In dentistry, tissue regeneration is a therapeutic aspect which cannot be disregarded for the
achievement of a complete and harmonious restoration of appearance and function.
During the past few years, several techniques have been sug- gested and many
types of materials have been as- sessed, all critically evaluated and revised in
clini- cal studies by means of histological investigations.

The ?Oral and Maxillofacial
Surgery Communi- ty? focused its attention on a set of scientific re- searches
discussing the possibility that a platelet de- rivative could be valid not only
for haemostasis, but also for the emerging field of ?bone grafting?1,2.

In 1997 Whitman et al2
determined the effec- tiveness of the ?Platelet Gel? in speeding up the wound
healing process. In 1998 Marx et al3 paved the way for a new era in ?bone
grafting?, stating that the use of PRP (Platelet Rich Plas- ma), together with
autologous bone, could allow a significantly enhanced result, in comparison with
the graft performed without its use.

The PRP, of which Marx et al
were talking about, consists of a plasma concentrate rich in platelets; it can
be obtained by centrifugation of the venous blood of the patient who will then
receive it as a bone grafting material. This technique had been already used in
dermatology and orthopedics4-6, with the well known growth factors capacity to
involve a wide spectrum of cellular events such as tissue healing, chemotaxis,
mitotic induction and cell differentiation, increase in collagen pro- duction
and angiogenesis.

In particular, PRP is rich in
the following growth factors: Transform- ing Growth Factor-beta (TGF-β),
Platelet-De- rived Growth Factor (PDGF) and Vascular En- dothelial Growth Factor

Their role in tissue recovery
and regeneration, as well as in bone regeneration, is confirmed by several
studies in this regard. Up to date, seven types of TGF-β are known: they are
generated by proteolysis of a higher molecular weight precur- sor, whereas their
activation can occur in a non- e n z y m a t i c w a y o r b y m e a n s o f p l
a s m i n o r cathepsin B7-10. The above-mentioned growth fac- tors are released
by platelets through the degran- ulation process, which occurs few hours after
contact with the receiving surface: in vitro exper- iments demonstrated that
this process exerts its effect even on cells metabolism actively involved in the
early stage of tissue regeneration (mes- enchymal stromal cells, fibroblasts,
osteoblasts and osteoclasts). In particular, the cellular events induced by PRP
three days after grafting in the receiving site, are:

1. Capillary regeneration
2. Increase in osteoblast proliferation
3. Increase in fibroblast proliferation
4. Induction of osteoblast differentiation process, with the consequent
stimulation of mineraliza- tion into the newly-formed bone matrix.

In this regard, it needs to
underline that these events properly occur if the platelet concentration
increases from 2 to 3.38 times the basal platelet count11. Indeed, the
assumption that tissue regen- eration results better in case of a higher
platelet concentration results widely disproven by a set of in vitro

Firstly reported by
international literature, PRP has been widely used in odontostomatology to
accelerate and improve the wound healing process: it was generally used after
radical onco- logic surgery, extractive and periodontal surgery whereas, in
recent times, the PRP is widely used in pre-implant GBR surgeries for the
rehabilita- tion of toothless areas, requiring an increase in bone volume. Data
reported in a large number of scientific reports are often not in accordance,
also because the use of PRP was made both alone or in combi- nation with
autogenous bone, anorganic bone material and organic bone substitutes.

Conflicting opinions also derive
from the analysis of studies aimed to evaluate, in terms of neo-apposition and
osteointegration, the im- provement which can be achieved in case of as-
sociation between PRP and anorganic bone mate- rial (Bio-Oss, Hydroxyapatite),
compared with the use of bone material alone.

In a research carried out in
2001, Marx et al5 stated that the percentage of bone formation was significantly
higher with autogenous bone than either Bio-Oss or Bio-Oss with PRP.

The effects of PRP alone have
been assessed during its use in periodontal and post-extraction surgery and in
the management of bone defects before implant. Also in this case, the reported
data did not allow to out- line a correct and unambiguous conclusion. In fact,
whereas some Authors reported an increase in the initial osteointegration, it
results not sta- tistically significant if using PRP or not. The con- flicting
opinions reported in literature could be partly due to the lack of agreement
about the used animal models.

Furthermore, a negative role
also played the absence of a validated model to obtain the concentrate mix,
together with a standardized procedure protocol, universally recognized and
assumed. Although the used techniques share the main phases procedure, there are
several sub- stances suggested to work within each phase and numerous versions
are proposed to optimize a way to obtain PRP25-28. The aim of this work is to
show the effective- ness of a protocol involving the use of PRP, as a grafting
material in bone regeneration, before dental implant rehabilitation.

Patients and Methods

The study protocol includes a cohort of 127 patients (72 women and 55 men aged
between 32 and 45) requiring maxillary sinus lift, before im- plant-prosthetic
rehabilitation, by using PRP in combination with autogenous bone, anorganic bone
material and organic bone substitutes in 63 patients (test group) and with the
only autoge- nous bone, anorganic bone material and organic bone substitutes in
the remaining 64 patients (control group).

The typology of grafting materi-
al was chosen according to the clinical conditions of each patient.
The bone loss was assessed by means of radiological examinations and
computerized tomography (CT): the choice of implants was case-spe- cific, on the
basis of the typology of rehabilita- tion necessity.
The protocol used to obtain PRP from each patient enrolled in our study was
developed in compliance with the indications reported in literature: PRP can be
obtained by centrifugation of the venous blood of the patient who will then
receive it as a bone grafting material.

First phase: venous blood was
taken (16-24 ml, according to the defect to treat); ? Second phase: coagulation
was promptly in- hibited by using sodium citrate, already pre- sent in the 3.5
ml vacuum venojet tubes which were used in the present study. The test tubes are
characterized by a smooth glass surface; ? Third phase: in all 127 cases, a
single centrifu- gation was performed (speed = 180 rpm = 1000 rpm for 20
minutes), in order to obtain a PRP with optimum physical and biological
properties, to use in the surgical site; ? Fourth phase: the activation of the
preparation was achieved by using calcium chloride, in a ra- tio of 1 g/10 ml of
PRP. This phase is funda- mental to guarantee the correct functioning of the
biochemical processes resulting from the haemostatic, regenerative and
osteoinductive action of PRP. However, before beginning this last phase, the
Authors considered it opportune to use liquid PRP: it was prepared for endosi-
nusal injection, and used for the imbibition of fibrin sponges and implant

Then, 1 g of calcium chloride/10
ml of PRP was mixed with the platelet concentrate and with the grafting
materials in order to fill the bone gaps. All patients accepted in this protocol
were non- smokers and practicing a good home oral hygiene. By selection
criteria, subjects with coagulation and calcium metabolism defects were
excluded, together with subjects with hepatopathies and im- munological
deficits. Before each surgical treatment in both the groups, a professional oral
hygiene was per- formed by supragingival and subgingival scaling; moreover, 2 g
of amoxicillin were administered 1 hour before surgery.

The treated patients of both the
groups were in- cluded in a follow-up plan, which established clin- ical and
radiological examinations on the day after surgery and six months later.

Therefore, the clinical progress
of the healing process, according with the literature, was supported by
radiological investigations. After a dental panoramic X-Ray, a computerized
tomographic examination was made, with the purpose to assess the variation in
bone density around the implant.

From a clinical point of view,
the Authors took in account the peri-implant tissue healing together with
eventual recessive phenomena of the soft tissues. Statistical Analysis At
peri-implant surfaces, changes in bone level (mm) between baseline and 6 months
after surgery, were recorded and analyzed.

Statistical analysis compared
data listed into Group 1 (treated patients) with data listed into Group 2 (non
treated patients = controls) by means of a two tailed paired t test. Results
show an high significance: p < 0.001 with t = 4.28 and df = 62 Mean of
differences resulted 0.724 (95%) with a confidence interval from 0.385 to 1.06.
R squared = 0.228 These data show that the differences between the two groups
result statistically significant indi- cating that, in the group treated with
the PRP, the quality of the response in bone tissue growth is higher than the
response quality found in ?con- trol group? patients.

The achieved results show the
ability of the PRP to promote neo-angiogene- sis during the period of bone
healing. In addition, the PRP acts as a scaffold able to guide the tissue
replacement in a functional manner.

Other clinical and radiological
criteria of Al- brektsson et al29 also confirmed the successful implant
reconstitution. Results In all 127 cases included in the protocol, the Authors
observed a successful implant-prosthetic rehabilitation, according with the
Albrektsson et al criteria29. In fact, there was an optimum im- plant stability
in the monitored patients; besides, no signs of tissue pain affecting the
peri-implant soft tissues were noticed.

The analysis of the vol- ume
ratio of regenerated bone was performed us- ing a personal computer and a
software called ?Master 3D?, which allowed a three-dimensional reconstruction of
the treated area. The different tomographic sections were then scanned and ana-
lyzed on a personal computer by a software of image management, comparing the
videos corre- sponding to both pre- and post-procedure condi- tions.

The radiographic evaluations,
made six months after the implant insertions, showed, in the Test group treated
with PRP, the presence of a newly-formed bone tissue, well amalgamated with the
residual bone, also revealing an average increase in the peri-implant bone
quality, accord- ing with Lekholm and Zarb classification30. Data obtained in
the compared groups resulted statistically significant and the radiographic
integration, at the moment of the implant placement, reached the 100% of all
treated cases.

Table I. Test group (n = 63):
changes in bone level at peri- implant surfaces between baseline and 6 months


bone                             Number level
(mm) at

surfaces patients


3.1 to
4.0                                                            1

2.1 to
3.0                                                            2

1.1 to
2.0                                                          10

0.1 to
1.0                                                          23

0.0 to
-0.9                                                         21

-1.0 to
-1.9                                                          5

-2.0 to
-2.9                                                          1

-3.0 to
-3.9                                                          0

= 63

Discussion Data reported in
literature result sometimes in contrast, also because these studies often sug-
gested the use of PRP both alone or in combina- tion with autogenous bone,
anorganic bone mate- rial and organic bone substitutes11,12,14-18.

The conflicting opinions
reported in literature are certainly caused by a lack of agreement into study
models and especially into an absence of one unique protocol universally
approved and ap- plied in order to obtain the platelet concentrate: the
repeatability and predictability of results could only be achieved by strictly
applying the standardized protocol.

Once the degranulation process
ends, platelets are no more the source of growth factor release. Macrophages,
derived from circulating mono- cytes, intervene in their place. They are drawn
back in large quantities into the graft, due to tissue hypoxia (pO2 between 0.3
and 5 mm/Hg instead pO2 between 35 and 40 mm/Hg in venous blood).

Table II. Control group (n =
64): changes in bone level (mm) at peri-implant surfaces between baseline and 6
months later.


bone                             Number level
at  of peri-implant
surfaces patients


3.1 to
4.0                                                           0

2.1 to
3.0                                                           0

1.1 to
2.0                                                           4

0.1 to
1.0                                                          18

0.0 to
-0.9                                                         32

-1.0 to
-1.9                                                        8

-2.0 to
-2.9                                                        1

-3.0 to
-3.9                                                        1

= 64

Thanks to their fast replication, macrophages are
remarkably efficient in growth factors production, being actively involved in
the second stage of tis- sue regeneration. As the induced revascularization
increases, a decrease in the oxygen pressures val- ues is found and,
consequently, with a significant reduction in macrophage action. Complete revas-
cularization occurs on the 14th day, when bone ma- trix deposition by stem cells
(endosteal os- teoblasts) is fully detected.

Up to now, the exact concentration able to optimize
the aforesaid processes has not yet been detected; medical liter- ature is only
in agreement with the above-men- tioned range12. Higher platelet concentrations,
in fact, are not able to sustain an increase in os- teoblast and fibroblast

Even though it is widely stressed the necessi- ty to continue this study on a
larger cohort of patients, the Authors verified and documented the positive
clinical-radiological examinations achieved by using the described protocol in
all treated cases. Therefore, this paper tested, with a standardized
methodology, the possibility to satisfactorily use PRP in the clinical practice
with a great predictability about its success.


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