Advanced reconstructive technologies for periodontal tissue repair.
ABSTRACT Reconstructive therapies to promote the regeneration of lost periodontal support have been investigated through both preclinical and clinical studies. Advanced regenerative technologies using new barrier-membrane techniques, cell-growth-stimulating proteins or gene-delivery applications have entered the clinical arena. Wound-healing approaches using growth factors to target the restoration of tooth-supporting bone, periodontal ligament and cementum are shown to significantly advance the field of periodontal-regenerative medicine. Topical delivery of growth factors, such as platelet-derived growth factor, fibroblast growth factor or bone morphogenetic proteins, to periodontal wounds has demonstrated promising results. Future directions in the delivery of growth factors or other signaling models involve the development of innovative scaffolding matrices, cell therapy and gene transfer, and these issues are discussed in this paper.
- [show abstract] [hide abstract]
ABSTRACT: Bone morphogenetic proteins (BMP) are secretory signal molecules which have a variety of regulatory functions during morphogenesis and cell differentiation. Teeth are typical examples of vertebrate organs in which development is controlled by sequential and reciprocal signaling between the epithelium and mesenchyme. In addition, tooth development is characterized by formation of mineralized tissues: the bone-like dentin and cementum as well as epithelially derived enamel. We have performed a comparative in situ hybridization analysis of the expression of six different Bmps (Bmp-2 to Bmp-7) starting from initiation of tooth development to completion of crown morphogenesis when dentine and enamel matrices are being deposited. Bmps-2, -4, and -7 were frequently codistributed and showed marked associations with epithelial-mesenchymal interactions. Their expression shifted between the epithelium and mesenchyme starting from the stage of tooth initiation. They were subsequently expressed in the enamel knot, the putative signaling center regulating tooth shape. Their expression domains prior to and during the differentiation of the dentine-forming odontoblasts and enamel-forming ameloblasts was in line with functions in regulation of cell differentiation and/or secretory activities of the cells. The expression of Bmp-3 was confined to mesenchymal cells, in particular to the dental follicle cells which give rise to the cementoblasts, forming the hard tissue covering the roots of teeth. Bmp-5 was expressed only in the epithelial ameloblasts. It was upregulated as the cells started to polarize and intense expression continued in the secretory ameloblasts. Bmp-6 was expressed only weakly in the dental mesenchyme during bud and cap stages. Our results are in line with regulatory functions of Bmps at all stages of tooth morphogenesis. Bmps-2, -4, and -7 are conceivably parts of signaling networks regulating tooth initiation and shape development. They as well as Bmp-5 may be involved in the induction and formation of dentine and enamel, and Bmp-3 in the development of cementum. The remarkable overlaps in the expression domains of different Bmp genes may implicate functional redundancy and/or formation of active heterodimers between different BMPs.Developmental Dynamics 01/1998; 210(4):383-96. · 2.59 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: Cementum, a mineralized tissue lining the tooth root surface, is destroyed during the inflammatory process of periodontitis. Restoration of functional cementum is considered a criterion for successful regeneration of periodontal tissues, including formation of periodontal ligament, cementum, and alveolar bone. Short-term administration of platelet-derived growth factor (PDGF) has been shown to partially regenerate periodontal structures. Nonetheless, the role of PDGF in cementogenesis is not well understood. The aim of the present study was to determine the effect of sustained PDGF gene transfer on cementum formation in an ex vivo ectopic biomineralization model. Osteocalcin (OC) promoter-driven SV40 transgenic mice were used to obtain immortalized cementoblasts (OCCM). The OCCM cells were transduced with adenoviruses (Ad) encoding either PDGF-A, an antagonist of PDGF signaling (PDGF-1308), a control virus (green fluorescent protein, GFP), or no treatment (NT). The transduced cells were incorporated into polymer scaffolds and implanted subcutaneously into severe combined immunodeficient (SCID) mice. The implants were harvested at 3 and 6 weeks for histomorphometric analysis of the newly formed mineralized tissues. Northern blot analysis was performed to determine the expression levels of mineral-associated genes including bone sialoprotein (BSP), OC, and osteopontin (OPN) in the cell-implant specimens at 3 and 6 weeks. The results indicated mineralization was significantly reduced in both the Ad/PDGF-A and Ad/PDGF-1308 treated specimens when compared to the NT or Ad/GFP groups at 3 and 6 weeks (P<0.01). In addition, the size of the implants treated with Ad/PDGF-A and Ad/PDGF-1308 was significantly reduced compared to implants from Ad/GFP and NT groups at 3 weeks (P<0.05). At 6 weeks, the size of implants and mineral formation increased in NT, Ad/GFP, and Ad/PDGF-A groups, while the Ad/PDGF-1308 treated implants continued to decrease in size and mineral formation (P<0.01). Northern blot analysis revealed that in the Ad/PDGF-A treated implants OPN was increased, whereas OC gene expression was downregulated at 3 weeks. In the Ad/PDGF-1308 treated implants, BSP, OC, and OPN were all downregulated at 3 weeks. At 3 weeks, the Ad/PDGF-A treated implants contained significantly higher multinucleated giant cell (MNGC) density compared to NT, Ad/GFP, and Ad/PDGF-1308 specimens. The MNGC density in NT, Ad/GFP, and Ad/PDGF-A treated groups reduced over time, while the Ad/PDGF-1308 transduced implants continued to exhibit significantly higher MNGC density compared with the other treatment groups at 6 weeks. The results showed that continuous exposure to PDGF-A had an inhibitory effect on cementogenesis, possibly via the upregulation of OPN and subsequent enhancement of MNGCs at 3 weeks. On the other hand, Ad/PDGF-1308 inhibited mineralization of tissue-engineered cementum possibly due to the observed downregulation of BSP and OC and a persistence of stimulation of MNGCs. These findings suggest that continuous exogenous delivery of PDGF-A may delay mineral formation induced by cementoblasts, while PDGF is clearly required for mineral neogenesis.Journal of Periodontology 03/2004; 75(3):429-40. · 2.40 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: Destruction of tooth support due to the chronic inflammatory disease periodontitis is a major cause of tooth loss. There are limitations with available treatment options to tissue engineer soft tissue periodontal defects. The exogenous application of growth factors (GFs) such as platelet-derived growth factor (PDGF) has shown promise to enhance oral and periodontal tissue regeneration. However, the topical administration of GFs has not led to clinically significant improvements in tissue regeneration because of problems in maintaining therapeutic protein levels at the defect site. The utilization of PDGF gene transfer may circumvent many of the limitations with protein delivery to soft tissue wounds. The objective of this study was to test the effect of PDGF-A and PDGF-B gene transfer to human gingival fibroblasts (HGFs) on ex vivo repair in three-dimensional collagen lattices. HGFs were transduced with adenovirus encoding PDGF-A and PDGF-B genes. Defect fill of bilayer collagen gels was measured by image analysis of cell repopulation into the gingival defects. The modulation of gene expression at the defect site and periphery was measured by RT-PCR during a 10-day time course after gene delivery. The results demonstrated that PDGF-B gene transfer stimulated potent (>4-fold) increases in cell repopulation and defect fill above that of PDGF-A and corresponding controls. PDGF-A and PDGF-B gene expression was maintained for at least 10 days. PDGF gene transfer upregulated the expression of phosphatidylinosital 3-kinase and integrin alpha5 subunit at 5 days after adenovirus transduction. These results suggest that PDGF gene transfer has potential for periodontal soft tissue-engineering applications.Tissue Engineering 09/2003; 9(4):745-56. · 4.07 Impact Factor
technologies for periodontal
CHRISTOPH A. RAMSEIER, GIULIO RASPERINI, SALVATORE BATIA &
WILLIAM V. GIANNOBILE
Regenerative periodontal therapy uses specific tech-
niques designed to restore those parts of the tooth-
supporting structures that have been lost as a result
of periodontitis or gingival trauma. The term ?regen-
eration? is defined as the reconstruction of lost or
injured tissues in such a way that both the original
structures and their function are completely restored.
Procedures aimed at restoring lost periodontal tissues
favor the creation of new attachment, including the
formation of a new periodontal ligament with its
fibers inserting in newly formed cementum and
Deep infrabony defects associated with periodontal
pockets are the classic indication for periodontal-
regenerative therapy. Different degrees of furcation
involvement in molars and upper first premolars are a
further indication for regenerative approaches as the
furcation area remains difficult to maintain through
instrumentation and oral hygiene. A third group of
indications for regenerative periodontal therapy are
localized gingival recession and root exposure be-
cause they may cause significant esthetic concern for
the patient. The denuding of a root surface with
resultant root sensitivity represents a further indica-
tion for regenerative periodontal therapy in order to
reduce root sensitivity and to improve esthetics.
Professional periodontal therapy and maintenance,
combined with risk-factor control, are shown to
effectively reduce periodontal disease progression (7,
128). In contrast to the conventional approaches of
anti-inflammatory periodontal therapy, however, the
regenerative procedures aimed at repairing lost
periodontal tissues, including alveolar bone, peri-
odontal ligament and root cementum, remain more
challenging (24). During the last few decades, peri-
odontal research has systematically attempted to
identify clinical procedures that are predictably suc-
cessful in regenerating periodontal tissues. Hence,
the extent to which various methods, in combination
with regenerative biomaterials, such as hard- and
soft-tissue grafts, or cell-occlusive barrier mem-
branes used in guided tissue-regeneration proce-
dures, are able to regenerate lost tooth support has
been investigated (162).
Periodontal regeneration is assessed using probing
measures, radiographic analysis, direct measure-
ments of new bone and histology (133). Many cases
that are considered clinically successful, including
those in which significant regrowth of alveolar bone
occurs, may histologically still show an epithelial
lining along the treated root surface, instead of newly
formed periodontal ligament and cementum (84). In
general, however, the clinical outcome of periodon-
tal-regenerative techniques is shown to depend on:
(i) patient-associated factors, such as plaque control,
smoking habits, residual periodontal infection, or
membrane exposure in guided tissue-regeneration
procedures, (ii) effects of occlusal forces that deliver
intermittent loads in axial and transverse dimensions,
as well as (iii) factors associated with the clinical skills
of the operator, such as lack of primary closure of the
surgical wound (93). Even though modified flap de-
signs and microsurgical approaches are shown to
positively affect the outcome of both soft- and hard-
tissue regeneration, the clinical success for peri-
odontal regeneration still remains limited in many
cases. Moreover, the surgical protocols for regenera-
tive procedures are skill-demanding and may there-
fore lack practicability for a number of clinicians.
Consequently, both clinical and preclinical research
approaches using new barrier-membrane techniques
Periodontology 2000, Vol. 59, 2012, 1–19
Printed in Singapore. All rights reserved
? 2012 John Wiley & Sons A/S
(69), cell-growth-stimulating proteins (28, 44, 70) or
gene-delivery applications (125) in order to simplify
and enhance the rebuilding of missing periodontal
support. The aim of our review was to compare these
regenerative techniques (Table 1). While the focus
will be on clinical applications for the delivery of
growth factors, the applications for gene delivery of
tissue growth factors are also reviewed.
Periodontal wound healing
Previous research on periodontal wound healing has
provided a basic understanding of the mechanisms
favoring periodontal tissue regeneration. A number of
valuable findings at both the cellular and molecular
levels was revealed and subsequently used to engi-
neer the regenerative biomaterials currently available
in periodontal medicine. In order to provide an
overview of the cellular and molecular events and
their association with periodontal tissue regenera-
tion, the course of periodontal wound healing is
briefly reviewed in this article.
The biology and principles of periodontal wound
healing have previously been reviewed (123). Based
on observations following experimental incisions in
periodontal soft tissues, the sequence of healing after
blood-clot formation is commonly divided into the
following phases: (i) soft-tissue inflammation, (ii)
granulation-tissue formation, and (iii) intercellular
matrix formation and remodeling (22, 150). Plasma
proteins, mainly fibrinogen, accumulate rapidly in
the bleeding wound and provide the initial basis for
the adherence of a fibrin clot (167). The inflammatory
phase of healing in the soft-tissue wound is initiated
by polymorphonuclear leukocytes infiltrating the fi-
brin clot from the wound margins, followed shortly
afterwards by macrophages (114). The major function
of the polymorphonuclear leukocytes is to debride
the wound by removing bacterial cells and injured
tissue particles through phagocytosis. The macro-
phages, in addition, have an important role to play in
the initiation of tissue repair. The inflammatory
phase progresses into its later stage as the amount of
decreases while the macrophage influx continues.
These macrophages contribute to the cleansing pro-
cess through the phagocytosis of used polymorpho-
nuclear leukocytes and erythrocytes. Additionally,
macrophages release a number of biologically active
molecules, such as inflammatory cytokines and tis-
sue growth factors, which recruit further inflamma-
tory cells as well as fibroblastic and endothelial cells,
thus playing an essential role in the transition of the
wound from the inflammatory stage to the granula-
tion tissue-formation stage. The influx of fibroblasts
and budding capillaries from the gingival connective
tissue and the periodontal ligament connective tissue
initiate the phase of granulation-tissue formation in
the periodontal wound approximately 2 days after
incision. At this stage, fibroblasts are responsible for
the formation of a loose new matrix of collagen,
fibronectin and proteoglycans (12). Eventually, cells
and matrix form cell-to-cell and cell-to-matrix links
that generate a concerted tension, resulting in tissue
contraction. The phase of granulation-tissue forma-
tion gradually develops into the final phase of heal-
ing, in which the reformed, more cell-rich tissue,
undergoes maturation and sequenced remodeling to
meet functional needs (22, 150).
The morphology of a periodontal wound comprises
the gingival epithelium, the gingival connective tis-
sue, the periodontal ligament and the hard-tissue
components, such as alveolar bone and cementum or
dentin on the dental root surface (Fig. 1). This par-
ticular composition ultimately affects the healing
events in each tissue component as well as those in
the entire periodontal site. While the healing of gin-
gival epithelia and their underlying connective
tissues concludes in a number of weeks, the regen-
eration of periodontal ligament, root cementum and
alveolar bone generally takes longer, occurring within
a number of weeks or months. Aiming for wound
closure, the final outcome of wound healing in the
epithelium is the formation of the junctional epi-
thelium surrounding the dentition (16). On the other
hand, the healing of gingival connective tissue results
in a significant reduction of its volume, thus clinically
creating both gingival recession and a reduction of
the periodontal pocket. Periodontal ligament is
shown to regenerate on newly formed cementum
created by cementoblasts that have originated from
periodontal ligament granulation tissue (73). Fur-
thermore, alveolar bone modeling occurs following
the stimulation of mesenchymal cells from the
gingival connective tissue that are transformed into
osteoprogenitor cells by locally expressed bone
morphogenetic proteins (78, 154).
A series of classical animal studies demonstrated
that the tissue derived from alveolar bone or gingival
connective tissue lacks cells with the potential to
produce a new attachment between the periodontal
ligament and newly formed cementum (74, 112).
Moreover, granulation tissue derived from the gingi-
Ramseier et al.
Table 1. Regenerative biomaterials currently available for use in periodontology
Regenerative biomaterials Trade name(s) References
Bone autogenous grafts (autografts)
Intra-oral autograftsn ⁄ a Renvert et al. (134)
Ellegaard & Lo ¨e (31)
Extra-oral autograftsn ⁄ aFroum et al. (39)
Bone allogenic grafts (allografts)
Freeze-dried bone allograftGrafton?(Osteotech, Eatontown, NJ, USA),
Lifenet?(LifeNet Health Inc., Virginia Beach,
Mellonig et al. (96)
Demineralized freeze-dried bone
Foundation Inc., Miami, FL, USA)
Gurinsky et al. (52)
Kimble et al. (76)
Trejo et al. (156)
Bone xenogenic grafts (xenografts)
Bovine mineral matrix Bio-Oss?(Geistlich Pharma AG, Wolhusen,
Switzerland), OsteoGraf?(Dentsply, Tulsa, OK,
USA), Pep-Gen P-15?(Dentsply GmbH,
Hartman et al. (55)
Camelo et al. (13)
Nevins et al. (108)
Richardson et al. (136)
Bone alloplastic grafts (alloplasts)
Hydroxyapatite (dense, porous,
Holliswood , NY, USA)
Meffert et al. (95)
Galgut et al. (41)
Beta tricalcium phosphateSynthograph?(Bicon, Boston, MA, USA),
alpha-BSM?(Etex Corp., Cambridge, MA,
Palti & Hoch (117)
Scher et al. (143)
Nery et al. (107)
Hard-tissue replacement polymersBioplant?(Kerr Corp., Orange, CA, USA)Dryankova et al. (29)
Bioactive glass (SiO2, CaO, Na2O,
PerioGlas?(Novabone, Jacksonville, FL, USA),
BioGran?(Biomet 3i, Palm Beach Gardens, FL,
Sculean et al. (146)
Reynolds et al. (135)
Trombelli et al. (158)
Fetner et al. (35)
Coral-derived calcium carbonateBiocoral?(Biocoral Inc., La Garenne Colombes,
Polimeni et al. (122)
Polymer and collagen sponges
CollagenHelistat?(Dental Implant Technologies Inc.,
Scottsdale, AZ, USA), Collacote?(Carlsbad, CA,
USA), Colla-Tec?(Colla-Tec Inc., Plainsboro,
NJ, USA), Gelfoam?(Baxter, Deerfield, IL, USA)
Poly lactide-copolyglycolide barrier membranes
Methylcellulosen ⁄ a Lioubavina-Hack et al. (83)
Hyaluronic acid estern ⁄ a Wikesjo ¨ et al. (163)
Chitosann ⁄ aYeo et al. (171)
Polyethylene glycoln ⁄ aJung et al. (69)
Nonresorbable cell-occlusive barrier membranes
PolytetrafluorethyleneGore-Tex?(W. L. Gore & Associates Inc., New-
ark, DE, USA)
Trombelli et al. (159)
Moses et al. (100)
Murphy & Gunsolley (102)
Needleman et al. (105)
Periodontal tissue-engineering technologies
val connective tissue or alveolar bone results in root
resorption or ankylosis when placed in contact with
the root surface. Therefore, it should be expected that
these complications would occur more frequently
following regenerative periodontal surgery, particu-
larly following those procedures that include the
placement of grafting materials to stimulate bone
formation. The reason for root resorption (which is
rarely observed), however, may be that following the
surgical intervention, the dento–gingival epithelium
migrates apically along the root surface, forming a
protective barrier towards the root surface (11, 75).
The findings from these animal experiments revealed
that ultimately the periodontal ligament tissue con-
tains cells with the potential to form a new connec-
tive tissue attachment (73).
Typically, the down-growth of the epithelium along
the tooth-root surface reaches the level of the peri-
odontal ligament before the latter has regenerated
with new layers of cementum and newly inserting
connective tissue fibers. Therefore, in order to enable
and promote healing towards the rebuilding of
cementum and periodontal ligament, the gingival
epithelium must be prevented from forming a long
junctional epithelium along the root surface down to
the former level of the periodontal ligament (Fig. 2).
This basic acquisition of knowledge has been the key
for the engineering of standard clinical procedures
for the placement of a fabricated membrane in gui-
ded tissue regeneration.
In summary, the principles of periodontal wound
healing presented provide a basic understanding of
the events following wounding in surgical interven-
tions. In order to obtain new connective tissue
attachment, the granulation tissue derived from
periodontal ligament cells has to be given both space
and time to produce and mature new cementum and
periodontal ligament. The conventional guided tis-
Table 1. Continued
Regenerative biomaterialsTrade name(s) References
Resorbable cell-occlusive barrier membranes
Polyglycolide ⁄ Polylactide (synthetic) Ossix?(ColBar LifeScience Ltd., Rehovot, Israel) Minenna et al. (98)
Stavropoulos et al. (153)
Parashis et al. (118)
Collagen membrane (xenogen)Bio-Gide?(Geistlich Pharma AG, Wolhusen,
Sculean et al. (144)
Owczarek et al. (116)
Camelo et al. (15)
Enamel matrix derivative Emdogain?(Straumann AG, Basel, Switzerland) Rasperini et al. (130)
Rosing et al. (139)
Sanz et al. (142)
Francetti et al. (38)
Tonetti et al. (155)
Esposito et al. (33)
Esposito et al. (32)
Esposito et al. (34)
Platelet-derived growth factor Gem 21S?(Osteohealth, Shirley, NY, USA) Nevins et al. (110)
Bone morphogenetic proteinInfuse?(Medtronic Inc., Minneapolis, MN,
Fiorellini et al. (36)
Fig. 1. Periodontal wound following flap surgery: (1)
gingival epithelium, (2) gingival connective tissue, (3)
periodontal ligament, (4) alveolar bone and (5) cementum
or dentin on the dental root surface.
Ramseier et al.
sue-regeneration techniques in periodontal practice
have shown their predictable, albeit limited, potential
to regenerate lost periodontal support. Consequently,
advanced regenerative technologies for periodontal
tissue repair aim to increase the current gold stan-
dards for success of periodontal regeneration. In
order to identify appropriate advanced repair tech-
niques for tooth-supporting periodontal tissues, a
number of combinations of conventional regenera-
tive techniques have been evaluated: guided tissue
regeneration and application of tissue growth fac-
tor(s); guided tissue regeneration and hard-tissue
graft and application of tissue growth factor(s); hard-
tissue graft and biomodification of the tooth-root
surface; and hard-tissue graft and application of tis-
sue growth factors.
Advanced repair of alveolar bone
The morphology of the alveolar infrabony defect was
a predictable outcome of regeneration of periodontal
attachment (124). Goldman & Cohen (50) originally
proposed a classification for infrabony defects that
referred to the number of osseous walls surrounding
the defect: one-wall, two-wall or three-wall.
In a number of clinical trials and animal experiments,
the periodontal flap approach was combined with the
placement of bone grafts or implant materials into
the curetted bony defects with the aim of stimulating
periodontal regeneration. The various graft and im-
plant materials evaluated to date are: (i) autogenous
graft: a graft transferred from one location to another
within the same organism; (ii) allogenic graft: a graft
transferred from one organism to another organism
of the same species; (iii) xenogenic graft: a graft taken
from an organism of a different species; and (iv)
alloplastic material: synthetic or inorganic implant
material used instead of the previously mentioned
The biologic rationale behind the use of bone grafts
or alloplastic materials for regenerative approaches is
the assumption that these materials may serve as a
scaffold for bone formation (osteoconduction) and
contain the bone-forming cells (osteogenesis) or
bone-inductive substances (osteoinduction).
Histological studies in both humans and animals
have demonstrated that grafting procedures often
result in healing with a long junctional epithelium
rather than a new connective tissue attachment (17,
84). Therefore, multiple studies have evaluated the
use of hard-tissue graft materials for periodontal
regeneration in infrabony defects when compared
with the periodontal flap approach alone.
Biomodification of the tooth-root surface
A number of studies have focused on the modifica-
tion of the periodontitis-involved root surface in or-
der to advance the formation of a new connective
tissue attachment. However, despite histological
evidence of regeneration
biomodification with citric acid, the outcome of
controlled clinical trials have failed to show any
improvements in clinical conditions compared with
nonacid-treated controls (40, 91, 99).
In recent years, biomodification of the root surface
with enamel matrix proteins during periodontal sur-
gery and following demineralization with EDTA has
been introduced to promote periodontal regenera-
tion. Based on the understanding of the biological
model, the application of enamel matrix proteins
Fig. 2. (A) Normal healing process following adaptation of
the periodontal flap with significant reduction of the
attachment apparatus. (B) In order to enable and promote
healing towards the rebuilding of cementum and peri-
odontal ligament, the gingival epithelium must be pre-
vented from forming a long junctional epithelium along
the root surface down to the former level of the peri-
odontal ligament (e.g., by placement of a bioresorbable
Periodontal tissue-engineering technologies
regeneration as it initiates events that occur during
the growth of periodontal tissues (43, 54). The com-
mercially available product Emdogain?, a purified
acid extract of porcine origin containing enamel
matrix derivates, is reported to be able to enhance
periodontal regeneration (Fig. 3). More basic re-
search, in addition to the clinical findings, indicates
that enamel matrix derivates have a key role in peri-
odontal wound healing (26, 32). Histological results
from both animal and human studies have shown
that the application of enamel matrix derivates pro-
motes periodontal regeneration and confidently
influences periodontal wound healing (147). Thus far,
enamel matrix derivates, either alone or in combi-
nation with grafts, have demonstrated their potential
to effectively treat intraosseous defects and the clin-
ical results appear to be stable long term (157).
isseen to promote periodontal
Periodontal tissue growth factors
Wound-healing approaches using growth factors to
target restoration of tooth-supporting bone, peri-
odontal ligament and cementum have been shown to
significantly advance the field of periodontal-regen-
erative medicine. A major focus of periodontal re-
search has studied the impact of tissue growth factor
on periodontal tissue regeneration (Table 2) (3, 44,
104, 126). Advances in molecular cloning have made
available unlimited quantities of recombinant growth
factors for applications in tissue engineering. Re-
combinant growth factors known to promote skin and
bone wound healing, such as platelet-derived growth
factors (14, 46, 67, 110, 115, 140), insulin-like growth
factors (44, 46, 58, 87), fibroblast growth factors (49,
101, 149, 77, 151) and bone morphogenetic proteins
(42, 59, 152, 164, 165), have been used in preclinical
and clinical trials for the treatment of large peri-
odontal or infrabony defects, as well as around dental
implants (36, 68, 110). The combined use of re-
combinant human platelet-derived growth factor-BB
and peptide P-15 with a graft biomaterial has shown
beneficial effects in intraosseous defects (157). How-
ever, contrasting results were reported for growth
factors such as platelet-rich plasma and graft combi-
nations, or the use of bioactive agents either alone or
in association with graft or guided tissue regeneration
for the treatment of furcation defects (157).
Biological effects of growth factors:
platelet-derived growth factor
Platelet-derived growth factor is a member of a
multifunctional polypeptide family that binds to two
cell-membrane tyrosine kinase receptors (platelet-
derived growth factor-Ra and platelet-derived growth
factor-Rb) and subsequently exerts its biological ef-
fects on cell proliferation, migration, extracellular
matrix synthesis and anti-apoptosis (56, 71, 138, 148).
Platelet-derived growth factor-a and -b receptors are
expressed in regenerating periodontal soft and hard
tissues (119). In addition, platelet-derived growth
factor initiates tooth-supporting periodontal liga-
ment cell chemotaxis (111), mitogenesis (113), matrix
synthesis (53) and attachment to tooth dentinal sur-
faces (172). More importantly, in vivo application of
platelet-derived growth factor alone or in combina-
Fig. 3. Periodontal regeneration of a three-wall infrabony
defect using Emdogain. (A) A 32-year-old male patient
(nonsmoker with severe periodontitis). Tooth 13 shows a
probing pocket depth of 10 mm disto-buccally and clinical
attachment loss of 14 mm. (B) Pretreatment radiograph
shows the infrabony defect distal to tooth 13. (C) After the
buccal incision of the papilla, the interdental tissue is
preserved attached to the palatal flap. After debridement
of the granulation tissue and the root surface, the in-
frabony defect is classified and measured: the predomi-
nant component is a 7-mm-deep three-wall defect. (D)
One year after surgical intervention the distal site of tooth
13 shows a probing pocket depth of 2 mm and clinical
attachment loss of 7 mm. Comparison with the initial
measurements indicates that a probing pocket depth gain
of 8 mm and a clinical attachment loss gain of 7 mm have
been achieved. (E) Radiograph 1 year postsurgery showing
filling of the defect.
Ramseier et al.
tion with insulin-like growth factor-1 results in the
partial repair of periodontal tissues (46, 47, 87, 88,
140). Platelet-derived growth factor has been shown
to have a significant regenerative impact on peri-
odontal ligament cells, as well as on osteoblasts (90,
92, 113, 115).
The clinical application of platelet-derived growth
factor was shown to successfully advance alveolar
bone repair and clinical attachment level gain. A first
clinical study reported the successful repair of class II
furcations using demineralized freeze-dried bone
allograft saturated with recombinant human platelet-
derived growth factor-BB (109). In a second study,
recombinant human platelet-derived growth factor-
BB mixed with a synthetic beta-tricalcium phosphate
matrix was shown to advance the repair of deep in-
frabony pockets in a large multicenter randomized
controlled trial (110). Both studies demonstrated that
the use of recombinant human platelet-derived
growth factor-BB was safe and effective in the treat-
ment of periodontal osseous defects. In a follow-up
trial, the same sample of patients was assessed 18 or
24 months following periodontal surgery. Substantial
radiographic changes in the appearance of the defect
fill were observed for patients treated with re-
combinant human platelet-derived growth factor-BB
Biological effects of growth factors:
bone morphogenetic proteins
Bone morphogenetic proteins are multifunctional
polypeptides belonging to the transforming growth
factor-beta superfamily of proteins (169). The human
genome encodes at least 20 bone morphogenetic
proteins (131). Bone morphogenetic proteins bind to
type I and type II receptors that function as serine-
threonine kinases. The type I receptor protein kinase
called Smads (the sma gene in Caenorhabditis elegans
and the Mad gene in Drosophila). The phosphory-
lated bone morphogenetic protein-signaling Smads
enter the nucleus and initiate the production of bone
matrix proteins, leading to bone morphogenesis. The
most remarkable feature of bone morphogenetic
proteins is their ability to induce ectopic bone for-
mation (160). Bone morphogenetic proteins are not
only powerful regulators of cartilage and bone for-
mation during embryonic development and regen-
eration in postnatal life, but they also participate in
the development and repair of other organs such as
the brain, kidney and nerves (132).
Sigurdsson et al.(149)
cementum formation following regenerative peri-
odontal surgery by the use of recombinant human
bone morphogenetic protein in surgically created
supra-alveolar defects in dogs (168). Histologic
analysis showed significantly more cementum for-
mation and regrowth of alveolar bone on bone
morphogenetic protein-treated sites compared with
Studies have demonstrated the expression of bone
morphogenetic proteins during tooth development
and periodontal repair, including alveolar bone (1, 2).
Investigations in animal models have shown the po-
tential repair of alveolar bony defects using re-
combinant human bone morphogenetic protein-12
(165) or recombinant human bone morphogenetic
protein-2 (86, 166). In a clinical trial by Fiorellini
et al. (36), recombinant human bone morphogenetic
protein-2, delivered by a bioabsorbable collagen
sponge, revealed significant bone formation in a
human buccal wall defect model following tooth
extraction when compared with collagen sponge
alone. Furthermore, bone morphogenetic protein-7,
Table 2. Effects of growth factors used for periodontal tissue engineering
Growth factor Effects
Platelet-derived growth factor Migration, proliferation and noncollagenous matrix synthesis of mesenchymal
Bone morphogenetic protein Proliferation, differentiation of osteoblasts and differentiation of periodontal lig-
ament cells into osteoblasts
Enamel matrix derivativeProliferation, protein synthesis and mineral nodule formation in periodontal lig-
ament cells, osteoblasts and cementoblasts
Transforming growth factor-betaProliferation of cementoblasts and periodontal ligament fibroblasts
Insulin-like growth factor-1Cell migration, proliferation, differentiation and matrix synthesis
Fibroblast growth factor-2 Proliferation and attachment of endothelial cells and periodontal ligament cells
Periodontal tissue-engineering technologies
also known as osteogenic protein-1, stimulates bone
regeneration around teeth, endosseous dental im-
plants and in maxillary sinus floor-augmentation
procedures (49, 141, 161).
Clinical application of growth factors for
use in periodontal regeneration
In general, the impact of topical delivery of growth
factors to periodontal wounds has been promising,
yet insufficient to promote predictable periodontal
tissue engineering (14, 23) (Fig. 4). Growth factor
proteins, once delivered to the target site, tend to
suffer from instability and quick dilution, presum-
ably because of proteolytic breakdown, receptor-
mediated endocytosis and solubility of the delivery
vehicle (3). Because their half-lives are significantly
reduced, the period of exposure may not be suf-
ficient to act on osteoblasts, cementoblasts or
methods of growth-factor delivery need to be
Investigations for periodontal bioengineering have
examined a variety of methods that combine delivery
vehicles, such as scaffolds, with growth factors to
target the defect site in order to optimize bioavail-
ability (85). The scaffolds are designed to optimize
the dosage of the growth factor and to control its
Fig. 4. Periodontal regeneration using platelet-derived
growth factor and bone-graft materials. (A) A 27-year-old
patient at the re-evaluation visit after the initial nonsur-
gical therapy; three sites with a probing pocket depth of
>6 mm were identified. One of those sites, distal to tooth
44, shows a probing pocket depth of 7 mm and no gingival
recession. (B) The periapical radiograph shows a deep,
one-wall defect distal to tooth 44 and a lesion between
teeth 45 and 46. (C) Measurement of the one-wall defect
shows an infrabony component of 6 mm. (D) The grafting
material (GEM 21S?) is mixed with particles of autoge-
nous bone chips collected in the surgical area with a
Rhodes instrument and with the liquid component of the
GEM 21S?(platelet-derived growth factor). (E) The liquid
platelet-derived growth factor is placed in the defect
together with the graft to rebuild the lost bone. (F) A
second internal mattress suture is performed with a 7-0
Gore-Tex?suture, to allow for optimal adaptation of the
flap margin without the interference of the epithelium.
The two internal mattress sutures are tied and the knots
are performed only after a perfect free-tension closure of
the wound. Two additional interrupted 7-0 sutures are
placed to ensure stable contact between the connective
tissues of the edges of the flaps. The mesial and distal
papillae are stabilized with additional simple interrupted
sutures. (G) Nine months after surgery, the probing
pocket depth is 2 mm. (H) Nine months after surgery, the
periapical radiograph shows good bone fill of the one-
wall bony defect. (I) Nine months after surgery, the sur-
gical re-entry shows new bone formation.
Ramseier et al.
release pattern, which may be pulsatile, constant or
time-programmed (8). The kinetics of the release and
the duration of the exposure of the growth factor may
also be controlled (61).
A new polymeric system, permitting the tissue-
specific delivery (at a controlled dose and delivery
rate) of two ormore growth factors, was reported in an
animal study carried out by Richardson et al. (137).
The dual delivery of vascular endothelial growth fac-
tor with platelet-derived growth factor from a single,
structural polymer scaffold results in the rapid for-
mation of a mature vascular network (137).
Guided tissue regeneration
Histological findings from periodontal-regeneration
studies reveal that a new connective tissue attach-
ment could be predicted if the cells from the peri-
odontal ligament settle on the root surface during
healing. Hence, the clinical applications of guided
placement of a physical barrier membrane to enable
the previous periodontitis-affected tooth root surface
to be repopulated with cells from the periodontal
ligament. In the last few decades, guided tissue
regeneration has been applied in many clinical trials
for the treatment of various periodontal defects, such
as infrabony defects (25), furcation involvement (72,
89) and localized gingival recession (121). In a recent
membranes and grafting materials used in preclinical
models have been summarized. The analysis of 10
papers revealed that the combination of barrier
membranes and grafting materials may result in
histological evidence of periodontal regeneration,
predominantly bone repair. No additional histologi-
cal benefits of combination treatments were found in
animal models of three-wall intrabony, class II fur-
cation, or fenestration defects. In supra-alveolar and
two-wall intrabony defect models of periodontal
regeneration, the additional use of a grafting material
gave superior histological results of bone repair
compared with the use of barrier membranes alone
The types of barrier membranes evaluated in clin-
ical studies vary in design, configuration and com-
position. Nonresorbable membranes of expanded
polytetrafluoroethylene have been used successfully
in both animal experiments and human clinical trials.
In recent years, natural or synthetic bio-absorbable
barrier membranes have been used for guided tissue
regeneration in order to eliminate the need for fol-
low-up surgery for membrane removal. Collagen
membranes, as well as barrier materials of polylactic
acid, or copolymers of polylactic acid and poly-
glycolic acid, have been tested in animal and human
Following therapy, guided tissue regeneration has a
greater effect on the probing measures of periodontal
including increased attachment gain, reduction of
probing depth, less gingival recession and more gain
in hard-tissue probing at surgical re-entry. Referring
to the best evidence currently available, however, it is
difficult to draw general conclusions about the
clinical benefit of guided tissue regeneration. Al-
though there is evidence demonstrating that guided
tissue regeneration has significant benefits over
conventional open-flap surgery, the factors affecting
outcomes are unclear from the present literature
because they might be influenced by study conduct
issues, such as bias (106).
procedure used to achieve periodontal regeneration
in infrabony defects and in class II furcations. Further
benefit may be achieved by the additional use of
grafting materials (155).
Gene therapeutics for periodontal
Although encouraging results for periodontal regen-
eration have been found in various clinical investi-
gations using recombinant tissue growth factors,
there are limitations for topical protein delivery, such
as transient biological activity, protease inactivation
and poor bioavailability from existing delivery vehi-
cles. Therefore, newer approaches seek to develop
methodologies that optimize growth-factor targeting
to maximize the therapeutic outcome of periodontal-
periodontal tissue engineering show early progress in
achieving delivery of growth-factor genes, such as
platelet-derived growth factor or bone morphogenetic
protein, to periodontal lesions (Fig. 5). Gene-transfer
methods may circumvent many of the limitations
with protein delivery to soft-tissue wounds (10, 45). It
has been shown that the application of growth factors
(37, 63, 64, 78) or soluble forms of cytokine receptors
(21) by gene transfer provides greater sustainability
than the application of a single protein. Thus, gene
therapy may achieve greater bioavailability of growth
factors within periodontal wounds and hence provide
greater regenerative potential.
Periodontal tissue-engineering technologies
Methods for gene delivery in periodontal
Various gene-delivery methods are available to
administer growth factors to periodontal defects,
offering great flexibility for tissue engineering. The
delivery method can be tailored to the specific
characteristics of the wound site. For example, a
horizontal one- or two-walled defect may require the
use of a supportive carrier, such as a scaffold. Other
defect sites may be conducive to the use of an ade-
novirus vector embedded in a collagen matrix.
More importantly from a clinical point of view is
the risk associated with the use of gene therapy in
periodontal tissue engineering (51). As with maxi-
mizing growth-factor sustainability and accounting
for specific characteristics of the wound site, both the
DNA vector and delivery method need to be consid-
ered when assessing patient safety. In summary,
studies examining the use of specific delivery meth-
ods and DNA vectors in periodontal tissue engi-
neering aim to maximize the duration of growth
factor expression, optimize the method of delivery to
the periodontal defect and minimize patient risk.
A combination of an Adeno-Associated Virus-
delivered angiogenic molecule, such as vascular
endothelial growth factor, bone morphogenetic pro-
tein signaling receptor (caALK2) and receptor acti-
vator of nuclear factor-kappa B ligand, was demon-
strated to promote bone allograft turnover and
osteogenesis as a mode to enrich human bone allo-
grafts (62). To date, combinations of vascular endo-
thelial growth factor ⁄ bone morphogenetic protein
(120) and platelet-derived growth factor ⁄ vascular
endothelial growth factor (137) have had highly po-
sitive synergistic responses in bone repair.
Promising preliminary results from preclinical stud-
ies reveal that host modulation achieved through gene
delivery of soluble proteins, such as tumor necrosis
factor receptor 1 (TNFR1:Fc), reduces tumor necrosis
factor activity and therefore inhibits alveolar bone loss
(21). These results are comparable to the findings in the
research on rheumatoid arthritis where pathogenesis
includes high tumor necrosis factor activity and the
pathways for bone resorption are similar (127).
Preclinical studies evaluating growth
factor gene therapy for periodontal tissue
In order to overcome the short half-lives of growth
factor peptides in vivo, gene therapy using a vector
encoding the growth factor is advocated to stimulate
tissue regeneration. So far, two main strategies of
gene vector delivery have been applied to peri-
odontal tissue engineering. Gene vectors can be
introduced directly to the target site (in vivo tech-
nique) (63) or selected cells can be harvested, ex-
Fig. 5. Advanced approaches for re-
generating tooth-supporting struc-
tures. (A) Application of a graft
material (e.g. bone ceramic) and
growth factor into an infrabony de-
membrane. (B) Application of gene
Ramseier et al.
panded, genetically transduced and then re-im-
planted (ex vivo technique) (64). In vivo gene
transfer involves the insertion of the gene of interest
directly into the body anticipating the genetic
modification of the target cell. Ex vivo gene transfer
includes the incorporation of genetic material into
cells exposed from a tissue biopsy with subsequent
re-implantation into the recipient. Using the in vivo
technique, the potential inhibition of alveolar bone
loss has been studied in an experimental periodon-
titis model evaluating the inhibition of osteoclasto-
genesis by administering human osteoprotegerin, a
competitive inhibitor of the receptor activator of
nuclear factor-kappa B ligand-derived osteoclast
activation. Significant preservation of alveolar bone
volume was observed among osteoprotegerin:Fc-
treated animals compared with controls. Systemic
delivery of osteoprotegerin:Fc inhibits alveolar bone
resorption in experimental periodontitis, suggesting
that inhibition of receptor activator of nuclear fac-
tor-kappa B ligand may represent an important
therapeutic strategy for the prevention of progres-
sive alveolar bone loss (65).
Platelet-derived growth factor gene
Platelet-derived growth factor-gene transfer strate-
gies were originally used in tissue engineering to
improve healing in soft-tissue wounds such as skin
lesions (27). Both plasmid (57) and adenovirus ⁄
platelet-derived growth factor (125) gene delivery
have been evaluated in preclinical and human trials.
However, the latter exhibits greater safety in clinical
use (51). In a recent animal study reporting on safety
and distribution profiles, adenovirus ⁄ platelet-de-
rived growth factor-B applied for tissue engineering
of tooth-supporting alveolar bone defects was well
contained within the localized osseous defect area
without viremia or distant organ involvement (18).
Early studies in dental applications using re-
combinant adenoviral vectors encoding platelet-de-
rived growth factor demonstrated the ability of these
vector constructs to potently transduce cells isolated
from the periodontium (osteoblasts, cementoblasts,
periodontal ligament cells and gingival fibroblasts)
(48, 173). These studies revealed the extensive and
prolonged transduction of periodontal-derived cells.
Both Chen & Giannobile (19) and Lin et al. (81) were
able to demonstrate the effects of adenoviral delivery
of platelet-derived growth factor to understand, in
greater detail, sustained platelet-derived growth fac-
growth factor-B generally displays higher sustained
signal-transduction effects in human gingival fibro-
blasts compared to cells treated with recombinant
human platelet-derived growth factor-BB protein
alone. Their data on platelet-derived growth factor
gene deliverymay contribute to an improved
understanding of the pathways that are likely to play
a role in the control of clinical outcomes of peri-
In an ex vivo investigation by Anusaksathien et al.
(6), it was shown that the expression of platelet-de-
rived growth factor genes was prolonged for up to
10 days in gingival wounds. Adenovirus encoding
platelet-derived growth factor-B (adenovirus ⁄ plate-
let-derived growth factor-B) transduced gingival
fibroblasts and enhanced defect fill by inducing
human gingival fibroblast migration and proliferation
(6). On the other hand, continuous exposure of
cementoblasts to platelet-derived growth factor-A
had an inhibitory effect on cementum mineraliza-
tion, possibly via the upregulation of osteopontin and
the subsequent enhancement of multinucleated giant
cells in cementum-engineered scaffolds. Moreover,
adenovirus ⁄ platelet-derived growth factor-1308 (a
dominant-negative mutant of platelet-derived growth
factor) inhibited mineralization of tissue-engineered
cementum, possibly owing to the downregulation of
bone sialoprotein and osteocalcin and the persis-
tence of stimulation with multinucleated giant cells.
These findings suggest that continuous exogenous
delivery of platelet-derived growth factor-A may de-
lay mineral formation induced by cementoblasts,
while platelet-derived growth factor is clearly re-
quired for mineral neogenesis (5).
Jin et al. (63) demonstrated that direct in vivo gene
transfer of platelet-derived growth factor-B was able to
stimulate tissue regeneration in large periodontal de-
fects. Descriptive histology and histomorphometry
revealed that delivery of the human platelet-derived
growth factor-B gene promotes the regeneration of
both cementum and alveolar bone, while delivery of
platelet-derived growth factor-1308, a dominant-neg-
ative mutant of platelet-derived growth factor-A, has
minimal effects on periodontal tissue regeneration.
signaling. Genedelivery of platelet-derived
Delivery of the bone
morphogenetic protein gene
An experimental study in rodents by Lieberman et
al. (81) advanced gene therapy for bone regenera-
tion, with the results revealing that the transduction
Periodontal tissue-engineering technologies
of bone marrow stromal cells with recombinant
human bone morphogenetic protein 2 led to bone
formation within an experimental defect comparable
to skeletal bone. Another group was similarly able to
regenerate skeletal bone by directly administering
adenovirus5 ⁄ bone morphogenetic protein 2 into a
bony segmental defect in rabbits (9). Further ad-
vances in the area of orthopedic gene therapy using
viral delivery of bone morphogenetic protein 2 have
provided further evidence for the ability of both in
vivo and ex vivo bone engineering (20, 79, 80, 103).
Franceschi et al. (37) investigated in vitro and in vivo
adenovirus gene transfer of bone morphogenetic
protein 7 for bone formation. Adenovirus-trans-
duced nonosteogenic cells were also found to dif-
ferentiate into bone-forming cells and to produce
bone morphogenetic protein 7 (78) or bone mor-
phogenetic protein 2 (20) both in vitro and in vivo.
In another study by Huang et al. (60), plasmid DNA
encoding bone morphogenetic protein 4 adminis-
tered using a scaffold-delivery system was found to
enhance bone formation when compared with blank
In an early approach to regenerate alveolar bone in
an animal model, it was demonstrated that the
ex vivo delivery of an adenovirus encoding murine
bone morphogenetic protein 7 was found to promote
periodontal tissue regeneration in large mandibular
periodontal bone defects (64). Transfer of the bone
morphogenetic protein 7 gene enhanced alveolar
bone repair and also stimulated cementogenesis and
periodontal ligament fiber formation. Of interest,
alveolar bone formation was found to occur via a
cartilage intermediate. However, when genes encod-
ing the bone morphogenetic protein antagonist
noggin were delivered, inhibition of periodontal tis-
sue formation resulted (66). In a study by Dunn et al.
(30), it was shown that direct in vivo gene delivery of
adenovirus ⁄ bone morphogenetic protein 7 in a col-
lagen gel carrier promoted successful regeneration of
alveolar bone defects around dental implants. Fur-
thermore, an in vivo synergism was found of aden-
oviral-mediated coexpression of bone morphogenetic
protein 7 and insulin like growth factor 1 on human
periodontal ligament cells in up-regulating alkaline
phosphatase activity and the mRNA levels of collagen
type I and Runx2 (170). Implantation with scaffolds
illustrated that the transduced cells exhibited osteo-
genic differentiation and formed bone-like struc-
tures. It was concluded that the combined delivery of
bone morphogenetic protein 7 and insulin like
growth factor 1 genes using an internal ribosome
entry site-based strategy synergistically enhanced the
differentiation of human periodontal ligament cells
These experiments provide promising evidence
showing the feasibility of both in vivo and ex vivo
gene therapy for periodontal tissue regeneration and
Future perspectives: targeted gene
therapy in vivo
Major advances have been made over the past decade
in the reconstruction of complex periodontal and
alveolar bone wounds that have resulted from disease
or injury. Developments in scaffolding matrices for
cell, protein and gene delivery have demonstrated
significant potential to provide ?smart? biomaterials
that can interact with the matrix, cells and bioactive
factors.Thetargeting ofsignaling moleculesorgrowth
factors (via proteins or genes) to periodontal tissue
components has led to significant new knowledge
generation using factors that promote cell replication,
differentiation, matrix biosynthesis and angiogenesis.
A major challenge that has been studied less is the
modulation of the exuberant host response to micro-
bial contamination that plagues the periodontal
wound microenvironment. To achieve improvements
in the outcome of periodontal-regenerative medicine,
scientists will need to examine the dual delivery of
host modifiers or anti-infective agents to optimize the
results of therapy. Further advancements in the field
will continue to rely heavily on multidisciplinary ap-
and infectious disease specialists in repairing the
complex periodontal wound environment.
This work was supported by NIH ⁄ NIDCR DE13397
and NIH ⁄ NCRR UL1RR-024986. The authors thank
Mr Chris Jung for his assistance with the figures.
1. Aberg T, Wozney J, Thesleff I. Expression patterns of bone
morphogenetic proteins (BMPs) in the developing mouse
tooth suggest roles in morphogenesis and cell differenti-
ation. Dev Dyn 1997: 210: 383–396.
2. Amar S, Chung KM, Nam SH, Karatzas S, Myokai F, Van
Dyke TE. Markers of bone and cementum formation
accumulate in tissues regenerated in periodontal defects
treated with expanded polytetrafluoroethylene mem-
branes. J Periodontal Res 1997: 32: 148–158.
Ramseier et al.
3. Anusaksathien O, Giannobile WV. Growth factor delivery
to re-engineer periodontal tissues. Curr Pharm Biotechnol
2002: 3: 129–139.
4. Anusaksathien O, Jin Q, Ma PX, Giannobile WV. Scaf-
folding in periodontal engineering. In: Ma PX, Eliseeff J,
editors. Scaffolding in tissue engineering. Boca Raton, FL,
USA: CRC Press, 2005: 427–444.
5. Anusaksathien O, Jin Q, Zhao M, Somerman MJ, Gianno-
bile WV. Effect of sustained gene delivery of platelet-
derived growth factor or its antagonist (PDGF-1308) on
tissue-engineered cementum. J Periodontol 2004: 75: 429–
6. Anusaksathien O, Webb SA, Jin QM, Giannobile WV.
Platelet-derived growth factor gene delivery stimulates ex
vivo gingival repair. Tissue Eng 2003: 9: 745–756.
7. Axelsson P, Lindhe J. The significance of maintenance care
in the treatment of periodontal disease. J Clin Periodontol
1981: 8: 281–294.
8. Babensee JE, McIntire LV, Mikos AG. Growth factor deliv-
ery for tissue engineering. Pharm Res 2000: 17: 497–504.
9. Baltzer AW, Lattermann C, Whalen JD, Wooley P, Weiss K,
Grimm M, Ghivizzani SC, Robbins PD, Evans CH. Genetic
enhancement of fracture repair: healing of an experi-
mental segmental defect by adenoviral transfer of the
BMP-2 gene. Gene Ther 2000: 7: 734–739.
10. Baum BJ, Goldsmith CM, Kok MR, Lodde BM, van Mello
NM, Voutetakis A, Wang J, Yamano S, Zheng C. Advances
in vector-mediated gene transfer. Immunol Lett 2003: 90:
11. Bjorn H, Hollender L, Lindhe J. Tissue regeneration in
patients with periodontal disease. Odontol Revy 1965: 16:
12. Briggs SL. The role of fibronectin in fibroblast migration
during tissue repair. J Wound Care 2005: 14: 284–287.
13. Camelo M, Nevins ML, Lynch SE, Schenk RK, Simion
M, Nevins M. Periodontal regeneration with an autog-
enous bone-Bio-Oss composite graft and a Bio-Gide
membrane. Int J Periodontics Restorative Dent 2001: 21:
14. Camelo M, Nevins ML, Schenk RK, Lynch SE, Nevins M.
Periodontal regeneration in human Class II furcations
growth factor-BB (rhPDGF-BB) with bone allograft. Int J
Periodontics Restorative Dent 2003: 23: 213–225.
15. Camelo M, Nevins ML, Schenk RK, Simion M, Rasperini G,
Lynch SE, Nevins M. Clinical, radiographic, and histologic
evaluation of human periodontal defects treated with Bio-
Oss and Bio-Gide. Int J Periodontics Restorative Dent 1998:
16. Caton J, Nyman S, Zander H. Histometric evaluation of
periodontal surgery. II. Connective tissue attachment
levels after four regenerative procedures. J Clin Period-
ontol 1980: 7: 224–231.
17. Caton J, Zander HA. Osseous repair of an infrabony pocket
without new attachment of connective tissue. J Clin Peri-
odontol 1976: 3: 54–58.
18. Chang PC, Cirelli JA, Jin Q, Seol YJ, Sugai JV, D?Silva NJ,
Danciu TE, Chandler LA, Sosnowski BA, Giannobile WV.
Adenovirus encoding human platelet-derived growth fac-
tor-B delivered to alveolar bone defects exhibits safety and
biodistribution profiles favorable for clinical use. Hum
Gene Ther 2009: 20: 486–496.
19. Chen QP, Giannobile WV. Adenoviral gene transfer of
PDGF downregulates gas gene product PDGFalphaR and
prolongs ERK and Akt ⁄ PKB activation. Am J Physiol Cell
Physiol 2002: 282: C538–C544.
20. Cheng SL, Lou J, Wright NM, Lai CF, Avioli LV, Riew KD. In
vitro and in vivo induction of bone formation using a re-
combinant adenoviral vector carrying the human BMP-2
gene. Calcif Tissue Int 2001: 68: 87–94.
21. Cirelli JA, Park CH, MacKool K, Taba M Jr, Lustig KH,
Burstein H, Giannobile WV. AAV2 ⁄ 1-TNFR:Fc gene de-
livery prevents periodontal disease progression. Gene Ther
2009: 16: 426–436.
22. Clark RA. Biology of dermal wound repair. Dermatol Clin
1993: 11: 647–666.
23. Cochran DL, Wozney JM. Biological mediators for peri-
odontal regeneration. Periodontol 2000 1999: 19: 40–58.
24. Cortellini P. Reconstructive periodontal surgery: a chal-
lenge for modern periodontology. Int Dent J 2006: 56: 250–
25. Cortellini P, Bowers GM. Periodontal regeneration of in-
trabony defects: an evidence-based treatment approach.
Int J Periodontics Restorative Dent 1995: 15: 128–145.
26. Cortellini P, Tonetti MS. Clinical performance of a regen-
erative strategy for intrabony defects: scientific evidence
and clinical experience. J Periodontol 2005: 76: 341–350.
27. Crombleholme TM. Adenoviral-mediated gene transfer in
wound healing. Wound Repair Regen 2000: 8: 460–472.
28. Dereka XE, Markopoulou CE, Vrotsos IA. Role of growth
factors on periodontal repair. Growth Factors 2006: 24:
29. Dryankova MM, Popova CL. Regenerative therapy of fur-
cation defect. Folia Med (Plovdiv) 2001: 43: 64–68.
30. Dunn CA, Jin Q, Taba M Jr, Franceschi RT, Bruce Ruth-
erford R, Giannobile WV. BMP gene delivery for alveolar
bone engineering at dental implant defects. Mol Ther
2005: 11: 294–299.
31. Ellegaard B, Loe H. New attachment of periodontal tissues
after treatment of intrabony lesions. J Periodontol 1971:
32. Esposito M, Coulthard P, Thomsen P, Worthington HV.
Enamel matrix derivative for periodontal tissue regenera-
tion in treatment of intrabony defects: a Cochrane sys-
tematic review. J Dent Educ 2004: 68: 834–844.
33. Esposito M, Coulthard P, Worthington HV. Enamel matrix
derivative (Emdogain) for periodontal tissue regeneration
in intrabony defects. Cochrane Database Syst Rev 2003: 2:
34. Esposito M, Grusovin MG, Coulthard P, Worthington HV.
Enamel matrix derivative (Emdogain) for periodontal tis-
sue regeneration in intrabony defects. Cochrane Database
Syst Rev 2005: 4: CD003875.
35. Fetner AE, Hartigan MS, Low SB. Periodontal repair using
PerioGlas in nonhuman primates: clinical and histologic
observations. Compendium 1994: 938: 932, 935–938; quiz
36. Fiorellini JP, Howell TH, Cochran D, Malmquist J, Lilly LC,
Spagnoli D, Toljanic J, Jones A, Nevins M. Randomized
study evaluating recombinant human bone morphoge-
netic protein-2 for extraction socket augmentation. J Pe-
riodontol 2005: 76: 605–613.
37. Franceschi RT, Wang D, Krebsbach PH, Rutherford RB.
Gene therapy for bone formation: in vitro and in vivo
Periodontal tissue-engineering technologies
osteogenic activity of an adenovirus expressing BMP7.
J Cell Biochem 2000: 78: 476–486.
38. Francetti L, Del Fabbro M, Basso M, Testori T, Weinstein
R. Enamel matrix proteins in the treatment of intra-bony
defects. A prospective 24-month clinical trial. J Clin Peri-
odontol 2004: 31: 52–59.
39. Froum SJ, Thaler R, Scopp IW, Stahl SS. Osseous auto-
grafts. I. Clinical responses to bone blend or hip marrow
grafts. J Periodontol 1975: 46: 515–521.
40. Fuentes P, Garrett S, Nilveus R, Egelberg J. Treatment of
periodontal furcation defects. Coronally positioned flap
with or without citric acid root conditioning in class II
defects. J Clin Periodontol 1993: 20: 425–430.
41. Galgut PN, Waite IM, Brookshaw JD, Kingston CP. A 4-year
controlled clinical study into the use of a ceramic
hydroxylapatite implant material for the treatment of
periodontal bone defects. J Clin Periodontol 1992: 19: 570–
42. Gao Y, Yang L, Fang YR, Mori M, Kawahara K, Tanaka A.
The inductive effect of bone morphogenetic protein
(BMP) on human periodontal fibroblast-like cells in vitro.
J Osaka Dent Univ 1995: 29: 9–17.
43. Gestrelius S, Lyngstadaas SP, Hammarstrom L. Emdogain
– periodontal regeneration based on biomimicry. Clin
Oral Investig 2000: 4: 120–125.
44. Giannobile WV. Periodontal tissue engineering by growth
factors. Bone 1996: 19: 23S–37S.
45. Giannobile WV. What does the future hold for periodontal
tissue engineering? Int J Periodontics Restorative Dent
2002: 22: 6–7.
46. Giannobile WV, Finkelman RD, Lynch SE. Comparison of
canine and non-human primate animal models for peri-
odontal regenerative therapy: results following a single
administration of PDGF ⁄ IGF-I. J Periodontol 1994: 65:
47. Giannobile WV, Hernandez RA, Finkelman RD, Ryan S,
Kiritsy CP, D?Andrea M, Lynch SE. Comparative effects of
platelet-derived growth factor-BB and insulin-like growth
factor-I, individually and in combination, on periodontal
regeneration in Macaca fascicularis. J Periodontal Res
1996: 31: 301–312.
48. Giannobile WV, Lee CS, Tomala MP, Tejeda KM, Zhu Z.
Platelet-derived growth factor (PDGF) gene delivery for
application in periodontal tissue engineering. J Periodon-
tol 2001: 72: 815–823.
49. Giannobile WV, Ryan S, Shih MS, Su DL, Kaplan PL, Chan
TC. Recombinant human osteogenic protein-1 (OP-1)
stimulates periodontal wound healing in class III furcation
defects. J Periodontol 1998: 69: 129–137.
50. Goldman H, Cohen W. The infrabony pocket: classificas-
sion and treatment. J Periodontol 1958: 29: 272–291.
51. Gu DL, Nguyen T, Gonzalez AM, Printz MA, Pierce GF,
Sosnowski BA, Phillips ML, Chandler LA. Adenovirus
encoding human platelet-derived growth factor-B deliv-
ered in collagen exhibits safety, biodistribution, and
immunogenicity profiles favorable for clinical use. Mol
Ther 2004: 9: 699–711.
52. Gurinsky BS, Mills MP, Mellonig JT. Clinical evaluation of
demineralized freeze-dried bone allograft and enamel
matrix derivative versus enamel matrix derivative alone for
the treatment of periodontal osseous defects in humans.
J Periodontol 2004: 75: 1309–1318.
53. Haase HR, Clarkson RW, Waters MJ, Bartold PM. Growth
factor modulation of mitogenic responses and proteogly-
can synthesis by human periodontal fibroblasts. J Cell
Physiol 1998: 174: 353–361.
54. Hammarstrom L. Enamel matrix, cementum development
and regeneration. J Clin Periodontol 1997: 24: 658–668.
55. Hartman GA, Arnold RM, Mills MP, Cochran DL, Mellonig
JT. Clinical and histologic evaluation of anorganic bovine
bone collagen with or without a collagen barrier. Int J
Periodontics Restorative Dent 2004: 24: 127–135.
56. Heldin P, Laurent TC, Heldin CH. Effect of growth factors
on hyaluronan synthesis in cultured human fibroblasts.
Biochem J 1989: 258: 919–922.
57. Hijjawi J, Mogford JE, Chandler LA, Cross KJ, Said H, So-
snowski BA, Mustoe TA. Platelet-derived growth factor B,
but not fibroblast growth factor 2, plasmid DNA improves
survival of ischemic myocutaneous flaps. Arch Surg 2004:
58. Howell TH, Fiorellini JP, Paquette DW, Offenbacher S,
Giannobile WV, Lynch SE. A phase I ⁄ II clinical trial to
evaluate a combination of recombinant human platelet-
insulin-like growth factor-I in patients with periodontal
disease. J Periodontol 1997: 68: 1186–1193.
59. Huang KK, Shen C, Chiang CY, Hsieh YD, Fu E. Effects of
bone morphogenetic protein-6 on periodontal wound
healing in a fenestration defect of rats. J Periodontal Res
2005: 40: 1–10.
60. Huang YC, Simmons C, Kaigler D, Rice KG, Mooney DJ.
Bone regeneration in a rat cranial defect with delivery of
PEI-condensed plasmid DNA encoding for bone morpho-
genetic protein-4 (BMP-4). Gene Ther 2005: 12: 418–426.
61. Hutmacher DW, Teoh SH, Zein I, Ranawake M, Lau S.
Tissue engineering research: the engineer?s role. Med
Device Technol 2000: 11: 33–39.
62. Ito H, Koefoed M, Tiyapatanaputi P, Gromov K, Goater JJ,
Carmouche J, Zhang X, Rubery PT, Rabinowitz J, Samulski
RJ, Nakamura T, Soballe K, O?Keefe RJ, Boyce BF, Schwarz
EM. Remodeling of cortical bone allografts mediated by
adherent rAAV-RANKL and VEGF gene therapy. Nat Med
2005: 11: 291–297.
63. Jin Q, Anusaksathien O, Webb SA, Printz MA, Giannobile
WV. Engineering of tooth-supporting structures by deliv-
ery of PDGF gene therapy vectors. Mol Ther 2004: 9: 519–
64. Jin QM, Anusaksathien O, Webb SA, Rutherford RB, Gi-
annobile WV. Gene therapy of bone morphogenetic pro-
tein for periodontal tissue engineering. J Periodontol 2003:
65. Jin Q, Cirelli JA, Park CH, Sugai JV, Taba M, Kostenuik PJ,
Giannobile WV. RANKL inhibition through osteoproteg-
erin blocks bone loss in experimental periodontitis.
J Periodontol 2007: 78: 1300–1308.
66. Jin QM, Zhao M, Economides AN, Somerman MJ, Gian-
nobile WV. Noggin gene delivery inhibits cementoblast-
induced mineralization. Connect Tissue Res 2004: 45:
67. Judith R, Nithya M, Rose C, Mandal AB. Application of a
PDGF-containing novel gel for cutaneous wound healing.
Life Sci 2010: 87: 1–8.
68. Jung RE, Glauser R, Scharer P, Hammerle CH, Sailer HF,
Weber FE. Effect of rhBMP-2 on guided bone regener-
Ramseier et al.
ation in humans. Clin Oral Implants Res 2003: 14: 556–
69. Jung RE, Zwahlen R, Weber FE, Molenberg A, van Lenthe
GH, Hammerle CH. Evaluation of an in situ formed syn-
thetic hydrogel as a biodegradable membrane for guided
bone regeneration. Clin Oral Implants Res 2006: 17: 426–
70. Kaigler D, Cirelli JA, Giannobile WV. Growth factor deliv-
ery for oral and periodontal tissue engineering. Expert
Opin Drug Deliv 2006: 3: 647–662.
71. Kaplan DR, Chao FC, Stiles CD, Antoniades HN, Scher CD.
Platelet alpha granules contain a growth factor for fibro-
blasts. Blood 1979: 53: 1043–1052.
72. Karring T, Cortellini P. Regenerative therapy: furcation
defects. Periodontol 2000 1999: 19: 115–137.
73. Karring T, Isidor F, Nyman S, Lindhe J. New attachment
formation on teeth with a reduced but healthy periodontal
ligament. J Clin Periodontol 1985: 12: 51–60.
74. Karring T, Nyman S, Lindhe J. Healing following implan-
tation of periodontitis affected roots into bone tissue.
J Clin Periodontol 1980: 7: 96–105.
75. Karring T, Nyman S, Lindhe J, Sirirat M. Potentials for root
resorption during periodontal wound healing. J Clin Pe-
riodontol 1984: 11: 41–52.
76. Kimble KM, Eber RM, Soehren S, Shyr Y, Wang HL.
Treatment of gingival recession using a collagen mem-
brane with or without the use of demineralized freeze-
dried bone allograft for space maintenance. J Periodontol
2004: 75: 210–220.
77. Kitamura M, Akamatsu M, Machigashira M, Hara Y, Sa-
kagami R, Hirofuji T, Hamachi T, Maeda K, Yokota M, Kido
J, Nagata T, Kurihara H, Takashiba S, Sibutani T, Fukuda
M, Noguchi T, Yamazaki K, Yoshie H, Ioroi K, Arai T,
Nakagawa T, Ito K, Oda S, Izumi Y, Ogata Y, Yamada S,
Shimauchi H, Kunimatsu K, Kawanami M, Fujii T, Fur-
uichi Y, Furuuchi T, Sasano T, Imai E, Omae M, Yamada S,
Watanuki M, Murakami S. FGF-2 stimulates periodontal
regeneration: results of a multi-center randomized clinical
trial. J Dent Res 2011: 90: 35–40.
78. Krebsbach PH, Gu K, Franceschi RT, Rutherford RB. Gene
therapy-directed osteogenesis: BMP-7-transduced human
fibroblasts form bone in vivo. Hum Gene Ther 2000: 11:
79. Lee JY, Musgrave D, Pelinkovic D, Fukushima K, Cummins
J, Usas A, Robbins P, Fu FH, Huard J. Effect of bone
morphogenetic protein-2-expressing muscle-derived cells
on healing of critical-sized bone defects in mice. J Bone
Joint Surg Am 2001: 83-A: 1032–1039.
80. Lee JY, Peng H, Usas A, Musgrave D, Cummins J, Pe-
linkovic D, Jankowski R, Ziran B, Robbins P, Huard J.
Enhancement of bone healing based on ex vivo gene
therapy using human muscle-derived cells expressing
bone morphogenetic protein 2. Hum Gene Ther 2002: 13:
81. Lieberman JR, Daluiski A, Stevenson S, Wu L, McAllister P,
Lee YP, Kabo JM, Finerman GA, Berk AJ, Witte ON. The
effect of regional gene therapy with bone morphogenetic
protein-2-producing bone-marrow cells on the repair of
segmental femoral defects in rats. J Bone Joint Surg Am
1999: 81: 905–917.
82. Lin Z, Sugai JV, Jin Q, Chandler LA, Giannobile WV.
Platelet-derived growth factor-B gene delivery sustains
gingival fibroblast signal transduction. J Periodontal Res
2008: 43: 440–449.
83. Lioubavina-Hack N, Karring T, Lynch SE, Lindhe J. Methyl
cellulose gel obstructed bone formation by GBR: an
experimental study in rats. J Clin Periodontol 2005: 32:
84. Listgarten MA, Rosenberg MM. Histological study of repair
following new attachment procedures in human peri-
odontal lesions. J Periodontol 1979: 50: 333–344.
85. Lutolf MP, Hubbell JA. Synthetic biomaterials as instruc-
tive extracellular microenvironments for morphogenesis
in tissue engineering. Nat Biotechnol 2005: 23: 47–55.
86. Lutolf MP, Weber FE, Schmoekel HG, Schense JC, Kohler
T, Muller R, Hubbell JA. Repair of bone defects using
synthetic mimetics of collagenous extracellular matrices.
Nat Biotechnol 2003: 21: 513–518.
87. Lynch SE, de Castilla GR, Williams RC, Kiritsy CP, Howell
TH, Reddy MS, Antoniades HN. The effects of short-term
application of a combination of platelet-derived and
insulin-like growth factors on periodontal wound healing.
J Periodontol 1991: 62: 458–467.
88. Lynch SE, Williams RC, Polson AM, Howell TH, Reddy
MS, Zappa UE, Antoniades HN. A combination of
platelet-derived and insulin-like growth factors enhances
periodontal regeneration. J Clin Periodontol 1989: 16:
89. Machtei EE, Schallhorn RG. Successful regeneration of
mandibular Class II furcation defects: an evidence-based
treatment approach. Int J Periodontics Restorative Dent
1995: 15: 146–167.
90. Marcopoulou CE, Vavouraki HN, Dereka XE, Vrotsos IA.
Proliferative effect of growth factors TGF-beta1, PDGF-BB
and rhBMP-2 on human gingival fibroblasts and peri-
odontal ligament cells. J Int Acad Periodontol 2003: 5: 63–
91. Mariotti A. Efficacy of chemical root surface modifiers in
the treatment of periodontal disease. A systematic review.
Ann Periodontol 2003: 8: 205–226.
92. Matsuda N, Lin WL, Kumar NM, Cho MI, Genco RJ.
Mitogenic, chemotactic, and synthetic responses of rat
periodontal ligament fibroblastic cells to polypeptide
growth factors in vitro. J Periodontol 1992: 63: 515–525.
93. McCulloch CA. Basic considerations in periodontal wound
healing to achieve regeneration. Periodontol 2000 1993: 1:
94. McGuire MK, Kao RT, Nevins M, Lynch SE. rhPDGF-BB
promotes healing of periodontal defects: 24-month clini-
cal and radiographic observations. Int J Periodontics
Restorative Dent 2006: 26: 223–231.
95. Meffert RM, Thomas JR, Hamilton KM, Brownstein CN.
Hydroxylapatite as an alloplastic graft in the treatment of
human periodontal osseous defects. J Periodontol 1985:
96. Mellonig JT. Freeze-dried bone allografts in periodontal
reconstructive surgery. Dent Clin North Am 1991: 35: 505–
97. Mellonig JT. Human histologic evaluation of a bovine-
derived bone xenograft in the treatment of periodontal
osseous defects. Int J Periodontics Restorative Dent 2000:
98. Minenna L, Herrero F, Sanz M, Trombelli L. Adjunctive
effect of a polylactide ⁄ polyglycolide copolymer in the
Periodontal tissue-engineering technologies