materials

Review
Nanotechnology Scaffolds for Alveolar
Bone Regeneration
Goker Funda 1, * , Silvio Taschieri 1,2 , Giannì Aldo Bruno 1,3 , Emma Grecchi 3 ,
Savadori Paolo 2 , Donati Girolamo 4 and Massimo Del Fabbro 1,2
1 Department of Biomedical, Surgical and Dental Sciences, University of Milano, 20122 Milan, Italy;
silvio.taschieri@unimi.it (S.T.); aldo.gianni@unimi.it (G.A.B.); massimo.delfabbro@unimi.it (M.D.F.)
2 IRCCS Orthopedic Institute Galeazzi, Via Riccardo Galeazzi, 4, 20161 Milano MI, Italy;
paolo_savadori@yahoo.it
3 Dental and Maxillo-Facial Surgery Unit, IRCCS Ca Granda Ospedale Maggiore Policlinico di Milano,
Via Francesco Sforza 35, 20122 Milan, Italy; emma.grecchi@gmail.com
4 ASST Fatebenefratelli Sacco Hospital, Dentistry Department, Via Giovanni Battista Grassi, 74, 20157 Milan,
Italy; girolamo.donati@asst-fbf-sacco.it
* Correspondence: funda.goker@unimi.it; Tel.: +39-02-5031-9950




Received: 4 November 2019; Accepted: 31 December 2019; Published: 3 January 2020


Abstract: In oral biology, tissue engineering aims at regenerating functional tissues through a series
of key events that occur during alveolar/periodontal tissue formation and growth, by means of
scaffolds that deliver signaling molecules and cells. Due to their excellent physicochemical properties
and biomimetic features, nanomaterials are attractive alternatives offering many advantages for
stimulating cell growth and promoting tissue regeneration through tissue engineering. The main aim
of this article was to review the currently available literature to provide an overview of the different
nano-scale scaffolds as key factors of tissue engineering for alveolar bone regeneration procedures.
In this narrative review, PubMed, Medline, Scopus and Cochrane electronic databases were searched
using key words like “tissue engineering”, “regenerative medicine”, “alveolar bone defects”, “alveolar
bone regeneration”, “nanomaterials”, “scaffolds”, “nanospheres” and “nanofibrous scaffolds”.
No limitation regarding language, publication date and study design was set. Hand-searching
of the reference list of identified articles was also undertaken. The aim of this article was to give a
brief introduction to review the role of different nanoscaffolds for bone regeneration and the main
focus was set to underline their role for alveolar bone regeneration procedures.

Keywords: scaffolds; nanomaterials; tissue engineering; regenerative medicine; alveolar

bone regeneration

1. Introduction
The reconstruction and augmentation of the alveolar bone is a complex and challenging field for the
maxillofacial and periodontal surgeon. The crucial aim of the therapies in this field is mainly increasing
the bone mass in patients who have lost this tissue as a result of a consequence of several conditions
such as periodontal disease, aging, osteoporosis, trauma, neoplastic pathology and reconstructive
surgery or as a result of congenital defects [1].
At present auto-transplantation from a patient’s extra-oral or intra-oral donor site is accepted
as the gold standard and is the most frequently used method [2]. The critical limitations of this
conventional approach are donor site morbidity, inadequate blood supply of graft tissue, associated
pain and limited supply. For these reasons, autologous grafting is usually reserved for a restricted
number of cases [2,3].

Materials 2020, 13, 201; doi:10.3390/ma13010201 www.mdpi.com/journal/materials

Materials 2020, 13, 201 2 of 20

Alternative sources for bone grafts include allografts (grafts originating from another individual
of the same species) and xenografts (grafts originating from different species such as bovine or porcine).
However, these substitutes also have some certain disadvantages such as the possibility of immune
rejection and pathogen transmission from donor to host [2,3].
Another strategy for bone grafts is the use of synthetic alloplasts made from polymers, ceramics
or metals. Alloplasts represent some disadvantages including non-optimal integration with the native
tissue at the site of the defect and the potential failure due to fatigue or infection caused during
implantation. In addition to these, their applications are limited at locations of high stress or mechanical
load [2,3].
Owing to the drawbacks and limitations of bone grafts, over the last few decades, several
novel approaches involving tissue engineering and regenerative medicine (TE/RM) have emerged
as alternatives to conventional treatments. The fundamental concept underlying TE/RM was to use
scaffolds alone or in combination with growth factor, cell and/or gene delivery to form a so-called
“tissue engineering construct,” that stimulates tissue repair and/or regeneration [2–5].
In brief, the four crucial factors that must be considered with TE/RM are:

• Cells, which represent the fundamental structural unit of any tissue,

• A matrix such as “scaffolds”, as framework material supporting the growth of cells to form a fully
organized tissue,
• Biological factors like growth factors (GF) and bone morphogenetic proteins (BMPs) to guide
cellular activity and tissue formation,
• Vascularization to provide oxygen and nutrients for the cell metabolism, and to remove catabolic
waste products [6,7].

Current limitations in bone TE/RM strategies are the impaired cellular differentiation, inadequate
synthesis of extrinsic factors essential for an effective osteogenesis and, most importantly, insufficient
mechanical strength and physicochemical properties of the conventional scaffolds [7]. Conventional
scaffolds combined with growth factors and cells do not always achieve successful bone regeneration
in clinical settings, generally due to the limited capability of controlling framework degradation as
well as delivery of biological factors and drugs [6,8,9].
The focus of this review was to give a brief introduction to the nanoscaffolds in TE/RM and to
underline the role of different scaffolds in successful tissue formation for bone regeneration. The main
aim of this literature-based article is to provide an overview of the different nanoscale scaffolds as key
factors of the tissue-engineering paradigm, used for alveolar bone regeneration.
In order to find articles pertinent to this narrative review, PubMed/Medline, Scopus and Cochrane
electronic databases were searched using key terms such as “tissue engineering”, “regenerative
medicine”, “alveolar bone defects”, “alveolar bone regeneration”, “nanomaterials”, “scaffolds”,
“nanospheres” and “nanofibrous scaffolds”. No limitation regarding study design, publication date
and language was set. A hand search through the references of the identified articles and previous
reviews was also undertaken. The last electronic search was performed on 3 December 2019.

1.1. Importance of Scaffolds in TE/RM

Scaffold material plays a key role in tissue regeneration, as it provides a micro-environment
suitable for cell adhesion, proliferation and differentiation. In general, an ideal scaffold material should
be biocompatible, have a controllable degradation and appropriate physico-chemical features in order
to simulate the extracellular matrix (ECM) structure of the original tissues [7]. It should be able to
balance various combinations of materials with specific functions by means of engineered surface, cell
encapsulation and controlled release of chemicals [6,9]. Additionally, it should be able to endorse and
control peculiar events occurring at the cellular and tissue level [10].
In scaffold based tissue engineering, the scaffold is expected to perform several functions. It should
provide adequate mechanical strength and stiffness to replace the mechanical properties of the missing
Materials 2020, 13, 201 3 of 20

or the injured tissue [6,11]. Most importantly, a successful scaffold should stimulate not only the early
ingrowth of new tissue, but also the progressive maturation and remodeling by providing adequate
support and morphology [12]. Its design should also take into account degradation kinetics and
physico-chemical features [6,11].

1.2. Nanotechnology
One thousand nm is equal to 1 micrometer (µm) and nanoparticles are smaller than 100 nanometers
(nm). Nanomaterials compared with bulk materials possess features like quantum size, macroscopic
quantum tunneling, as well as small size effects, resulting in altered physiochemical properties [7].
Current areas of research in nanotechnology for tissue regeneration are as follows:

i. Nanoparticle-based techniques for delivering bioactive molecules,

ii. Nanoparticle-mediated cells tagging and targeting,
iii. Nanoparticle-based scaffold manufacturing [7].

Additionally, the half-life and distribution of nanoparticles can be affected by their size.
Their surface properties can determine their stability and their localization in the tissues [7]. A decreased
size in nanomaterial particles is beneficial in terms of stiffness, effective surface area and area-to-volume
ratio [7]. The charge of the nanoparticles is also another important factor affecting the internalization
of nanosized particles into different target cells [7].
In oral biology, nanotechnology applications are mainly focused on augmentation procedures
for bone tissue regeneration and enhancement of osseointegration of oral implants [7]. Natural bone
itself is constituted by a highly organized extracellular matrix (ECM) in a nanometric scale, and the
application of nanoscaffolds can represent an intrinsic advantage for tissue engineering regeneration
procedures of the musculoskeletal apparatus [8,13]. Nanomaterials can overcome the main problems
encountered with the current scaffolds used for bone regeneration such as: Inadequate mechanical
strength, instability of growth factors released and impaired cellular differentiation [7,10].

1.3. Rationale of Nanotechnology Scaffolds in Tissue Engineering

Advanced materials play a crucial role in promoting the bench-to-bedside translation of tissue
engineering technologies [14,15]. Direct application of therapeutic substances may be affected by
some limitations of the conventional scaffold materials including non-specific targeting, insufficient
physiological stability and low cell membrane permeability. Usually, supra-physiological doses are
needed to compensate for the poor pharmacokinetics of such agents, which in return increase the
potential risk of adverse effects [7]. Currently, nanotechnology has allowed the production of structures
having the same size as the naturally occurring tissues and has opened a new era for TE/RM [8,16].
Nanoscaffolds can be produced so as to be extremely similar to tissue-specific ECM. The reduced
size of the nanoparticles permits a fast response to external stimuli coming from the environment, like
ultrasounds, magnetic fields, pH and iX-ray exposure.
Nanoscaffold materials may be used to deliver drugs, genetic material or biological factors in a
controlled way, both systemically and locally [8]. Nanoscaffolds can stabilize the bioactive agents by
means of encapsulation or surface attachment, promoting molecule internalization, targeting their
delivery from cells and allowing to control biological factor release at the intended target [7]. Controlled
and sustained delivery by nanoparticles mainly depends on their reduced size and related high specific
surface area [17]. Thus they may represent stimulus-sensitive delivery vehicles for chemically or
biologically active substances, which will provide a triggered delivery as a response to an external
stimulus [7,16,17].
Nanoscaffolds have a considerable drug loading capability, high mobility of drug loaded particles,
and efficient in vivo reactivity toward nearby tissues [17]. They can be used for labeling cells, in order
to enable continuous cell tracking and monitoring [8,16]. Additionally, nanoparticles can provide
enhanced osseointegration, osteoconduction and osteoinduction [16].
Materials 2020, 13, x FOR PEER REVIEW 4 of 20

cells, in2020,
Materials order to enable
13, 201continuous cell tracking and monitoring [8,16]. Additionally, nanoparticles
4 of 20
can provide enhanced osseointegration, osteoconduction and osteoinduction [16].

1.4.
1.4. Nanofibrous
Nanofibrous Scaffold
Scaffold Systems
Systems
Fibrous
Fibrous nanoscaffolds
nanoscaffolds areare extremely-thin
extremely-thin uninterrupted
uninterrupted fibers with a short short diffusional
diffusional path,
path, aa
considerable
considerablesurface
surfacearea to unit
area massmass
to unit ratio and
ratiohigh
andporosity. The porous
high porosity. Thestructure
porousofstructure
the nanofibrous
of the
drug deliverydrug
nanofibrous systems is highly
delivery interconnected
systems and represents an
is highly interconnected and adequate
representssubstrate for cell adhesion
an adequate substrate
and transport
for cell of nutrients.
adhesion One ofofthe
and transport limitations
nutrients. Oneofof
these
the systems is the
limitations ofinitial
these burst release
systems phenomenon,
is the initial burst
which
releasedepends on the which
phenomenon, large surface
depends area
onand the short
the large diffusional
surface area and path.
the short diffusional path.
Fibrous
Fibrousnanoscaffolds,
nanoscaffolds,besides
besidestheir role
their as as
role drug carriers,
drug provide
carriers, mechanical
provide strength.
mechanical There
strength. had
There
been worries
had been that that
worries drugdrug
incorporation
incorporationinto into
the nanofibers
the nanofibers might impair
might the the
impair mechanical
mechanical features of
features
the
of the same nanofibers [8]. As an example for a solution to this problem, Ionescu et al. developed aa
same nanofibers [8]. As an example for a solution to this problem, Ionescu et al. developed
microsphere-laden
microsphere-laden fibrous nanoscaffold structure in which the microspheres were to deliver drugs,
while
while the
the nanofibers
nanofibersonly
onlyworked
workedas asan
anengineered
engineeredscaffold
scaffold[18].
[18].

1.5.
1.5. Nanosphere
Nanosphere Scaffold
Scaffold Systems
Systems
Nanospheres
Nanospheres can
can deliver
deliver drugs,
drugs, growth
growth factors
factors or
or genetic
genetic material
material [7].
[7]. The
The advantages
advantages of
of
nanospheres over conventional monolithic bulk scaffolds can be listed as follows:
nanospheres over conventional monolithic bulk scaffolds can be listed as follows:
•• Mechanically
Mechanically weakweak scaffolds
scaffolds in
in load-bearing
load-bearing applications
applications may be be reinforced
reinforced by by the
the adjunct
adjunct ofof
nanospheres
nanospheresas ascross-linking
cross-linkingagents,
agents,
•• The porosity
The porosityofofthe thematerials
materialsconstituting
constitutingtraditional
traditional scaffolds
scaffolds maymaybe be significantly
significantly enhanced,
enhanced, by
by adding
adding spheres
spheres like like porogen.
porogen. Increased
Increased porosity
porosity means means allowing
allowing interior
interior tissuetissue infiltration
infiltration into
intoscaffold,
the the scaffold,
•• Nanospheres can stimulate
Nanospheres stimulate the
the creation
creation of of apatite
apatite crystals
crystals and
and the
thefollowing
followingmineralization
mineralization of of
hydrogels through
hydrogels through the the release
release of
ofthe
thecorresponding
correspondingminerals,
minerals,so sothat
thatself-hardening
self-hardeningbiomaterials
biomaterials
adapted to
adapted to the
the regeneration
regenerationof ofbone
bonetissue
tissuecan
canbebeproduced.
produced.
•• Injectable and/or
Injectable and/or moldable
moldable materials
materials can can be
be developed
developed thanks
thanks toto the
the spherical
spherical nature
nature of
of some
some
nanomaterials, so that their application is possible by means of minimally invasive
nanomaterials, so that their application is possible by means of minimally invasive surgery [17]. surgery [17].

1.6. Classification
1.6. Classification of
of Nanoparticle
Nanoparticle (NP)
(NP) Materials
Materials
Nanoparticles can
Nanoparticles can be
be classified
classifiedas
asinorganic
inorganic(Figure
(Figure1)
1)and
andorganic
organicnanoparticles
nanoparticles(Figure
(Figure2).
2).

Figure 1.
Figure 1. Inorganic
Inorganic nanomaterials.
nanomaterials.
Materials 2020, 13,
Materials 2020, 13, 201
x FOR PEER REVIEW 55 of 20
20

Figure 2.
Figure 2. Organic
Organic nanomaterials.
nanomaterials.

1.6.1.
1.6.1. Inorganic
Inorganic NPs
NPs
Synthetic Polymers
Synthetic Polymers
Synthetic polymers have some advantages, such as sufficient supply, easy fabrication, easy
Synthetic polymers have some advantages, such as sufficient supply, easy fabrication, easy
adaptation, high safety profile and reasonable costs. Additionally, they have adjustable physiochemical
adaptation, high safety profile and reasonable costs. Additionally, they have adjustable
and morphological features, which are valuable for wide-scale production and application [19,20].
physiochemical and morphological features, which are valuable for wide-scale production and
The most common synthetic polymers for biomedical applications are poly-α-hydroxyesters
application [19,20].
(poly-glycolic acid (PGA), poly-lactic acid (PLA), poly-lactic-glycolic acid (PLGA) and poly-caprolactone
The most common synthetic polymers for biomedical applications are poly-α-hydroxyesters
(PCL)) [20]. However, there are also disadvantages of poly-α-hydroxyesters, which can be listed as
(poly-glycolic acid (PGA), poly-lactic acid (PLA), poly-lactic-glycolic acid (PLGA) and poly-caprolactone
follows: (i) Hydrophobicity, which causes failure of loading hydrophilic drugs or molecules and poor
(PCL)) [20]. However, there are also disadvantages of poly-α-hydroxyesters, which can be listed as
cell adhesion, (ii) degradation by autocatalysis, which causes unpredictable degradation behavior,
follows: (i) Hydrophobicity, which causes failure of loading hydrophilic drugs or molecules and
(iii) an acidic degradation product, which leads to denaturation of bioactive proteins and inflammatory
poor cell adhesion, (ii) degradation by autocatalysis, which causes unpredictable degradation
tissue response, and (iv) low capacity of loading therapeutic agents, which limits their penetration into
behavior, (iii) an acidic degradation product, which leads to denaturation of bioactive proteins and
the polymer network [20,21].
inflammatory tissue response, and (iv) low capacity of loading therapeutic agents, which limits their
Various combinations of PLGA/PLLA/PEG/PCL were tested by several researchers with promising
penetration into the polymer network [20,21].
results for alveolar bone regeneration applications [1,22–24].
Various combinations of PLGA/PLLA/PEG/PCL were tested by several researchers with
•promising results for alveolar bone regeneration applications [1,22–24].
Dendrimers
• Dendrimers are synthetic polymers with branching treelike structures. They are biocompatible
Dendrimers
and biodegradable with uniform morphology. They have a high number of surface functional groups,
Dendrimers are synthetic polymers with branching treelike structures. They are biocompatible
which makes them suitable for several applications [16,25]. Dendrimers can cause an improvement on
and biodegradable with uniform morphology. They have a high number of surface functional
drug solubility [16,26] and are mostly used for targeted drug delivery reasons [16,27]. However, they
groups, which makes them suitable for several applications [16,25]. Dendrimers can cause an
have a few disadvantages, like cytotoxicity and possible poor drug retention inside the branches of the
improvement on drug solubility [16,26] and are mostly used for targeted drug delivery reasons
dendrimer [16].
[16,27]. However, they have a few disadvantages, like cytotoxicity and possible poor drug retention
inside the
Ceramic branches of the dendrimer [16].
NPs
Ceramic
Ceramic NPs materials are “synthethic crystalline, solid, inorganic non-metallic materials” [28].
Bioceramics include bioactive glass, bioactive glass–ceramic, calcium phosphate groups and
Ceramic materials are “synthethic crystalline, solid, inorganic non-metallic materials” [28].
alumina [16,29,30].
Bioceramics include bioactive glass, bioactive glass–ceramic, calcium phosphate groups and
alumina [16,29,30].
Materials 2020, 13, 201 6 of 20

Bioceramics possess certain advantages like biocompatibility, nontoxicity and dimensional

stability [29]. They are added to the scaffolds to improve their structural properties and delivery
performance. Many ceramic materials such as calcium phosphate groups and mineral trioxide
aggregate materials have been used and are currently in use for bone regeneration procedures [16].
Bioactive glass nanoscaffold was investigated and was reported to be beneficial for formation of
new alveolar bone tissue [31].
• Calcium Phosphate (CP) Groups
Normal bone tissue is composed of 30% w/v organic collagen fibers and 70% inorganic matter,
mainly CP crystals. The latter represents a model for mimicking natural bone material at the
macro- and nanoscale level [7]. For this reason, CP nanoparticles (nano-tricalcium phosphate (nTCP)
and nano-hydroxyapatite (nHA)) have been the most commonly applied materials for bone tissue
regeneration [32].
CP has many favorable properties such as similarity to the inorganic portion of natural bone tissue,
biocompatibility, biosafety, osteoconductivity, low cost and ease of manufacturing. The drawbacks
are poor regulation of drug delivery and degradation rate [20,33]. However, when CP is used as a
single component, it might represent some limitations due to its poor mechanical features and very low
toughness. In order to overcome such drawbacks, researchers recently introduced composite scaffolds
composed of nTCP/nHA plus further biological materials or synthetic polymeric materials [33].
Several applications of nanoHA [34–39] and nanoTCP [40,41] combinations were investigated in
the literature and were reported as successful for alveolar bone tissue regeneration.
• Mineral Trioxide Aggregate (MTA)
MTA is prepared by bismuth oxide reaction, and is composed of tricalcium aluminate, tricalcium
silicate, dicalcium silicate and calcium sulfate dehydrate [42,43]. During hydration, MTA releases
Ca2+ and OH ions, which increases pH in order to neutralize the acidic metabolites produced by
macrophages and osteoclasts. Nanoscale particle sized tricalcium aluminate may increase bone
regeneration, inflammatory response and foreign body reactions [43].

Silica NPs
Silica, either itself or as a coating of other compounds, has been used for applications in the
biomedical field, like imaging and drug delivery [16]. According to their application purposes,
silica nanoparticles can be synthesized as bulk particles, core/shell silica NPs and mesoporous silica
nanoparticles (MSNPs) [16]. In particular, MSNPs have attracted the attention of researchers for
applications of controlled release. MSNPs can be synthesized in different ways to obtain particles with
various dimensions and physical properties. MSNPs have unique pore structure, high surface area
and large pore volume. Additionally, because of their honeycomb-like porous structure, they are able
to encapsulate and absorb a number of biomolecules [16,44,45].
The typical advantages of silica nanoparticles are their uniform morphology, biocompatibility and
chemical stability. Their disadvantage is their variable toxicity [16].

Metallic NPs
Metallic NPs are biocompatible materials with reduced cytotoxicity. Their functionalization is
easy; however, they need long term cytotoxicity testing and their coating is advised [16]. In biomedical
applications, gold NPs are considered as the safest and the most commonly utilized among other
metallic NPs. Gold NPs can be synthesized as spheres, rods or cages and they can be applied in areas
such as drug delivery, biosensing, bioimaging and photothermal therapy [16,46,47].
Combinations of gold and silver nanoparticles with chitosan for alveolar bone regeneration were
reported for gold [48] and silver [22] with successful outcomes for enhanced mineralization and
implant osseointegration.
Materials 2020, 13, 201 7 of 20

Magnetic NPs
In biomedical applications, nickel, iron, cobalt and their oxides may be used as magnetic
nanoparticles for drug delivery, three-dimensional cell organization, cell tracking, imaging procedures
and as biosensors [16,49]. Some magnetic nanoparticles offer super paramagnetic properties with
long-lasting and non-invasive tracking possibility. However, magnetic nanoparticles are cytotoxic and
coating is required due to the safety concerns for overcoming their cytotoxicity and for producing
biocompatible magnetic NPs [16].

1.6.2. Organic Nanoparticles

Liposomes
Liposomes are vesicles composed of bilayers of phospholipids [50]. Their properties are affected by
their lipid size, surface charge, composition and preparation method [16]. Liposomes are biodegradable,
stimuli responsive and can be incorporated with hydrophilic/hydrophobic drugs [16,51]. They are
mainly used in drug delivery and imaging [52]. They are the most clinically tested nanosystems,
with several commercialized formulations [51]. However, their rapid clearance from circulation and
scale-up issues might represent some disadvantages [16,51].

Natural Polymeric NPs

Natural polymers are biocompatible and biodegradable materials with bulk physical properties.
They are divided mainly as proteins and polysaccharides [53] and can be listed as collagen, alginate,
fibrin, gelatin and chitosan [17]. Due to their intrinsic biocompatibility and biodegradability, natural
polymers are very important for bone tissue engineering procedures [54]. The synthesis methods
can be flexible and polymer chains can show a wide diversity of composition and properties [53].
Generally they have high drug loading capability, but the scale-up issues might represent some
disadvantages [16,53].
• Collagen
Collagen is the most frequently tested natural polymer since the ECM of the natural bone
is mainly composed of collagen. It has excellent biocompatibility, optimal biodegradability and
negligible immunogenicity [17,55]. The limitations of the collagen can be listed as weak mechanical
stability, possible denaturation on processing and the intrinsic risk of causing immune reactions and/or
transmitting infectious agents. As an alternative solution, instead of collagen extracted from animals,
recombinant collagen was proposed for tissue engineering procedures [17].
• Gelatin
Gelatin is a denatured form of collagen with similar beneficial biological properties and limitations.
Gelatin degradation is controllable by tailoring the crosslinking procedures. The existing presence of
functional groups additionally allow functionalization by chemical transformation [17].
• Chitosan
Chitosan has many advantageous biological and physicochemical properties. It is biocompatibile,
antibacterial and its production is easy. Various studies have tested chitosan as a factor and/or drug
delivery system for pharmaceutical and tissue engineering applications [5,17].
Chitosan nanomaterial was tested by researchers with different combinations of nanoparticles
such as PLGA/bioactive glass [1], PCL/Sr-doped calcium phosphate [56], PLGA/silver [22] and gold [48]
for alveolar bone regeneration procedures with successful results in favor of alveolar bone regeneration.
Materials 2020, 13, 201 8 of 20

• Alginate
Alginates are biocompatible, hydrophilic, non-immunogenic and cost effective materials.
Their intrinsic features can induce in vivo calcification without the use of any additives and they are
suitable for a number of tissue engineering and drug delivery applications [17]. However their rather
slow degradation when implanted in bone defects is poorly controllable and they do not have cell
attachment sites for osteoblast anchorage [17].

Carbon
• Carbon Nanotubes
Carbon nanotubes (CNTs) are carbon allotropes with a long cylindrical structure. They might be
single-walled or multi-walled graphitic hollow tubular structures. They have mechanical and chemical
stability with optimum electrical properties. However, their good chemical stability might represent a
drawback for the covalent functionalization process. Carbon materials are added to the composite
scaffolds, in order to reinforce the structural properties [16].
Carbon nanotubes are effective on bone tissue regeneration as they promote cell proliferation
and osteogenic differentiation [57]. Several carbon nanomaterials like CNTs, graphene oxide (GO),
fullerenes, carbon dots (CDs), nano-diamonds and their derivatives were successfully tested as a
scaffold for bone tissue engineering [57].
• Graphene and Its Derivates
Graphene is a film monolayer one-atom-thick, with a honeycomb-like structure made of carbon
atoms, and arranged in a two-dimensional hexagonal structure [58]. Nanomaterials of the graphene
family include a great variety of graphene derivatives such as, graphene oxide (GO), reduced graphene
oxide (rGO) and graphene nanosheets [59,60]. Graphene and its derivatives represent advantageous
mechanical, electrochemical and physical properties such as, small size, large surface area, thermal
stability, electrical conductivity and mechanical strength. Additionally, graphene materials may be
functionalized and combined with different biomaterials and biomolecules, such as small molecules,
polymers or nanoparticles, by means of covalent or non-covalent interaction [50–61]. Graphene
materials are favorable for cell differentiation, proliferation and osteogenic differentiation. Additionally
they are biocompatible and represent antibacterial properties. However, the behavior of graphene is
dependent on its size, surface functionalization and its coating [59–61]. Biomedical applications of
graphene oxide can produce adverse effects mostly dependent on the dose and time. Currently, the
cytotoxicity of the GO material represents a challenging situation for clinical applications. The quality
of graphene plays a major role since the presence of impurities can cause undesired events [62].
Interactions between body and GO are still being investigated for in vivo applications and it is most
likely that their primary clinical applications shall be the ones for topical use or short-time transient
implantations [63].
GOs are able to enter into cytoplasm and nuclei. They can induce cell membrane damage and
apoptosis. GO can induce lung diseases by causing severe toxicity. GOs also stay for a long time
in kidneys since they are cleaned in the kidney by a very complex process [64]. Further studies are
lacking in the literature to explore their toxicity and effect on cells/tissues.
Graphene oxide combined with silk fibroin [65,66], GO-coating of titanium implants [67] and
GO-coating of collagen membranes [60,61] for guided bone regeneration were investigated by various
researchers. The results showed that these applications can improve cell proliferation and osteogenic
differentiation with success.

1.6.3. Composite Scaffolds

Inspired by the natural organic/inorganic composition of bone, different organic and/or inorganic
particle combinations were tested as composite scaffolds that increase the advantages and decrease
Materials 2020, 13, 201 9 of 20

the drawbacks of each component [17]. In order to overcome the disadvantages of a single particle,
strategies of different composite scaffold designs containing diverse materials have been developed by
many researchers [68].
Combining nHA with polymers of high molecular weight was evaluated to overcome the
disadvantages of a single material. PLA [19], PLGA [69], collagen [70], polyamide [71], coralline [72],
chitosan and PCL [73] have been combined with nHA and were found to improve scaffold
biocompatibility as well as mechanical strength. As an example for bone regeneration, several
studies have shown that the combination of nHA and collagen scaffolds has favorable mechanical
properties [19,70]. Collagen fiber provides an osteoinductive and absorbable scaffold for infiltration
of osteoblast cells. However, the nonabsorbable nHA can reduce this effect. For this reason, the
percentage of nHA and collagen in composite materials play a crucial role for tissue regeneration [19].

1.7. Manufacturing Methods to Fabricate Scaffolds

Strategies of scaffold designs utilizing particles can be listed as follows: (i) Particles as different
single components embedded into matrices, solid polymers, hydrogels or calcium phosphate cements,
(ii) particles combined into blocks for fabrication of scaffolds, (iii) haphazard packing, (iv) rapid
prototyping and self-assembling scaffolds (assembly driven by electrostatic, magnetic interactions or
hydrophobic interactions) [17].
Various methods have been developed to produce scaffolds with porous properties for bone
regeneration, such as freeze-drying [74], gas foaming [75], salt leaching [76], phase transformation [77],
sponge replication [78], electrospinning [79] and additive manufacturing methods such as 3D printing
(selective laser sintering (SLS), stereolithography, laser assisted printing, inkjet printing and extrusion
printing) [32,68,80].
The sol-gel phase, co-precipitation as well as chemo-mechanical techniques have been utilized for
nHA production [81]. However, these procedures are complex with high cost and poor reproducibility.
Additionally, the manufacturing variables, the low-yield end-products and high byproducts are difficult
to control, and the produced nHA is different from the biological HA in terms of physicochemical
properties [81].

1.8. Biphasic and Multiphasic Scaffolds

The nanoparticles have many limitations such as a high agglomeration rate and an uneven
distribution pattern. Biphasic and multiphasic preparations might partially overcome the limitations
of single-phase biomaterials by improving the bioactivity and biocompatibility [32] since they have the
ability to combine different material compositions and physical structures [82–84].
The rationale of utilizing multiphasic/biphasic scaffolds in oral science mostly depends on the
expectations from an ideal scaffold. An ideal scaffold should have various functions, such as the
support for cell colonization, migration, differentiation and growth. The design should also take
into account suitable physical and physicochemical properties together with degradation kinetics.
Multiphasic scaffolds are needed for an appropriate control of the spatiotemporal events for periodontal
tissue regeneration since they can allow compartmentalized tissue healing [82,83]. Manufacturing of
multiphasic nanoscaffolds for periodontal tissue engineering can be divided into three approaches:
(i) Bilayered occlusive membrane + bone compartment [84,85], (ii) compartmentalized biphasic
scaffolds [86–89] and (iii) compartmentalized triphasic scaffold [90]. In vitro and in vivo investigations
have reported promising results for biphasic/multiphasic scaffold designs. However, the methods still
need to be refined for future clinical applications.

1.9. Three-Dimensional (3D) Porous Scaffolds

Three-dimensional porous scaffolds have been developed and tested in many tissues, including
bone, cartilage, skin, muscle and vasculature [17]. The importance of 3D porous scaffolds comes from
their ability to simulate extracellular matrices, which can sustain cellular activity. They can provide a
Materials 2020, 13, 201 10 of 20

pool of bioactive signaling molecules through a sustained release to the natural environment. In this
way, they might regulate cell function and trigger tissue repair [17].

1.10. Three-Dimensional Bio-Printing

Three-dimensional bio-printing (3D additive manufacturing) is a precise layering of biomaterials,
biochemical substances and living cells, with spatial control of the positioning of each component [91].
Tissue bio-printing strategies include inkjet, micro-extrusion and laser-assisted bio-printing.
Three-dimensional printing is one of the hot topics in TE/RM and nanomaterials play a crucial
role in this manufacturing method [92,93]. The major challenge in these technologies is to provide
multiple cell types and micro-architecture of extracellular matrix (ECM) in an adequate resolution to
reproduce biological function [92,93].
In periodontal regeneration, the main challenge is the scaffold design, which might mimic
the periodontal tissue nature and organization. While 3D bio-printing may help to solve this
challenge, the long-term success of these applications mostly depends on the properties of the current
biomaterials [82,94].

1.11. Influence of Magnetic Fields on Biological Systems

Magnetism might play a significant role in the control of cell responses. A magnetic field can be
applied to cells externally or internally; cells can be embedded in a scaffold material with magnetic
properties [95]. Yun et al. investigated the additional effects of the outer static magnetic field (SMF)
with magnetic polycaprolactone nanocomposite scaffold for bone regeneration. According to the
results of this experiment, the combination of external and internal magnetism can be advantageous
for new bone formation [95].

2. Nanoscaffold Applications for alveolar Bone Regeneration

In TE/RM, one of the major research topics is to create bioconstructs that can integrate with the
in vivo tissues [10]. Current systems represent limitations such as restricted diffusion and uneven
cell-matrix distribution. In order to overcome limitations of the current materials, during the last years
different types of scaffolds and bioreactors have been designed and tested [10]. The regeneration of oral
tissues is very challenging [96] and nanomaterials might represent a great potential for future TE/RM
applications of various craniofacial and oral defects. The most frequently investigated nanomaterials for
oral tissues can be listed as nanofibers, nanoparticles, nanosheets, nanotubesand nanospheres. Recently,
the multi-layer nanoscaffolds for oral tissue regeneration were also tested by researchers [97,98].
In general, nanomaterials have excellent physiochemical properties and biomimetic features for
enhancing cell growth and tissue regeneration [96,97]. Nanomaterials, due to their size, can have an
effective control of the release rate of the each agent upon matrix degradation. Furthermore, they
can be used for delivery of therapeutic agents in alveolar bone and tooth regeneration. Nanofibers
represent an ideal environment for cell migration due to their similarity to extracellular matrices
and their high porosity. In addition, nanotubes and nanoparticles might enhance the chemical and
mechanical features of the scaffold, increase cell migration and proliferation, and tissue regeneration [96].
While the application of nanomaterials in TE/RM is still in its infancy phase with several limitations
to be addressed, the recent results of investigations indicate their potential role for future oral tissue
engineering applications [93].
Three fabricating modes exist for TE/RM scaffolds: Growth factor-scaffold, cell-scaffold and cell
growth factor-scaffold. In cell-scaffold, and cell growth factor-scaffold materials, cells are inserted on a
biocompatible scaffold, in order to proliferate into the scaffold and to remodel the natural environment
by secreting ECM. Biomaterials in periodontal tissue engineering are mainly growth factor-scaffold,
used as the carrier of exogenous growth factors. The main challenge is the necessity to provide an
adequate number of seeded cells integrated with the scaffold material in vitro and implanted into a
defect position [96,99].
Materials 2020, 13, 201 11 of 20

There is a huge number of experimental studies reported in the literature, searching for the best
nanoscaffold material for various periodontal tissue regeneration applications. Ogawa et al. [41] tested
nano-ß-TCP/collagen loaded with fibroblast growth factor-2 (FGF-2) on periodontal wound healing.
In the study, nano-ß-TCP scaffold, nano-ß-TCP scaffold loaded with FGF-2 and non-coated collagen
scaffold were implanted into one-wall infrabony defect models. nano-ß-TCP scaffold was fabricated
by surface coating of collagen scaffold with nanosize ß-TCP dispersion. According to the four week
post-surgery histological results, nano-ß-TCP scaffold loaded with FGF-2 displayed nearly five-times
greater periodontal tissue repair when compared with the collagen scaffold [41].
Xue et al. [22] produced nanoparticles of chitosan, PLGA and silver (Ag) for investigating
the optimal combination ratio for mineralization of periodontal membrane cells, and periodontal
tissue regeneration. The antibacterial properties of each nanoparticle were also evaluated. Different
combination ratios of nPLGA and chitosan were tested. According to the results, single nanoparticles
did not show any cytotoxicity and were able to enhance cell mineralization. Additionally, chitosan
and nAg showed antibacterial properties, while nAg limited cell proliferation. The 3:7 ratio of
nPLGA/chitosan and 50 µg/mL nAg was the optimal proportion [22].
Hydrogels are soft materials with (3D) polymer structure, water-absorbability and
adjustable physical and chemical properties [100]. Natural and synthetic hydrogels composed
of micro-/nanostructures have been shown to imitate the chemical and physical properties of
natural ECM for bone and periodontal tissue regeneration [37–101]. Gelatin can be modified by
methacryloyl (methacrylamide and methacrylate) side groups and the final product is GelMA [84].
Gelatin methacrylate (GelMA) is a biocompatible and photocrosslinkable hydrogel. Chen et al. [37]
tested fabrication of GelMA/nanohydroxylapatite microgel arrays using a photocrosslinkable method.
The group evaluated the regeneration and osteogenic proliferation of human periodontal ligament stem
cells (hPDLSCs) encapsulated in microgels. According to the results GelMA/nHA microgels (10%/2%
w/v) enhanced periodontal tissue regeneration. Since GelMA/nHA microgels enhanced hPDLSCs
proliferation and osteogenic differentiation in vitro and supported bone regeneration in vivo [37].
In recent years, researchers proposed and tested graphene-based nanomaterials for oral tissue
engineering because of the many advantageous properties and antibacterial capacities of the
graphene [60,61,102].
Sowmya et al. [1] tested a scaffold system for the regeneration of oral tissues (periodontal ligament,
cementum and alveolar bone) in a rabbit study [1]. The structure of the scaffold consisted of three
layers. For bone tissue a layer of chitosan/PLGA/nano-sized bioactive glass layer enriched with PRP,
for periodontal ligament a layer of chitosan/PLGA with the adjunct of fibroblast growth factor 2
(FGF-2), and finally for cementum a layer of chitosan/PLGA/nano-sized bioactive glass layer loaded
with cementum protein 1 (CEMP1). According to the histological and tomographic evaluation results,
new alveolar bone formation and complete periodontal healing was obtained in three months [1].
Zhang et al. [103] tested the incorporation of growth factors in nanomaterial-based silk fibroin
scaffolds in a dog study for tissue regeneration. Nanomaterial-based silk fibroin scaffolds loaded with
BMP-7 and/or platelet derived growth factor (PDGF)-ß adenovirus were used to test tissue regeneration.
According to the results, the scaffolds loaded with BMP-7 mainly enhanced alveolar bone regeneration,
while the scaffolds incorporated with PDGF-ß adenovirus produced a partial regeneration of the
periodontal ligament. The combination of growth factors created a synergistic effect showing up to
two times greater alveolar bone, periodontal ligament, and cementum formation, as compared with
each factor alone [103].
Vaquette et al. [40] tested PCL and ß–TCP for periodontal ligament and alveolar bone regeneration.
The scaffold design was composed of a flexible electrospun component for the periodontal ligament
section and a fused deposition modeled component for the bone section. Cell sheet technology was
utilized to manufacture biphasic scaffolds additively. While the results were promising with increased
mechanical stability of the cell sheets and mineralization, the bone section did not provide sufficient
ectopic bone proliferation [40].
Materials 2020, 13, 201 12 of 20

Rasperini et al. [104] tested an individualized custom-made 3D-biomanufactured scaffold for

regeneration of a periodontal osseous defect in a clinical study. For this purpose, a customized scaffold
containing PCL powder and HA was produced by using selective laser sintering technology, according
to the computed tomography scan of the patient’s defect. While the scaffold became exposed after
12 months, which led to failure, the results of this study are promising in terms of further research [104].
Nanomaterials seem to be promising for alveolar bone regeneration applications as they mostly
provide favorable results, as listed in Table 1. However, the decreased size of nanomaterials can
cause some potential problems. Nanomaterials possess sizes close to biological molecules, peptides,
deoxyribonucleic acid and viruses. Thus, they can provoke adverse events by moving throughout the
body, depositing in target organs (liver, lungs, kidney, heart and spleen), penetrating cell membranes
and staying in the mitochondria [105–108]. Manganese oxide, titanium dioxide, aluminum oxide, zinc
oxide and silver might accumulate and elicit harmful responses [109]. The brain is partly shielded
by the blood–brain barrier, but the nanoscale size of these materials might cause them to cross this
barrier or they might penetrate through the olfactory and sensory nerves. The biological toxicity of
nanomaterials is thought to be induced by their oxidative properties [110,111] and further research
should be conducted to evaluate and develop a science-based risk evaluation of the nanomaterials [109].

Table 1. Nanoscaffold applications for alveolar bone regeneration.

Material Reference Outcome

3 layer chitosan/PLGA/nano-sized Complete periodontal healing and new
Sowmya et al. [1]
bioactive glass alveolar bone deposition after three months
Favourable on promoting osteoblastic
GO-coating of collagen membranes Radunovic et al. [60]
differentiation process
Improved biocompatibility of collagen
GO-coating of collagen membranes De Marco et al. [61] membranes on in vitro human primary
gingival fibroblast model
Enhanced mechanical stability of the cell
PCL containing ß–TCP Vaquette et al. [40] sheets, and mineralization. However,
ectopic bone ingrowth was not sufficient
nano-ß-TCP/collagen scaffolds loaded with
nano-ß- TCP/collagen scaffolds Ogawa et al. [41] fibroblast growth factor-2 (FGF-2) improved
periodontal tissue wound healing results
Chitosan, PLGA, and silver (Ag) Contributed to cell mineralization without
Xue et al. [22]
nanoparticles complex cytotoxicity
Promoted in vivo osteogenesis of hPDLSCs
GelMA/nHAmicrogels Chen et al. [37]
encapsulated in microgels
nanomaterial-based silk fibroin scaffolds
Zhang et al. [103] Promoted periodontal healing
incorporating BMP-7 and/or PDGF-ß
Clinical study with failure due exposure of
PCLpowder containing HA Rasperini et al. [104]
the scaffold
Favourable on osteogenic differentiation but
Graphene Xie et al. [112]
not on osteoblastic differentiation
Favourable on mechanical resistance and
Graphene Oxide combined with silk
Rodríguez-Lozano et al. [65] hPDLSC proliferation and showed
fibroin
biocompatibility
PDLSC proliferation rate into
Graphene Oxide combined with silk
Vera-Sánchez et al. [66] osteo/cementoblast like cells improved with
fibroin
these combinations
Improved cell proliferation, osteogenic
GO-coating of titanium implants Ren et al. [67] differentiation and biocompatibility of
implants
Citric Acid-Based Nano Hydroxyapatite Dayashankar et al. [39] Significant bone regeneration
Nano-bioactive glass loaded with Good osteoconductivity for promoting the
Zhang et al. [31]
NELL1 gene formation of new alveolar bone tissue
Materials 2020, 13, 201 13 of 20

Table 1. Cont.

Material Reference Outcome

Poly(l-lactic acid) (PLLA) nanofibrous
spongy microspheres, In a mouse model of periodontitis, the
PLLA/polyethylene glycol (PEG) injectable and biomolecule-delivering PLLA
co-functionalized mesoporous silica Liu et al. [23] lead to Treg enrichment, expansion, and
nanoparticles, and poly(lactic Treg-mediated immune therapy against
acid-co-glycolic acid) (PLGA) bone loss
microspheres
Promoted bone formation in critical sized
Nanofibrous yarn reinforced HA-gelatin Manju et al. [34]
alveolar defects in rabbit model
Induced osteogenic differentiation of
Silver nanoparticle-coated collagen
Chen et al. [35] mesenchymal stem cells that guide bone
membrane
regeneration.
Chitosan-gold nanoparticles mediated Enchanced osseointegration of dental
Takanche et al. [48]
gene delivery implant even in osteoporotic condition
Hydroxyapatite nanowires modified Promoted bone regeneration in a rat
Han et al. [36]
polylactic acid membrane mandible defect model
PCL/chitosan/Sr-doped calcium Higher ALP activity level and a better
Ye et al. [56]
phosphate electrospun nanocomposite matrix mineralization
Nano hydroxyapatite mineralized silk
Nie et al. [38] Improved osteogenesis
fibroin
PLGA/PCL Modification Including
Enhanced alveolar bone regeneration
Silver Impregnation, Collagen Coating, Qian et al. [24]
(31.8%)
and Electrospinning

One of the limitations of the nanoscaffold systems is the incorporation of nanoscale material
for reinforcement. In the regeneration of bone, utilizing micro- and nano-length scale materials can
be beneficial for increasing the intrinsic fracture resistance of the bone by affecting bone strength
and supporting plasticity. However, macro-length scale materials are also beneficial, influencing
toughness and creating a resistance to fracture. In brief, nanoscale materials increase the capacity to
withstand pressure, but toughness drops dramatically [6] and it is not realistic to expect both high
strength and high toughness properties from one single material [113]. As an example, in ceramic
based materials, toughness is provided majorly by their large scales. Ceramic based materials are
able to shield crack by supporting uncracked material bridging the crack wake, which is impossible
for micro/nano-scale materials [114,115]. Nanotechnology alone cannot be the optimum solution for
improving the mechanical properties of the scaffolds. Incorporation of different scale materials with a
hierarchical design exhibiting many scales seems to be beneficial and should be considered as a future
perspective [6,116].

3. Conclusions
This review mainly focused on the basic information about the nanomaterials for bone regeneration
to give researchers a general knowledge about their possible applications in the oral field. Currently
their application in oral and maxillofacial clinics is very limited but interest is increasing for their
possible applications in order to overcome the current problems associated with conventional materials.
As a conclusion, it can be underlined that nanomaterials have excellent physico-chemical properties
and biomimetic features for promoting cell growth and stimulating tissue regeneration, and oral tissue
engineering with nanomaterials seems to represent a great potential with vital importance as future
treatment modalities.
However, nanomaterials should not be evaluated as optimum solutions for every current problem.
It should be understood that there are also disadvantages of nanomaterials such as toughness, which
can be solved with the incorporation of a hierarchical design encompassing many length-scales to
generate stronger and tougher scaffold materials. They can also provoke adverse events by moving
all over the body or by depositing in organs due to their nanosize, which is similar to biological
Materials 2020, 13, 201 14 of 20

molecules and viruses. Further research needs to be done in order to provide solutions for possible
future applications.

Author Contributions: Databases were searched and data was collected by G.F., M.D.F., E.G., G.A.B., S.T., S.P.,
D.G. All the authors contributed on analysis and interpretation of data for the work. G.F. drafted the work and
wrote the manuscript with input from all authors. G.F. and M.D.F. revised the work critically for intellectual
content. Integrity of the work was appropriately investigated and resolved by all authors. All authors contributed
and approved equally to the final version of the manuscript. All authors have read and agreed to the published
version of the manuscript.
Funding: This research received no external funding.
Acknowledgments: None.
Conflicts of Interest: All the authors have no conflict of interest to disclose.

Abbreviations
TE/RM Tissue engineering and regenerative medicine
GF Growth factors
BMP Bone morphogenetic proteins
mRNA Micro ribonucleic acid
TCP Tricalcium phosphate
BMMSC Bone marrow mesenchymal stem cells
MSC Mesenchymal stem cells
PRP Platelet rich plasma
ECM Extracellular matrix
nm Nanometers
µm Micrometers
NP Nanoparticles
PLA Poly-lactic acid
PGA Poly-glycolic acid,
PLGA Poly-lactic-co-glycolic acid
PLLA poly(l-lactic acid)
PEG PLLA/polyethylene glycol
PCL Poly-caprolactone
CP Calcium Phosphate
HA Hydroxyapatite
TCP Tricalcium phosphate
nHA Nano hydroxyapatite
nTCP Nano tricalcium phosphate
MTA Mineral trioxide aggregate
MSNP Mesoporous silica nanoparticle
CNT Carbon nanotube
GO Graphene oxide
CD Carbon dots
GO Graphene oxide
rGO Reduced graphene oxide
SLS Selective laser sintering
3D 3 dimensional
SMF Static magnetic field
FGF-2 Fibroblast growth factor-2
Ag Silver
GelMA Gelatin methacrylate
hPDLSC Human periodontal ligament stem cells
CEMP1 Cementum protein 1
PDGF Platelet derived growth factor
DNA Deoxyribonucleic acid
Materials 2020, 13, 201 15 of 20

References
1. Sowmya, S.; Mony, U.; Jayachandran, P.; Reshma, S.; Kumar, R.A.; Arzate, H.; Nair, S.V.; Jayakumar, R.
Tri-Layered Nanocomposite Hydrogel Scaffold for the Concurrent Regeneration of Cementum, Periodontal
Ligament, and Alveolar Bone. Adv. Healthc. Mater. 2017, 6, 1601251. [CrossRef] [PubMed]
2. Pilipchuk, S.P.; Plonka, A.B.; Monje, A.; Tau, A.D.; Lanisdro, A.; Kang, B.; Giannobile, W.V. Tissue Engineering
for Bone Regeneration and Osseointegration in the Oral Cavity. Dent. Mater. 2015, 31, 317–338. [CrossRef]
[PubMed]
3. Mudda, J.A.; Bajaj, M. Stem cell therapy: A challenge to periodontist. Indian J. Dent. Res. 2011, 22, 132–139.
[CrossRef] [PubMed]
4. Rios, H.F.; Lin, Z.; Oh, B.; Park, C.H.; Giannobile, W.V. Cell- and Gene-Based Therapeutic Strategies for
Periodontal Regenerative Medicine. J. Periodontol. 2011, 82, 1223–1237. [CrossRef]
5. Goker, F.; Ersanlı, S.; Arısan, V.; Cevher, E.; Güzel, E.E.; İşsever, H.; Ömer, B.; Altun, G.D.; Morina, D.;
Yılmaz, T.E.; et al. Combined effect of parathyroid hormone and strontiumranelate on bone healing in
ovariectomized rats. Oral Dis. 2018, 24, 1255–1269. [CrossRef]
6. Saiz, E.; Zimmermann, E.A.; Lee, J.S.; Ulrike, G.K.; Wegst, U.G.K.; Tomsia, A.P. Perspectives on the Role of
Nanotechnology in Bone Tissue Engineering. Dent. Mater. 2013, 29, 103–115. [CrossRef]
7. Walmsley, G.G.; Mc Ardle, A.; Tevlin, R.; Momeni, A.; Atashroo, D.; Hu, M.S.; Feroze, A.H.; Wong, V.W.;
Lorenz, P.H.; Longaker, M.T.; et al. Nanotechnology in bone tissue engineering. Nanomedicine 2015,
11, 1253–1263. [CrossRef]
8. Carbone, E.J.; Tao Jiang, T.; Nelson, C.; Henry, N.; Lo, K.W.H. Small molecule delivery through nanofibrous
scaffolds for musculoskeletal regenerative engineering. Nanomedicine 2014, 10, 1681–1699. [CrossRef]
9. Hosseinpour, S.; Ahsaie, M.G.; Rad, M.R.; Baghani, M.; Motamedian, S.R.; Khojasteh, A. Application of
selected scaffolds for bone tissue engineering: A systematic review. Oral Maxillofac. Surg. 2017, 21, 109–129.
[CrossRef]
10. D’Amora, U.; Russo, T.; Gloria, A.; Rivieccio, V.; D’Ant, V.; Negri, G.; Ambrosio, L.; De Santis, R. 3D
additive-manufactured nanocomposite magnetic scaffolds: Effect of the application mode of a time-dependent
magnetic field on hMSCs behavior. Bioact. Mater. 2017, 2, 138–145. [CrossRef]
11. Ferracane, J.L.; Giannobile, W.V. Novel Biomaterials and Technologies for the Dental, Oral, and Craniofacial
Structures. J. Dent. Res. 2014, 93, 1185–1186. [CrossRef] [PubMed]
12. Zhang, Y.; Sun, H.; Song, X.; Gu, X.; Sun, C. Biomaterials for periodontal tissue regeneration. Rev. Adv. Sci.
2015, 40, 209–214.
13. Deng, M.; James, R.; Laurencin, C.T.; Kumbar, S.G. Nanostructured polymeric scaffolds for orthopaedic
regenerative engineering. IEEE Trans. Nanobiosci. 2012, 11, 3–14. [CrossRef] [PubMed]
14. Laurencin, C.T.; Khan, Y. Regenerative engineering. Sci. Transl. Med. 2012, 14, 160–169. [CrossRef]
15. Laurencin, C.T.; Ashe, K.M.; Henry, N.; Kan, H.M.; Lo, K.W.H. Delivery of small molecules for bone
regenerative engineering: Preclinical studies and potential clinical applications. Drug Discov. Today. 2014,
19, 794–800. [CrossRef]
16. Vieira, S.; Vial, S.; Reis, R.L.; Oliveira, J.M. Nanoparticles for Bone Tissue Engineering. Biotechnol. Prog. 2017,
33, 590–611. [CrossRef]
17. Wang, H.; Leeuwenburgh, S.C.G.; Li, Y.; Jansen, J.A. The Use of Micro- and Nanospheres as Functional
Components for Bone Tissue Regeneration. Tissue Eng. Part B 2012, 18, 24–39. [CrossRef]
18. Ionescu, L.C.; Lee, G.C.; Sennett, B.J.; Burdick, J.A.; Mauck, R.L. An anisotropic nanofiber/microsphere
composite with controlled release of biomolecules for fibrous tissue engineering. Biomaterials 2010,
31, 4113–4120. [CrossRef]
19. Wang, X.; Xing, H.; Zhang, G.; Wu, X.; Zou, X.; Feng, L.; Wang, D.; Li, M.; Zhao, J.; Du, J.; et al.
Restoration of a critical mandibular bone defect using human alveolar bone-derived stem cells and porous
nano-HA/Collagen/PLA Scaffold. Stem Cells Int. 2016, 8741641. [CrossRef]
20. Wang, X.; Li, W. Biodegradable mesoporous bioactive glass nanospheres for drug delivery and bone tissue
regeneration. Nanotechnology 2016, 27, 225102. [CrossRef]
21. Mundargi, R.C.; Babu, V.R.; Rangaswamy, V.; Patel, P.; Aminabhavi, T.M. Nano/micro technologies
for delivering macromolecular therapeutics using poly(d,l-lactide-coglycolide) and its derivatives.
J. Control. Release 2008, 125, 193–209. [CrossRef] [PubMed]
Materials 2020, 13, 201 16 of 20

22. Xue, Y.; Hong, X.; Gao, J.; Shen, R.; Ye, Z. Preparation and biological characterization of the mixture of
poly(lactic-co-glycolic acid)/chitosan/Ag nanoparticles for periodontal tissue engineering. Int. J. Nanomed.
2019, 14, 483–498. [CrossRef] [PubMed]
23. Liu, Z.; Chen, X.; Zhang, Z.; Zhang, X.; Saunders, L.; Zhou, Y.; Ma, P.X. Nanofibrous Spongy Microspheres to
Distinctly Release miRNA and Growth Factors To Enrich Regulatory T Cells and Rescue Periodontal Bone
Loss. ACS Nano 2018, 12, 9785–9799. [CrossRef]
24. Qian, Y.; Zhou, X.; Zhang, F.; Diekwisch, T.G.H.; Luan, X.; Yang, J. Triple PLGA/PCL Scaffold Modification
Including Silver Impregnation, Collagen Coating, and Electrospinning Significantly Improve Biocompatibility,
Antimicrobial, and Osteogenic Properties for Orofacial Tissue Regeneration. ACS Appl. Mater. Interfaces
2019, 11, 37381–37396. [CrossRef] [PubMed]
25. Menjoge, A.R.; Kannan, R.M.; Tomalia, D.A. Dendrimer-based drug and imaging conjugates: Design
considerations for nanomedical applications. Drug Discov. Today 2010, 15, 171–185. [CrossRef] [PubMed]
26. Milhem, O.M.; Myles, C.; McKeown, N.B.; Attwood, D.; D’Emanuele, A. Polyamidoamine Starburst
dendrimers as solubility enhancers. Int. J. Pharm. 2000, 197, 239–241. [CrossRef]
27. Gillies, E.R.; Frechet, J.M.J. Dendrimers and dendritic polymers in drug delivery. Drug Discov. Today. 2005,
10, 35–43. [CrossRef]
28. Yamamuro, T. Bioceramics. In Biomechanics and Biomaterials in Orthopedics; Poitout, D.G., Ed.; Springer:
London, UK, 2004; pp. 22–23. [CrossRef]
29. Raghavendra, S.S.; Jadhav, G.R.; Gathani, K.M.; Kotadia, P. Bioceramics in endodontics–A review. J. Istanb.
Univ. Fac. Dent. 2017, 51, 128–137. [CrossRef]
30. Pina, S.; Oliveira, J.M.; Reis, R.L. Natural-based nanocomposites for bone tissue engineering and regenerative
medicine: A review. Adv. Mater. 2015, 27, 1143–1169. [CrossRef]
31. Zhang, J.; Chen, Y.; Xu, J.; Wang, J.; Li, C.; Wang, L. Tissue engineering using 3D printed nano-bioactive glass
loaded with NELL1 gene for repairing alveolar bone defects. Regen. Biomater. 2018, 5, 213–220. [CrossRef]
32. Ebrahimi, M.; Botelho, M.; Lu, W.; Monmaturapoj, N. Synthesis and characterization of biomimetic bioceramic
nanoparticleswith optimized physicochemical properties for bone tissue engineering. J. Biomed. Mater. Res. A
2019, 27. [CrossRef]
33. Liu, L.; Liu, J.; Wang, M.; Min, S.; Cai, Y.; Zhu, L.; Yao, J. Preparation and characterization of
nano-hydroxyapatite/silk fibroin porous scaffolds. J. Biomater. Sci. Polym. 2008, 19, 325–328. [CrossRef]
[PubMed]
34. Manju, V.; Anitha, A.; Menon, D.; Iyer, S.; Nair, S.V.; Nair, M.B. Nanofibrous yarn reinforced HA-gelatin
composite scaffolds promote bone formation in critical sized alveolar defects in rabbit model. Biomed. Mater.
2018, 13, 065011. [CrossRef] [PubMed]
35. Chen, P.; Wu, Z.; Leung, A.; Chen, X.; Landao-Bassonga, E.; Gao, J.; Chen, L.; Zheng, M.; Yao, F.; Yang, H.; et al.
Fabrication of a silver nanoparticle-coated collagen membrane with anti-bacterial and anti-inflammatory
activities for guided bone regeneration. Biomed. Mater. 2018, 13, 065014. [CrossRef] [PubMed]
36. Han, J.; Ma, B.; Liu, H.; Wang, T.; Wang, F.; Xie, C.; Li, M.; Liu, H.; Ge, S. Hydroxyapatite nanowires modified
polylactic acid membrane plays barrier/osteoinduction dual roles and promotes bone regeneration in a rat
mandible defect model. J. Biomed. Mater. Res. A 2018, 106, 3099–3110. [CrossRef] [PubMed]
37. Chen, X.; Shi, Z.; Bai, S.; Li, B.; Huan Liu, H.; Guo, F.; Wu, G.; Liu, S.; Zhao, Y. Fabrication of gelatin
methacrylate/nanohydroxyapatite microgel arrays for periodontal tissue regeneration. Int. J. Nanomed. 2016,
11, 4707–4718.
38. Nie, L.; Zhang, H.; Ren, A.; Li, Y.; Fu, G.; Cannon, R.D.; Ji, P.; Wu, X.; Yang, S. Nano-hydroxyapatite mineralized
silk fibroin porous scaffold for tooth extraction site preservation. Dent. Mater. 2019, 35, 1397–1407. [CrossRef]
39. Dayashankar, C.P.; Deepika, P.C.; Siddaramaiah, B. Clinical and Radiographic Evaluation of Citric Acid-Based
Nano Hydroxyapatite Composite Graft in the Regeneration of Intrabony Defects—A Randomized Controlled
Trial. Contemp. Clin. Dent. 2017, 8, 380–386. [CrossRef]
40. Vaquette, C.; Fan, W.; Xiao, Y.; Hamlet, S.; Hutmacher, D.W.; Ivanovski, S. A biphasic scaffold design
combined with cell sheet technology for simultaneous regeneration of alveolar bone/periodontal ligament
complex. Biomaterials 2012, 33, 5560–5573. [CrossRef]
41. Ogawa, K.; Miyaji, H.; Kato, A.; Kosen, Y.; Momose, T.; Yoshida, T.; Nishida, E.; Miyata, S.; Murakami, S.;
Takita, H.; et al. Periodontal tissue engineering by nano beta-tricalcium phosphate scaffold and fibroblast
growth factor-2 in one-wall infrabony defects of dogs. J. Periodontal. Res. 2016, 51, 758–767. [CrossRef]
Materials 2020, 13, 201 17 of 20

42. Camilleri, J. Is mineral trioxide Aggregate a Bioceramic. ODOVTOS Int. J. Dent. Sci. 2016, 18, 13–17.
[CrossRef]
43. Saghiri, M.A.; Orangi, J.; Tanideh, N.; Asatourian, A.; Janghorban, K.; Garcia-Godoy, F.; Sheibani, N. Repair
of bone defect by nano-modifed white mineral trioxide aggregates in rabbit: A histopathological study. Med.
Oral. Patol. Oral Cir. Bucal 2015, 20, 525–531. [CrossRef] [PubMed]
44. Lipski, A.M.; Pino, C.J.; Haselton, F.R.; Chen, I.W.; Shastri, V.P. The effect of silica nanoparticle-modified
surfaces on cell morphology, cytoskeletal organization and function. Biomaterials 2008, 29, 3836–3846.
[CrossRef] [PubMed]
45. Moller, K.; Kobler, J.; Bein, T. Colloidal suspensions of nanometer-sized mesoporous silica. Adv. Funct. Mater.
2007, 17, 605–612. [CrossRef]
46. Yeh, Y.C.; Creran, B.; Rotello, V.M. Gold nanoparticles: Preparation, properties, and applications in
bionanotechnology. Nanoscale 2012, 4, 1871–1880. [CrossRef] [PubMed]
47. Vigderman, L.; Khanal, B.P.; Zubarev, E.R. Functional gold nanorods: Synthesis, self-assembly, and sensing
applications. Adv. Mater. 2012, 24, 4811–4841. [CrossRef]
48. Takanche, J.S.; Kim, J.E.; Kim, J.S.; Lee, M.H.; Jeon, J.G.; Park, I.S.; Yi, H.K. Chitosan-gold nanoparticles
mediated gene delivery of c-myb facilitates osseointegration of dental implants in ovariectomized rat.
Artif. Cells Nanomed. Biotechnol. 2018, 46, S807–S817. [CrossRef]
49. Colombo, M.; Carregal-Romero, S.; Casula, M.F.; Gutiérrez, L.; Morales, M.P.; Böhm, I.B.; Heverhagen, J.T.;
Prosperi, D.; Parak, W.J. Biological applications of magnetic nanoparticles. Chem. Soc. Rev. 2012, 41, 4306–4334.
[CrossRef]
50. Bangham, A.D. Surrogate cells or Trojan horses. The discovery of liposomes. Bioessays 1995, 17, 1081–1088.
[CrossRef]
51. Immordino, M.L.; Dosio, F.; Cattel, L. Stealth liposomes: Review of the basic science, rationale, and clinical
applications, existing and potential. Int. J. Nanomed. 2006, 1, 297–315.
52. Allen, T.M.; Cullis, P.R. Liposomal drug delivery systems: From concept to clinical applications. Adv. Drug
Deliv. Rev. 2013, 65, 36–48. [CrossRef] [PubMed]
53. Nicolas, J.; Mura, S.; Brambilla, D.; Mackiewicz, N.; Couvreur, P. Design, functionalization strategies and
biomedical applications of targeted biodegradable/biocompatible polymer-based nanocarriers for drug
delivery. Chem. Soc. Rev. 2013, 42, 1147–1235. [CrossRef] [PubMed]
54. Dhandayuthapani, B.; Yoshida, Y.; Maekawa, T.; Kumar, D.S. Polymeric scaffolds in tissue engineering
application: A review. Int. J. Polym. Sci. 2011, 290602. [CrossRef]
55. Lutolf, M.P.; Weber, F.E.; Schmoekel, H.G.; Schense, J.C.; Kohler, T.; Müller, R.; Hubbell, J.A. Repair of bone
defects using synthetic mimetics of collagenous extracellular matrices. Nat. Biotechnol. 2003, 21, 513–518.
[CrossRef] [PubMed]
56. Ye, H.; Zhu, J.; Deng, D.; Jin, S.; Li, J.; Man, Y. Enhanced osteogenesis and angiogenesis by
PCL/chitosan/Sr-doped calcium phosphate electrospun nanocomposite membrane for guided bone
regeneration. J. Biomater. Sci. Polym. Ed. 2019, 30, 1505–1522. [CrossRef] [PubMed]
57. Eivazzadeh-Keihan, R.; Maleki, A.; de la Guardia, M.; Bani, M.S.; Chenab, K.K.; Pashazadeh-Panahi, P.;
Baradaran, B.; Mokhtarzadeh, A.; Hamblin, M.R. Carbon based nanomaterials for tissue engineering of bone:
Building new bone on small black scaffolds: A review. J. Adv. Res. 2019, 18, 185–201. [CrossRef]
58. La, W.G.; Jin, M.; Park, S.; Yoon, H.H.; Jeong, G.J.; Bhang, S.H.; Park, H.; Char, K.; Kim, B.S. Delivery of bone
morphogenetic protein-2 and substance P using graphene oxide for bone regeneration. Int. J. Nanomed. 2014,
9, 107–116.
59. Zhou, Q.; Yang, P.; Li, X.; Liu, H.; Ge, S. Bioactivity of periodontal ligament stem cells on sodium titanate
coated with graphene oxide. Sci. Rep. 2016, 6, 19343. [CrossRef]
60. Radunovic, M.; De Colli, M.; De Marco, P.; Di Nisio, C.; Fontana, A.; Piattelli, A.; Cataldi, A.; Zara, S. Graphene
oxide enrichment of collagen membranes improves DPSCs differentiation and controls inflammation
occurrence. J. Biomed. Mater. Res. A. 2017, 105, 2312–2320. [CrossRef]
61. De Marco, P.; Zara, S.; De Colli, M.; Radunovic, M.; Lazovi’c, V.; Ettorre, V.; Di Crescenzo, A.; Piattelli, A.;
Cataldi, A.; Fontana, A. Graphene oxide improves the biocompatibility of collagen membranes in an
in vitromodel of human primary gingival fibroblasts. Biomed. Mater. 2017, 12, 055005. [CrossRef]
62. Liao, C.; Li, Y.; Tjong, S.C. Graphene Nanomaterials: Synthesis, Biocompatibility, and Cytotoxicity. Int. J.
Mol. Sci. 2018, 19, 3564. [CrossRef] [PubMed]
Materials 2020, 13, 201 18 of 20

63. Bullock, C.J.; Bussy, C. Biocompatibility Considerations in the Design of Graphene Biomedical Materials.
Adv. Mater. Interfaces 2019, 6, 1900229. [CrossRef]
64. Wang, K.; Ruan, J.; Song, H.; Zhang, J.; Wo, Y.; Guo, S.; Cui, D. Biocompatibility of Graphene Oxide.
Nanoscale Res. Lett. 2011, 6, 8. [CrossRef] [PubMed]
65. Rodríguez-Lozano, F.J.; García-Bernal, D.; Aznar-Cervantes, S.; Ros-Roca, M.A.; Algueró, M.C.; Atucha, N.M.;
Lozano-García, A.A.; Moraleda, J.M.; Cenis, J.L. Effects of composite films of silk fibroin and graphene oxide
on the proliferation, cell viability and mesenchymal phenotype of periodontal ligament stem cells. J. Mater.
Sci. Mater. Med. 2014, 25, 2731–2741. [CrossRef]
66. Vera-Sánchez, M.; Aznar-Cervantes, S.; Jover, E.; García-Bernal, D.; Oñate-Sánchez, R.E.;
Hernández-Romero, D.; Moraleda, J.M.; Collado-González, M.; Rodríguez-Lozano, F.J.; Cenis, J.L.
Silk-Fibroin and Graphene Oxide Composites Promote Human Periodontal Ligament Stem Cell Spontaneous
Differentiation into Osteo/Cementoblast-Like Cells. Stem Cells Dev. 2016, 25, 1742–1754. [CrossRef]
67. Ren, N.; Li, J.; Qiu, J.; Yan, M.; Liu, H.; Ji, D.; Huang, J.; Yu, J.; Liu, H. Growth and accelerated differentiation
of mesenchymal stem cells on graphene-oxide-coated titanate with dexamethasone on surface of titanium
implants. Dent. Mater. 2017, 33, 525–535. [CrossRef]
68. Kinoshita, Y.; Maeda, H. Recent developments of functional scaffolds for craniomaxillofacial bone tissue
engineering applications. Sci. World J. 2013, 21, 863157. [CrossRef]
69. Huang, Y.; Ren, J.; Ren, T.; Gu, S.; Tan, Q.; Zhang, L.; Lv, K.; Pan, K.; Jiang, X. Bone marrow stromal cells
cultured on poly (lactide-co-glycolide)/nanohydroxyapatite composites with chemical immobilization of
Arg-Gly-Asp peptide and preliminary bone regeneration of mandibular defect thereof. J. Biomed. Mater.
Res. A 2010, 95, 993–1003. [CrossRef]
70. Han, X.; Liu, H.; Wang, D.; Su, F.; Zhang, Y.; Zhou, W.; Li, S.; Yang, R. Alveolar bone regeneration around
immediate implants using an injectable nHAC/CSH loaded with autogenic blood-acquired mesenchymal
progenitor cells: An experimental study in the dog mandible. Clin. Implant Dent. Relat. Res. 2013, 15,
390–401. [CrossRef]
71. Zhang, J.C.; Lu, H.Y.; Lv, G.Y.; Mo, A.C.; Yan, Y.G.; Huang, C. The repair of critical-size defects with
porous hydroxyapatite/polyamide nanocomposite: An experimental study in rabbit mandibles. Int. J. Oral
Maxillofac. Surg. 2010, 39, 469–477. [CrossRef]
72. Du, B.; Liu, W.; Deng, Y.; Li, S.; Liu, X.; Gao, Y.; Zhou, L. Angiogenesis and bone regeneration of porous
nano-hydroxyapatite/coralline blocks coated with rhVegF165 in critical-size alveolar bone defects in vivo.
Int. J. Nanomed. 2015, 10, 2555–2565.
73. Su, J.; Xu, H.; Sun, J.; Gong, X.; Zhao, H. Dual delivery of BMP-2 and bFGF from a new nano-composite
scaffold, loaded with vascular stents for large size mandibular defect regeneration. Int. J. Mol. Sci. 2013,
14, 12714–12728. [CrossRef] [PubMed]
74. Zhao, F.; Yin, Y.; Lu, W.W.; Leong, J.C.; Zhang, W.; Zhang, J.; Zhang, M.; Yao, K. Preparation and histological
evaluation of biomimetic three-dimensional hydroxyapatite/chitosan-gelatin network composite scaffolds.
Biomaterials 2002, 23, 3227–3234. [CrossRef]
75. Xynos, I.; Hukkanen, M.; Batten, J.; Buttery, L.; Hench, L.; Polak, J. Bioglass® 45S5 stimulates osteoblast
turnover and enhances bone formation in vitro: Implications and applications for bone tissue engineering.
Calcif. Tissue Int. 2000, 67, 321–329. [CrossRef]
76. Kim, H.D.; Valentini, R.F. Retention and activity of BMP-2 in hyaluronic acid-based scaffolds in vitro.
J. Biomed. Mater. Res. 2002, 59, 573–584. [CrossRef] [PubMed]
77. El-Ghannam, A.R. Advanced bioceramic composite for bone tissue engineering: Design principles and
structure—Bioactivity relationship. J. Biomed. Mater. Res. Part A. 2004, 69, 490–501. [CrossRef]
78. Iviglia, G.; Saeid Kargozar, S.; Baino, F. Biomaterials, Current Strategies, and Novel Nano-Technological
Approaches for Periodontal Regeneration. J. Funct. Biomater. 2019, 10, 3. [CrossRef]
79. Li, G.; Zhang, T.; Li, M.; Fu, N.; Fu, Y.; Ba, K.; Deng, S.; Jiang, Y.; Hu, J.; Peng, Q.; et al. Electrospun fibers for
dental and craniofacial applications. Curr. Stem Cell Res. 2014, 9, 187–195. [CrossRef]
80. Gmeiner, R.; Deisinger, U.; Schönherr, J.; Lechner, B.; Detsch, R.; Boccaccini, A.; Stampfl, J. Additive
manufacturing of bioactive glasses and silicate bioceramics. J. Ceram. Sci. Technol. 2015, 6, 75–86.
81. Kosachan, N.; Jaroenworaluck, A.; Jiemsirilers, S.; Jinawath, S.; Stevens, R. Hydroxyapatite nanoparticles
formed under a wet mechanochemical method. J. Biomed. Mater. Res. Part B Appl. Biomater. 2017, 105, 679–688.
[CrossRef]
Materials 2020, 13, 201 19 of 20

82. Ivanovski, S.; Vaquette, C.; Gronthos, S.; Hutmacher, D.W.; Bartold, P.M. Multiphasic Scaffolds for Periodontal
Tissue Engineering. J. Dent. Res. 2014, 93, 1212–1221. [CrossRef]
83. Carter, S.D.; Costa, P.F.; Vaquette, C.; ivanovski, S.; Hutmacher, D.W.; Malda, J. Additive Biomanufacturing:
An Advanced Approach for Periodontal Tissue Regeneration. Ann. Biomed. Eng. 2017, 45, 12–22. [CrossRef]
[PubMed]
84. Carlo-Reis, E.C.; Borges, A.P.; Araújo, M.V.; Mendes, V.C.; Guan, L.; Davies, J.E. Periodontal regeneration
using a bilayered PLGA/calcium phosphate construct. Biomaterials 2011, 32, 9244–9253. [CrossRef] [PubMed]
85. Requicha, J.F.; Viegas, C.A.; Muñoz, F.; Azevedo, J.M.; Leonor, I.B.; Reis, R.L.; Gomes, M.E. A tissue
engineering approach for periodontal regeneration based on a biodegradable double-layer scaffold and
adipose-derived stem cells. Tissue Eng. Part A 2014, 20, 2483–2492. [CrossRef] [PubMed]
86. Park, C.H.; Rios, H.F.; Jin, Q.; Bland, M.E.; Flanagan, C.L.; Hollister, S.J.; Giannobile, W.V. Biomimetic hybrid
scaffolds for engineering human tooth ligament interfaces. Biomaterials 2010, 31, 5945–5952. [CrossRef]
87. Park, C.H.; Rios, H.F.; Jin, Q.; Sugai, J.V.; Padial-Molina, M.; Taut, A.D.; Flanagan, C.L.; Hollister, S.J.;
Giannobile, W.V. Tissue engineering bone-ligament complexes using fiber-guiding scaffolds. Biomaterials
2012, 33, 137–145. [CrossRef]
88. Vaquette, C.; Cooper-White, J. A simple method for fabricating 3-D multilayered composite scaffolds. Acta
Biomater. 2013, 9, 4599–4608. [CrossRef]
89. Costa, P.F.; Vaquette, C.; Zhang, Q.; Reis, R.L.; Ivanovski, S.; Hutmacher, D.W. Advanced tissue engineering
scaffold design for regeneration of the complex hierarchical periodontal structure. J. Clin. Periodontol. 2014,
41, 283–294. [CrossRef]
90. Lee, C.H.; Hajibandeh, J.; Suzuki, T.; Fan, A.; Shang, P.; Mao, J.J. Three dimensional printed multiphase
scaffolds for regeneration of periodontium complex. Tissue Eng. Part A 2014, 20, 1342–1351. [CrossRef]
91. Murphy, S.V.; Atala, A. 3D bioprinting of tissues and organs. Nat. Biotechnol. 2014, 32, 773–785. [CrossRef]
92. Dababneh, A.B.; Ozbolat, I.T. Bioprinting technology: A current state-of-the-art review. J. Manuf. Sci. Eng.
2014, 136, 061016. [CrossRef]
93. Berthiaume, F.; Maguire, T.J.; Yarmush, M.L. Tissue engineering and regenerative medicine: History, progress,
and challenges. Annu. Rev. Chem. Biomol. Eng. 2011, 2, 403–430.
94. Zhang, Y.S.; Yue, K.; Aleman, J.; Mollazadeh-Moghaddam, K.; Bakht, S.M.; Yang, J.; Weitao, J.A.; Dell’Erba, V.;
Assawes, P.; Shin, S.R.; et al. 3D Bioprinting for Tissue and Organ Fabrication. Ann. Biomed. Eng. 2017,
45, 148–163. [CrossRef] [PubMed]
95. Yun, H.M.; Ahn, S.J.; Park, K.R.; Kim, M.J.; Kim, J.J.; Jin, G.Z.; Kim, H.W.; Ahn, S.J.; Kim, H.W.; Kim, E.C.
Magnetic nanocomposite scaffolds combined with static magnetic field in the stimulation of osteoblastic
differentiation and bone formation. Biomaterials 2016, 85, 88–98. [CrossRef] [PubMed]
96. Sculean, A.; Nikolidakis, D.; Nikou, G.; Ivanovic, A.; Chapple, I.L.; Stavropoulos, A. Biomaterials for
promoting periodontal regeneration in human intrabony defects: A systematic review. Periodontology 2000
2015, 68, 182–216. [CrossRef]
97. Kargozar, S.; Mozafari, M. Nanotechnology and Nanomedicine: Start small, think big. Mater. Today Proc.
2018, 5, 15492–15500. [CrossRef]
98. Neel, E.A.A.; Chrzanowski, W.; Salih, V.M.; Kim, H.W.; Knowles, J.C. Review Tissue engineering in dentistry.
J. Dent. 2014, 42, 915–928. [CrossRef]
99. Zhang, Y.; Miron, R.J.; Li, S.; Shi, B.; Sculean, A.; Cheng, X. Novel Meso Porous BioGlass/silk scaffold
containing ad PDGF-B and ad BMP 7 for the repair of periodontal defects in beagle dogs. J. Clin. Periodontol.
2015, 42, 262–271. [CrossRef]
100. Yue, K.; Trujillo-de Santiago, G.; Alvarez, M.M.; Tamayol, A.; Annabi, N.; Khademhosseini, A. Synthesis,
properties, and biomedical applications of gelatin methacryloyl (GelMA) hydrogels. Biomaterials 2015,
73, 254–271. [CrossRef]
101. Chen, X.; Liu, Y.; Miao, L.; Wang, Y.; Ren, S.; Yang, X.; Hu, Y.; Sun, W. Controlled release of recombinant human
cementum protein 1 from electrospun multiphasic scaffold for cementum regeneration. Int. J. Nanomed. 2016,
11, 3145–3158.
102. Guazzo, R.; Gardin, C.; Bellin, G.; Sbricoli, L.; Ferroni, L.; Ludovichetti, F.S.; Piattelli, A.; Antoniac, I.;
Bressan, E.; Zavan, B. Graphene-Based Nanomaterials for Tissue Engineering in the Dental Field. Nanomaterials
2018, 8, 349. [CrossRef]
Materials 2020, 13, 201 20 of 20

103. Zhang, Y.; Song, J.; Shia, B.; Wanga, Y.; Chena, X.; Huanga, C.; Yanga, X.; Xua, D.; Cheng, X.; Chen, X.
Combination of scaffold and adenovirus vectors expressing bone morphogenetic protein-7 for alveolar bone
regeneration at implant defects. Biomaterials 2007, 28, 4635–4642. [CrossRef] [PubMed]
104. Rasperini, G.; Pilipchuk, S.P.; Flanagan, C.L.; Park, C.H.; Pagni, G.; Hollister, S.J.; Giannobile, W.V. 3D-printed
bioresorbable scaffold for periodontal repair. J. Dent. Res. 2015, 94, 153–157. [CrossRef] [PubMed]
105. Feng, X.; Chen, A.; Zhang, Y.; Wang, J.; Shao, L.; Wei, L. Application of dental nanomaterials: Potential
toxicity to the central nervous system. Int. J. Nanomed. 2015, 10, 3547–3565.
106. Lee, C.M.; Jeong, H.J.; Yun, K.N.; Kim, D.W.; Sohn, M.H.; Lee, J.K.; Jeong, J.; Lim, S.T. Optical imaging to
trace near infrared fluorescent zinc oxide nanoparticles following oral exposure. Int. J. Nanomed. 2012,
7, 3203–3209.
107. Wang, Y.; Chen, Z.; Ba, T.; Pu, J.; Chen, T.; Song, Y.; Gu, Y.; Qian, Q.; Xu, Y.; Xiang, K.; et al. Susceptibility
of young and adult rats to the oral toxicity of titanium dioxide nanoparticles. Small 2013, 9, 1742–1752.
[CrossRef] [PubMed]
108. Fernandez-Urrusuno, R.; Fattal, E.; Rodrigues, J.M., Jr.; Féger, J.; Bedossa, P.; Couvreur, P. Effect of polymeric
nanoparticle administration on the clearance activity of the mononuclear phagocyte system in mice. J. Biomed.
Mater. Res. 1996, 31, 401–408. [CrossRef]
109. Karmakar, A.; Zhang, Q.; Zhang, Y. Neurotoxicity of nanoscale materials. J. Food Drug Anal. 2014, 22, 147–160.
[CrossRef]
110. Ma, L.; Liu, J.; Li, N.; Wang, J.; Duan, Y.; Yan, J.; Liu, H.; Wang, H.; Hong, F. Oxidative stress in the brain
of mice caused by translocated nanoparticulate TiO2 delivered to the abdominal cavity. Biomaterials 2010,
31, 99–105. [CrossRef]
111. Adamcakova-Dodd, A.; Stebounova, L.V.; Kim, J.S.; Vorrink, S.U.; Ault, A.P.; O’Shaughnessy, P.T.;
Grassian, V.H.; Thorne, P.S. Toxicity assessment of zinc oxide nanoparticles using sub-acute and sub-chronic
murine inhalation models. Part Fibretoxicol. 2014, 1, 11–15. [CrossRef]
112. Xie, H.; Chua, M.; Islam, I.; Bentini, R.; Cao, T.; Viana-Gomes, J.C.; Castro Neto, A.H.; Rosa, V. CVD-grown
monolayer graphene induces osteogenic but not odontoblastic differentiation of dental pulp stem cells.
Dent. Mater. 2017, 33, e13–e21. [CrossRef]
113. Launey, M.E.; Munch, E.; Alsem, D.H.; Barth, H.B.; Saiz, E.; Tomsia, A.P.; Ritchie, R.O. Designing
highly toughened hybrid composites through nature-inspired hierarchical complexity. Acta Mater. 2009,
57, 2919–2932. [CrossRef]
114. Dzenis, Y. Materials science—Structural nanocomposites. Science 2008, 319, 419–420. [CrossRef] [PubMed]
115. Ritchie, R.O. The quest for stronger, tougher materials. Science 2008, 320, 448. [CrossRef] [PubMed]
116. Evans, A.G. Perspective on the development of high-toughness ceramics. J. Am. Ceram Soc. 1990, 73, 187–206.
[CrossRef]

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).