) H. Matsuo, K. Uchida, H. Nakajima, A. R. Guerrero,
be reduced by chemical means to introduce graphene char- S. Watanabe, N. Takeura, D. Sugita, S. Shimada, T. Nakatsuka,
acteristics with enhanced electronic conductivity. As we con- H. Baba, Pain 2014, 155, 1888; e)C. Xu, Y. Kou, P. Zhang, N. Han,
tinue to refine graphene composites that are either inorganic X. Yin, J. Deng, B. Chen, B. Jiang, Plos One 2014, 9, e105045.
www.advmat.de
REVIEW
[11] M. Malone, D. Gary, I. H. Yang, A. Miglioretti, T. Houdayer, [32] H. Yue, W. Wei, Z. G. Yue, B. Wang, N. N. Luo, Y. J. Gao, D. Ma,
N. Thakor, J. McDonald, Glia 2013, 61, 843. G. H. Ma, Z. G. Su, Biomaterials 2012, 33, 4013.
[12] a) V. Aas, S. Torbla, M. H. Andersen, J. Jensen, A. C. Rustan, in Lipids [33] H. Zhang, C. Peng, J. Z. Yang, M. Lv, R. Liu, D. N. He, C. H. Fan,
and Insulin Resistance: The Role of Fatty Acid Metabolism and Fuel Q. Huang, ACS Appl. Mater. Interfaces 2013, 5, 1761.
Partitioning, Annals of the New York Academy of Sciences, Vol. 967 [34] Q. X. Mu, G. M. Su, L. W. Li, B. O. Gilbertson, L. H. Yu, Q. Zhang,
(Eds: I. Klimes, E. Sebokova, B. V. Howard, E. Ravussin), New Y. P. Sun, B. Yan, ACS Appl. Mater. Interfaces 2012, 4, 2259.
York Academy of Sciences, New York 2002, p. 506; b) S. Ahadian, [35] G. P. Kotchey, B. L. Allen, H. Vedala, N. Yanamala, A. A. Kapralov,
J. Ramon-Azcon, S. Ostrovidov, G. Camci-Unal, V. Hosseini, Y. Y. Tyurina, J. Klein-Seetharaman, V. E. Kagan, A. Star, ACS Nano
H. Kaji, K. Ino, H. Shiku, A. Khademhosseini, T. Matsue, Lab 2011, 5, 2098.
Chip 2012, 12, 3491; c) S. Bayol, C. Brownson, P. T. Loughna, Cell [36] a) E. Murray, S. Sayyar, B. C. Thompson, R. Gorkin, D. L. Officer,
Biochem. Funct. 2005, 23, 361; d) A. Khodabukus, K. Baar, Tissue G. G. Wallace, unpublished; b)S. Sayyar, R. Cornock, E. Murray,
Eng., Part C 2012, 18, 349; e) M. H. M. Thelen, W. S. Simonides, S. Beirne, D. L. Officer, G. G. Wallace, Mater. Sci. Forum 2014 ,
C. vanHardeveld, Biochem. J. 1997, 321, 845. 773–774, 496 .
[13] a) D. A. Lake, Sports Med. 1992, 13, 320; b) A. Delitto, S. J. Rose, [37] S. Sayyar, E. Murray, B. C. Thompson, J. Chung, D. L. Officer,
J. M. McKowen, R. C. Lehman, J. A. Thomas, R. A. Shively, Phys. G. Spinks, G. G. Wallace, J. Mater. Chem. B 2015, 3, 481.
Ther. 1988, 68, 660. [38] S. Sayyar, E. Murray, B. C. Thompson, S. Gambhir, D. L. Officer,
[14] a) B. J. Kirschbaum, H. B. Kucher, A. Termin, A. M. Kelly, D. Pette, G. G. Wallace, Carbon 2013, 52, 296.
J. Biol. Chem. 1990, 265, 13974; b) R. L. Lieber, P. D. Silva, [39] I. Levental, P. C. Georges, P. A. Janmey, Soft Matter 2007, 3, 299.
D. M. Daniel, J. Orthop. Res. 1996, 14, 131; c) H. Fujita, T. Nedachi, [40] J.-H. Liu, S.-T. Yang, H. Wang, Y. Chang, A. Cao, Y. Liu, Nanomedi-
M. Kanzaki, Exp. Cell Res. 2007, 313, 1853. cine 2012, 7, 1801.
[15] S. Ahadian, J. Ramon-Azcon, H. Chang, X. Liang, H. Kaji, [41] D. T. Butcher, T. Alliston, V. M. Weaver, Nat. Rev. Cancer 2009, 9,
H. Shiku, K. Nakajima, M. Ramalingam, H. Wu, T. Matsue, 108.
A. Khademhosseini, RSC Adv. 2014, 4, 9534. [42] P. C. Sherrell, B. C. Thompson, J. K. Wassei, A. A. Gelmi,
[16] A. F. Quigley, J. M. Razal, M. Kita, R. Jalili, A. Gelmi, A. Penington, M. J. Higgins, R. B. Kaner, G. G. Wallace, Adv. Funct. Mater. 2014,
R. Ovalle-Robles, R. H. Baughman, G. M. Clark, G. G. Wallace, 24, 769.
R. M. I. Kapsa, Adv. Healthcare Mater. 2012, 1, 801. [43] C. Heo, J. Yoo, S. Y. Lee, S. Lee, E. Y. Joo, S. B. Hong, Y. H. Lee,
[17] a) D. M. Ciombor, R. K. Aaron, Foot Ankle Clinics 2005, 10, 579; M. Suh, J. Nanosci. Nanotechnol. 2012, 12, 5222.
b) C. P. Huang, X. M. Chen, Z. Q. Chen, Med. Hypotheses 2008, [44] H. B. Park, H. G. Nam, H. G. Oh, J. H. Kim, C. M. Kim, K. S. Song,
70, 287; c) M. Griffin, A. Bayat, Eplasty 2011, 11, e34; d) J. Z. Hu, K. H. Jhee, J. Microbiol. Biotechnol. 2013, 23, 274.
J. Qu, D. Q. Xu, T. Zhang, L. Qin, H. B. Lu, J. Orthop. Res. 2014, [45] S. Y. Park, J. Park, S. H. Sim, M. G. Sung, K. S. Kim, B. H. Hong,
32, 204. S. Hong, Adv. Mater. 2011, 23, H263.
[18] No More Fillings as Dentists Reveal New Tooth Decay Treat- [46] a) B. P. Barna, M. A. Judson, M. J. Thomassen, Nanomaterials
ment (16/6/14), in The Guardian, http://www.theguardian.com/ 2014, 4, 508; b) H. Ali-Boucetta, K. Kostarelos, Adv. Drug Delivery
society/2014/jun/16/fillings-dentists-tooth-decay-treatment ; Rev. 2013, 65, 2111; c) S. Lanone, P. Andujar, A. Kermanizadeh,
accessed: July 2014. J. Boczkowski, Adv. Drug Delivery Rev. 2013, 65, 2063; d) K. Yang,
[19] a) C. D. McCaig, B. Song, A. M. Rajnicek, J. Cell Sci. 2009, 122, Y. Li, X. Tan, R. Peng, Z. Liu, Small 2013, 9, 1492.
4267; b) B. Reid, B. Song, C. D. McCaig, M. Zhao, FASEB J. 2005, [47] K. Wang, J. Ruan, H. Song, J. Zhang, Y. Wo, S. Guo, D. Cui,
19, 379. Nanoscale Res. Lett. 2011, 6, 8.
[20] M. Quarta, M. Scorzeto, M. Canato, M. Dal Maschio, D. Conte, [48] X. Y. Zhang, J. L. Yin, C. Peng, W. Q. Hu, Z. Y. Zhu, W. X. Li,
B. Blaauw, S. Vassanelli, C. Reggiani, Biomaterials 2011, 32, C. H. Fan, Q. Huang, Carbon 2011, 49, 986.
4228. [49] H. Ali-Boucetta, D. Bitounis, R. Raveendran-Nair, A. Servant, J. Van
[21] M. Bustamante, R. Fernandez-Verdejo, E. Jaimovich, S. Buvinic, den Bossche, K. Kostarelos, Adv. Healthcare Mater. 2013, 2, 433.
Am. J. Physiol.: Endocrinol. Metab. 2014, 306, E869. [50] N. Li, X. M. Zhang, Q. Song, R. G. Su, Q. Zhang, T. Kong,
[22] S. M. Richardson-Burns, J. L. Hendricks, B. Foster, L. K. Povlich, L. W. Liu, G. Jin, M. L. Tang, G. S. Cheng, Biomaterials 2011, 32,
D.-H. Kim, D. C. Martin, Biomaterials 2007, 28, 1539. 9374.
[23] D. Mawad, E. Stewart, D. L. Officer, T. Romeo, P. Wagner, [51] D. Sahni, A. Jea, J. A. Mata, D. C. Marcano, A. Sivaganesan,
K. Wagner, G. G. Wallace, Adv. Funct. Mater. 2012, 22, 2692. J. M. Berlin, C. E. Tatsui, Z. Z. Sun, T. G. Luerssen, S. Y. Meng,
[24] J. P. Donoghue, Nat. Neurosci. 2002, 5, 1085. T. A. Kent, J. M. Tour, J. Neurosurg.: Pediatr. 2013, 11, 575.
[25] G. G. Wallace, S. E. Moulton, G. M. Clark, Science 2009, 324, 185. [52] A. Bendali, L. H. Hess, M. Seifert, V. Forster, Anne-Fleur Stephan,
[26] P. Fattahi, G. Yang, G. Kim, M. R. Abidian, Adv. Mater. 2014, 26, J. A. Garrido, S. Picaud, Adv. Healthcare Mater. 2013, 2, 929.
1846. [53] N. Li, Q. Zhang, S. Gao, Q. Song, R. Huang, L. Wang, L. W. Liu,
[27] V. C. Sanchez, A. Jachak, R. H. Hurt, A. B. Kane, Chem. Res. Toxicol. J. W. Dai, M. L. Tang, G. S. Cheng, Sci. Rep. 2013, 3, 1604.
2012, 25, 15. [54] J. Yuan, H. Gao, J. Sui, H. Duan, W. N. Chen, C. B. Ching, Toxicol.
[28] a) S. Gambhir, E. Murray, S. Sayyar, G. G. Wallace, D. L. Officer, Sci. 2012, 126, 149.
Carbon, 2014, 76, 368; b) D. Li, M. B. Muller, S. Gilje, R. B. Kaner, [55] S. H. Ku, C. B. Park, Biomaterials 2013, 34, 2017.
G. G. Wallace, Nat. Nanotechnol. 2008, 3, 101. [56] A. Sasidharan, L. S. Panchakarla, P. Chandran, D. Menon, S. Nair,
[29] C. Bussy, H. Ali-Boucetta, K. Kostarelos, Acc. Chem. Res. 2013, 46, C. N. R. Rao, M. Koyakutty, Nanoscale 2011, 3, 2461.
692. [57] H. N. Lim, N. M. Huang, S. S. Lim, I. Harrison, C. H. Chia, Int. J.
[30] a) O. Akhavan, E. Ghaderi, H. Emamy, F. Akhavan, Carbon 2013, Nanomed. 2011, 6, 1817.
54, 419; b) Y. Li, H. Yuan, A. von dem Bussche, M. Creighton, [58] H. Hu, Z. B. Zhao, W. B. Wan, Y. Gogotsi, J. S. Qiu, Adv. Mater.
R. H. Hurt, A. B. Kane, H. Gao, Proc. Natl. Acad. Sci. USA 2013, 2013, 25, 2219.
110, 12295. [59] S. M. Kim, P. Joo, G. Ahn, I. H. Cho, D. H. Kim, W. K. Song,
[31] K. Yang, J. M. Wan, S. A. Zhang, Y. J. Zhang, S. T. Lee, Z. A. Liu, B. S. Kim, M. H. Yoon, J. Biomed. Nanotechnol. 2013, 9, 403.
ACS Nano 2011, 5, 516. [60] Z. Y. Sui, X. T. Zhang, Y. Lei, Y. J. Luo, Carbon 2011, 49, 4314.
www.advmat.de
REVIEW
[61] a) M. F. Zhang, M. Yudasaka, Yakugaku Zasshi 2013, 133, 151; [89] a) V. Carriel, J. Garrido-Gomez, P. Hernandez-Cortes, I. Garzon,
b) X. Y. Zhang, W. B. Hu, J. Li, L. Tao, Y. Wei, Toxicology Res. 2012, S. Garcia-Garcia, J. A. Saez-Moreno, M. D. C. Sanchez-Quevedo,
1, 62. A. Campos, M. Alaminos, J. Neural Eng. 2013, 10, 026022;
[62] S. Zhang, K. Yang, L. Feng, Z. Liu, Carbon 2011, 49, 4040. b) W. Zhao, X. Jin, Y. Cong, Y. Y. Liu, J. Fu, J. Chem. Technol. Bio-
[63] Y. X. Xu, K. X. Sheng, C. Li, G. Q. Shi, ACS Nano 2010, 4, 4324. technol. 2013, 88, 327.
[64] G. Y. Ni, Y. J. Wang, X. L. Wu, X. F. Wang, S. Chen, X. S. Liu, [90] A. Gloria, R. De Santis, L. Ambrosio, J. Appl. Biomater. Biomech.
Immunol. Lett. 2012, 148, 126. 2010, 8, 57.
[65] K. Yang, H. Gong, X. Z. Shi, J. M. Wan, Y. J. Zhang, Z. Liu, Bioma- [91] a) I. W. Frank, D. M. Tanenbaum, A. M. Van der Zande,
terials 2013, 34, 2787. P. L. McEuen, J. Vac. Sci. Technol. B 2007, 25, 2558; b) C. Lee,
[66] W. Miao, G. Shim, S. Lee, S. Lee, Y. S. Choe, Y.-K. Oh, Biomaterials X. Wei, J. W. Kysar, J. Hone, Science 2008, 321, 385.
2013, 34, 3402. [92] a) A. J. Engler, S. Sen, H. L. Sweeney, D. E. Discher, Cell 2006, 126,
[67] Y. Zhang, T. R. Nayak, H. Hong, W. B. Cai, Nanoscale 2012, 4, 677; b) D. E. Ingber, FASEB J. 2006, 20, 811.
3833. [93] K. Van de Velde, P. Kiekens, Polym. Test. 2002, 21, 433.
[68] a) H. Li, K. A. Khor, V. Chow, P. Cheang, J. Biomed. Mater. Res., [94] a) L. Hua, W. Kai, Y. Inoue, J. Appl. Polym. Sci. 2007, 106, 1880;
Part A 2007, 82A, 296; b) L. L. Hench, J. M. Polak, Science 2002, b) W. Kai, Y. Hirota, L. Hua, Y. Inoue, J. Appl. Polym. Sci. 2008,
295, 1014; c) J. T. K. Melton, R. Mayahi, S. E. Baxter, M. Facek, 107, 1395; c) C. Wan, B. Chen, Biomed. Mater. 2011, 6, 055010;
C. Glezos, J. Bone Jt. Surg., Br. Vol. 2012, 94B, 1067. d) O. J. Yoon, C. Y. Jung, I. Y. Sohn, H. J. Kim, B. Hong, M. S. Jhon,
[69] T.-G. Eom, G.-R. Jeon, C.-M. Jeong, Y.-K. Kim, S.-G. Kim, I.-H. Cho, N.-E. Lee, Composites, Part A 2011, 42, 1978; e) O. J. Yoon,
Y.-S. Cho, J.-S. Oh, Oral Surg., Oral Med., Oral Pathol., Oral Radiol. I. Y. Sohn, D. J. Kim, N.-E. Lee, Macromol. Res. 2012, 20, 789.
2012, 114, 411. [95] J. M. Zhu, R. E. Marchant, Expert Rev. Med. Devices 2011, 8, 607.
[70] Y. Liu, J. Huang, H. Li, J. Mater. Chem. B 2013, 1, 1826. [96] A. Di Martino, M. Sittinger, M. V. Risbud, Biomaterials 2005, 26,
[71] M. Wu, Q. Wang, X. Liu, H. Liu, Carbon 2013, 51, 335. 5983.
[72] S. Kim, S. H. Ku, S. Y. Lim, J. H. Kim, C. B. Park, Adv. Mater. 2011, [97] H. L. Fan, L. L. Wang, K. K. Zhao, N. Li, Z. J. Shi, Z. G. Ge,
23, 2009. Z. X. Jin, Biomacromolecules 2010, 11, 2345.
[73] H. Y. Liu, P. X. Xi, G. Q. Xie, Y. J. Shi, F. P. Hou, L. Huang, [98] D. Depan, R. D. K. Misra, Acta Biomater. 2013, 9, 6084.
F. J. Chen, Z. Z. Zeng, C. W. Shao, J. Wang, J. Phys. Chem. C 2012, [99] X. L. Wang, H. Bai, Z. Y. Yao, A. R. Liu, G. Q. Shi, J. Mater. Chem.
116, 3334. 2010, 20, 9032.
[74] G. Goncalves, S. M. A. Cruz, A. Ramalho, J. Gracio, [100] a) J. Liang, Y. Huang, L. Zhang, Y. Wang, Y. Ma, T. Guo, Y. Chen,
P. A. A. P. Marques, Nanoscale 2012, 4, 2937. Adv. Funct. Mater. 2009, 19, 2297; b) L. Zhang, Z. Wang,
[75] P. A. A. P. Marques, G. Goncalves, M. K. Singh, J. Gracio, C. Xu, Y. Li, J. Gao, W. Wang, Y. Liu, J. Mater. Chem. 2011, 21,
J. Nanosci. Nanotechnol. 2012, 12, 6686. 10399.
[76] H. B. Ma, W. X. Su, Z. X. Tai, D. F. Sun, X. B. Yan, B. Liu, Q. J. Xue, [101] B. Adhikari, A. Banerjee, Soft Matter 2011, 7, 9259.
Chin. Sci. Bull. 2012, 57, 3051. [102] T. Ramanathan, A. A. Abdala, S. Stankovich, D. A. Dikin,
[77] K. Kandiah, P. Muthusamy, S. Mohan, R. Venkatachalam, Mater. M. Herrera-Alonso, R. D. Piner, D. H. Adamson, H. C. Schniepp,
Sci. Eng. C 2014, 38, 252. X. Chen, R. S. Ruoff, S. T. Nguyen, I. A. Aksay, R. K. Prud’homme,
[78] M. Wojtoniszak, X. Chen, R. J. Kalenczuk, A. Wajda, J. Lapczuk, L. C. Brinson, Nat. Nanotechnol. 2008, 3, 327.
M. Kurzewski, M. Drozdzik, P. K. Chu, E. Borowiak-Palen, Colloids [103] a) P. M. Ajayan, J. M. Tour, Nature 2007, 447, 1066; b) X. Shi,
Surf.: B 2012, 89, 79. J. L. Hudson, P. P. Spicer, J. M. Tour, R. Krishnamoorti,
[79] K.-H. Liao, Y.-S. Lin, C. W. Macosko, C. L. Haynes, ACS Appl. A. G. Mikos, Biomacromolecules 2006, 7, 2237; X. Shi,
Mater. Interfaces 2011, 3, 2607. B. Sitharaman, Q. P. Pham, F. Liang, K. Wu, W. E. Billups,
[80] V. Stone, H. Johnston, R. P. F. Schins, Crit. Rev. Toxicol. 2009, 39, L. J. Wilson, A. G. Mikos, Biomaterials 2007, 28, 4078;
613. c) B. Sitharaman, X. Shi, L. A. Tran, P. P. Spicer, I. Rusakova,
[81] A. Sahu, W. I. Choi, G. Tae, Chem. Commun. 2012, 48, 5820. L. J. Wilson, A. G. Mikos, J. Biomater. Sci., Polym. Ed. 2007, 18, 655;
[82] R. Deepachitra, M. Chamundeeswari, B. S. Kumar, G. Krithiga, d) B. Sitharaman, X. Shi, X. F. Walboomers, H. Liao, V. Cuijpers,
P. Prabu, M. P. Devi, T. P. Sastry, Carbon 2013, 56, 64. L. J. Wilson, A. G. Mikos, J. A. Jansen, Bone 2008, 43,
[83] A. G. A. Coombes, E. Verderio, B. Shaw, X. Li, M. Griffin, 362.
S. Downes, Biomaterials 2002, 23, 2113. [104] G. Lalwani, A. M. Henslee, B. Farshid, L. Lin, F. K. Kasper,
[84] a) C. Mota, D. Puppi, D. Dinucci, M. Gazzarri, F. Chiellini, Y.-X. Qin, A. G. Mikos, B. Sitharaman, Biomacromolecules 2013, 14,
J. Bioact. Compat. Polym. 2013, 28, 320; b) Z. Y. Wang, E. Y. Teo, 900.
M. S. K. Chong, Q. Y. Zhang, J. Lim, Z. Y. Zhang, M. H. Hong, [105] Y. Y. Qi, Z. X. Tai, D. F. Sun, J. T. Chen, H. B. Ma, X. B. Yan, B. Liu,
E. S. Thian, J. K. Y. Chan, S. H. Teoh, Tissue Eng., Part C 2013, 19, 538. Q. J. Xue, J. Appl. Polym. Sci. 2013, 127, 1885.
[85] S. J. Kim, D. H. Yang, H. J. Chun, G. T. Chae, J. W. Jang, Y. B. Shim, [106] B. Girase, J. S. Shah, R. D. K. Misra, Adv. Eng. Mater. 2012, 14,
Macromol. Res. 2013, 21, 931. B101.
[86] a) R. P. F. Lanao, A. M. Jonker, J. G. C. Wolke, J. A. Jansen, [107] D. Lahiri, R. Dua, C. Zhang, I. de Socarraz-Novoa, A. Bhat,
J. C. M. Van Hest, S. C. G. Leeuwenburgh, Tissue Eng., Part B 2013, S. Ramaswamy, A. Agarwal, ACS Appl. Mater. Interfaces 2012, 4,
19, 380; b)C. Cicek, E. Cakmakci, N. Kayaman-Apohan, M. Arslan, 2234.
S. E. Kuruca, Int. J. Polym. Mater. Polym. Biomater. 2013, 62, 719. [108] T. D. Dao, K. M. Oh, J. T. Choi, H.-i. Lee, H. M. Jeong,
[87] a) R. H. Mendonca, T. D. Meiga, M. F. da Costa, R. Thire, J. Appl. Y. S. Kim, S.-J. Park, B. K. Kim, J. Nanosci. Nanotechnol. 2012, 12,
Polym. Sci. 2013, 129, 614; b) J. A. Sowjanya, J. Singh, T. Mohita, 8420.
S. Sarvanan, A. Moorthi, N. Srinivasan, N. Selvamurugan, Colloids [109] G. G. Wallace, S. E. Moulton, R. M. I. Kapsa, M. J. Higgins, Organic
Surf.: B 2013, 109, 294. Bionics, Wiley-VCH, Weinheim, Germany 2012.
[88] S. C. Rodrigues, C. L. Salgado, A. Sahu, M. P. Garcia, [110] a) L. Ouyang, C. L. Shaw, C.-C. Kuo, A. L. Griffin, D. C. Martin,
M. H. Fernandes, F. J. Monteiro, J. Biomed. Mater. Res., Part A J. Neural Eng. 2014, 11; b) S. M. Richardson-Burns, J. L. Hendricks,
2013, 101A, 1080. D. C. Martin, J. Neural Eng. 2007, 4, L6; c) D. Khodagholy,
www.advmat.de
REVIEW
T. Doublet, M. Gurfinkel, P. Quilichini, E. Ismailova, P. Leleux, [114] Q. Zhang, J. Xu, Q. Song, N. Li, Z. Zhang, K. Li, Y. Du, L. Wu,
T. Herve, S. Sanaur, C. Bernard, G. G. Malliaras, Adv. Mater. 2011, M. Tang, L. Liu, G. Cheng, J. Liu, J. Mater. Chem. B 2014, 2, 4331.
23, H268. [115] C.-H. Chen, C.-T. Lin, W.-L. Hsu, Y.-C. Chang, S.-R. Yeh, L.-J. Li,
[111] C. Heo, J. Yoo, S. Lee, A. Jo, S. Jung, H. Yoo, Y. H. Lee, M. Suh, D.-J. Yao, Nanomedicine 2013, 9, 600.
Biomaterials 2011, 32, 19. [116] T. Cohen-Karni, Q. Qing, Q. Li, Y. Fang, C. M. Lieber, Nano Lett.
[112] S. Lee, C. Heo, S. Y. Lee, Y. H. Lee, M. Suh, The Enhancement of 2010, 10, 1098.
Neuronal Cells Wound Healing with Non-contact Electric Field [117] L. H. Hess, M. Jansen, V. Maybeck, M. V. Hauf, M. Seifert,
Stimulation by Graphene Electrodes, Proc. SPIE 2013, 8879, DOI: M. Stutzmann, I. D. Sharp, A. Offenhäusser, J. A. Garrido, Adv.
10.1117/12.2020770. Mater. 2011, 23, 5045.
[113] M. Tang, Q. Song, N. Li, Z. Jiang, R. Huang, G. Cheng, Biomate- [118] C. M. Girish, A. Sasidharan, G. S. Gowd, S. Nair, M. Koyakutty,
rials 2013, 34, 6402. Adv. Healthcare Mater. 2013, 2, 1489.

石墨烯生物材料到仿生学

Uploaded by

Document Information

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

石墨烯生物材料到仿生学

Uploaded by

Copyright:

Available Formats

www.advmat.

Graphite Oxide to Graphene. Biomaterials to Bionics

synthetic scaffolds to make tissue engi-

While the concept of biocompatibility is loosely defined

2.1.2. Muscle 2.2.1. Stimulation Mechanisms

Figure 4. Action potentials in neural, skeletal, and cardiac muscles,

recordings) or sealed onto a small area of the membrane of

2.2.3. Requirements for Electrical Stimulation and Recording

cellular uptake and excretion in vivo.[30–32] A systematic study

This review will primarily cover the most recent progress

In vivo/In vitro Graphene type Biological model tested Comments References

the haemolytic activity of GO to almost zero.[79] More impor-

osteoconductive and osteoinductive capabilities, it is com-

Received: January 26, 2015

[1] L. E. Freed, F. Guilak, in Principles of Tissue Engineering (3rd ed.),

You might also like