Review

Platelets’ Role in Dentistry: From Oral Pathology to

Regenerative Potential
Serena Bianchi 1, Diana Torge 1, Fabiola Rinaldi 2, Maurizio Piattelli 2, Sara Bernardi 1,3,* and Giuseppe Varvara 2

1 Department of Life, Health and Environmental Sciences, University of L’Aquila, 67100 L’Aquila, Italy;
serena.bianchi@univaq.it (S.B.); diana.torge@graduate.univaq.it (D.T.)
2 Department of Innovative Technologies in Medicine & Dentistry, University of Chieti—Pescara ‘Gabriele

d’Annunzio’, Via dei Vestini 11, 66100 Chieti, Italy; f.rinaldi0993@gmail.com (F.R.);
maurizio.piattelli@unich.it (M.P.); gvarvara@unich.it (G.V.)
3 Center of Microscopy, University of L’Aquila, 67100 L’Aquila, Italy

* Correspondence: sara.bernardi@univaq.it

Abstract: Platelets are a cellular subgroup of elements circulating in the bloodstream, responsible
for the innate immunity and repairing processes. The diseases affecting this cellular population,
depending on the degree, can vary from mild to severe conditions, which have to be taken into
Citation: Bianchi, S.; Torge, D.;
consideration in cases of minor dental procedures. Their secretion of growth factors made them
Rinaldi, F.; Piattelli, M.; Bernardi, S.; useful in the regenerative intervention. The aim of this review is to examine the platelets from bio-
Varvara, G. Platelets’ Role in logical, examining the biogenesis of the platelets and the biological role in the inflammatory and
Dentistry: From Oral Pathology to reparative processes and clinical point of view, through the platelets’ pathology and their use as
Regenerative Potential. platelets concentrates in dental regenerative surgery.
Biomedicines 2022, 10, 218.
https://doi.org/10.3390/ Keywords: platelets; PRF; PRP; bleeding disorder
biomedicines10020218

Academic Editor: Raffaele Capasso

Received: 21 December 2021

1. Introduction
Accepted: 19 January 2022
Published: 20 January 2022
Platelets represent a cellular subgroup of the elements circulating in the bloodstream.
A human adult has nearly one trillion circulating platelets, with a lifespan of 8–10 days
Publisher’s Note: MDPI stays
[1]. These cellular circulating elements play important key roles: responding to vessel in-
neutral with regard to jurisdictional
juries, regulating angiogenesis, and the innate immunity [2].
claims in published maps and
The functional roles in hemostasis, inflammation, and regeneration place the plate-
institutional affiliations.
lets as characters of human physiology (Figure 1); eventual pathologies affecting them
influence the daily life of patients, as well as their survival in severe cases [3].
In dental medicine, platelets are studied and well-acknowledged due to the disorders
Copyright: © 2022 by the authors. Li-
that can affect the hemostasis [4] and the regenerative potential in oral surgery procedures
censee MDPI, Basel, Switzerland.
[5].
This article is an open access article From the place of their origin, the bone marrow, through the bloodstream, to the
distributed under the terms and con- periphery, platelets’ journey is worthy of being examined.
ditions of the Creative Commons At- This review aims to go through the platelets’ biology, describing their origin, mor-
tribution (CC BY) license (https://cre- phology, physiology, and pathologies relevant to oral surgery procedures and the related
ativecommons.org/licenses/by/4.0/). therapies. Finally, we examine the recent regenerative potentiality in dental surgery
driven by the use of autologous platelet concentrates.

Biomedicines 2022, 10, 218. https://doi.org/10.3390/biomedicines10020218 www.mdpi.com/journal/biomedicines

Biomedicines 2022, 10, 218 2 of 14

Figure 1. From the bone marrow (A) through the bloodstream, the megakaryoblast (B) evolves into
megakaryocyte (C,D) to finally acquire the platelet form (E). Platelets are activated during the in-
jures (F) by means of the coagulation cascade to block the blood loss and to start the very first steps
of healing.

1.1. Biological Characteristics of Platelets

Platelets are small, anucleate, and disc-shaped cells whose main structural elements
are plasma membrane, open canalicular system (OCS), dense tubular system (DTS), a
spectrin-based membrane skeleton, and actin-based cytoskeletal network. They also pre-
sent a peripheral band of microtubules and several organelles, such as α-granules, dense
granules, peroxisomes, lysosomes, and mitochondria [6].
The plasma membrane owns a bilayer of phospholipids, in which cholesterol, glyco-
lipids, and glycoproteins are immersed. In this plasma membrane, sodium and calcium-
adenosine triphosphate ensure the intracellular ionic environment of the platelet, while
phospholipids’ task is to regulate the coagulation process. Other crucial components of
the platelet plasma membrane are lipid rafts, cholesterol, and sphingolipid-rich microdo-
mains, essential for signaling and intracellular trafficking. In addition, the platelet plasma
membrane also includes surface receptors, which control signal-dependent platelet acti-
vation and participate in coagulation, inflammation, antimicrobial host defense, angio-
genesis, and wound repair [7].
The OCS regulates the flux of external elements into the platelet, and at the same
time, it is essential for the release of granule contents. Additionally, OCS is also an exten-
sive internal membrane reservoir that promotes filopodia formation and platelet adhe-
sion. Lastly, the OCS is a storage site for plasma membrane glycoproteins [7]. DTS is a
closed-channel network of endoplasmic reticulum residues, whose primary function is to
capture ionized calcium (Ca2+) supported by calreticulin, a calcium-binding protein.
The cytoskeleton of inactive platelet is well-defined and highly specialized, which
protects the morphology and integrity of platelets. Platelet cytoskeleton is composed of a
spectrin-based membrane skeleton, an actin cytoskeleton, and a microtubule coil. Spec-
trin-based membrane skeleton consists of self-assembled or elongated spectrin strands
that laminate the cytoplasmic surface of OCS and plasma membrane systems. In the actin
cytoskeleton, forty percent of the actin subunits assemble to create 2000–5000 linear fila-
ments in the resting platelets, while tubulin is equally distributed in dimer and polymer
fraction in the resting microtubule coil [6,7].
Concerning granules distribution, in the inactive platelet, they are located near the
OCS membranes, and during the activation process, they fuse and secrete into the OCS.
In this context, platelets have two primary storage granules: α-granules and dense gran-
ules [7]. Alpha granules are the most widespread and most prominent secretory granules:
according to their function, they present proteins essential for platelet adhesion during
the vascular repair. With a 200–500 nm diameter, these granules are characterized by a
spherical shape and a dark central core. They arise from the trans-Golgi network and in-
clude several membrane proteins essential for platelet function (e.g., P-selectin and CD36).
Alpha granules also present over 30 angiogenetic proteins, regulators of the new blood
Biomedicines 2022, 10, 218 3 of 14

vessel. Dense granules are 250 nm in size, visible on electron microscopy as dense bodies,
with an electron-dense core. In this context, dense granules are helpful to collect addi-
tional platelets in the site of vascular damage, and they also contain active hemostatic
molecules, such as catecholamines, serotonin, calcium, ADP, and ATP [1].
Platelets show a small number of mitochondria, valid for energy production. They
also present lysosomes, which contain hydrolytic enzymes, including acid phosphatase,
arylsulfatase, and peroxisomes, dispersed in their cytoplasm [6].
Daily platelets production starts from bone marrow megakaryocytes, rare myeloid
cells, which represent less than 0.1% of the cellular population of the bone marrow.
Megakaryocytes originate from pluripotent hematopoietic stem cells, which create burst
and colony-forming precursors and are subjected to maturation. Thrombopoietin (TPO),
Interleukin-3 (IL-3), stem cell factor (SCF), Interleukin-6 (IL-6), and Interleukin-11 (IL-11)
are the main regulating factors of megakaryocytes’ differentiation process [8].
During the maturation process, megakaryocytes are subjected to endomitosis, a chro-
mosomal duplication without cell division, which generates a unique polylobulated nu-
cleus without completing the M phase. Ploidy is essential for megakaryocytes’ cytoplas-
mic volume, as it is determinant in lipid and protein synthesis, considered propaedeutic
events for platelets’ production. Lipid and protein synthesis contributes to increasing
megakaryocyte’s size (approximately to a diameter of 100 µm), supporting the production
of platelet-specific organelles, granules, cytoskeletal proteins, and the invaginated mem-
brane system, an extensive internal membrane reservoir, essential for platelet synthesis
[6]. Regarding the maturation dynamics, during this process, megakaryocytes move from
the osteoblastic to the vascular niche, a dynamic scaffold, which regulates megakaryocyte
maturation and, consequently, platelets synthesis.
Another significant stage, which contributes to the complex nature of platelet syn-
thesis, is proplatelet elongation. This process starts with the erosion of megakaryocytes,
which leads to the genesis of pseudopod-like structures that elongate tubular projections.
A complex network of protein filaments regulates proplatelet elongation: the continuous
polymerization of tubulin bundles determines the elongation process at their free plus
end, and simultaneously, during this window of time, there is also the sliding of overlap-
ping microtubules induced by dynein.
As regards the actin contributing to proplatelet production, limited evidence con-
firms its pivotal role in this process [1]. In addition, another significant regulator of plate-
lets’ synthesis is the hematopoietic β–1 tubulin isoform, expressed by mature megakaryo-
cytes that elicits an essential role not only in platelets’ production but also in the definition
of their structure and function.
Due to the migration process from the osteoblastic to the vascular niche, which in-
volves several megakaryocytes, bone marrow stroma plays a fundamental role in
megakaryocyte maturation is reserved for [6]. The bone marrow influences proplatelet
synthesis by expressing specific matrix-receptor signals: type I collagen, for example, an
extracellular protein widely diffused in the osteoblastic niche. Simultaneously, bone mar-
row prevents proplatelet synthesis and enhances megakaryocyte spreading. These dy-
namics are governed by the interaction between type collagen I and integrin α2β1, acti-
vating the small GTPase Rho and Rho-kinase ROCK. In addition, Rho-kinase ROCK is
involved in the phosphorylation of the regulatory light chain of myosin IIA to reduce my-
osin ATPase activity and overpower proplatelet production. Moreover, among the most
significant extracellular matrix proteins with a regulating potential on platelet production
are type IV collagen, fibrinogen, and vitronectin [6].
In conclusion, proplatelet production results from a complex balance between mo-
lecular and protein factors, which influence the cellular environment of platelets and de-
termine their varied biological function.
Biomedicines 2022, 10, 218 4 of 14

1.2. Platelets’ Activation

As previously described, platelets arise from megakaryocytes and are rapidly re-
cruited during vessel damage to create a hemostatic clot in the vessel wall. Moreover, the
platelet adhesion and aggregation process require specific platelet–endothelium and
platelet–platelet interactions. The adhesion process is the first step of primary hemostasis:
at this point, platelets interact with extracellular matrix proteins through adhesive glyco-
proteins (GP). Therefore, platelets’ activation results from a complex cascade of signals
and receptor cross-linking, a process that induces significant changes in the assembly of
cytoskeletal proteins [7]. At the same time, when the endothelium of a blood vessel is
damaged, damaged cells produce endothelin, enhancing vessel contraction to stop blood
loss. At this level, plasma glycoprotein von Willebrand factor (vWF) binds to the exposed
collagen through GPIIb/IIIa receptors on the surface of the subendothelium as a substrate
for platelet adhesion.
Alternative adhesion substrate in this step is also the modular glycoprotein throm-
bospondin-1 (TSP1), while among the most involved collagen receptors, there are
GPIa/IIa, CD36, and GPVI, essential in promoting collagen-induced signaling [1,7].
After their activation, platelets release different granule components, which ensure
the interaction between platelets and vascular cells [9]. Platelet dense bodies release sec-
ondary agonists, such as ADP and serotonin. ADP is considered a prominent amplifier of
platelet activation, while serotonin exerts a procoagulant role. As regards serotonin, this
vasoconstrictor binds to Gq-coupled 5HT2A receptor and enhances the platelet response
through ADP. At the same time, the dense tubular system contains a Ca 2+ pool, whose
fluxes are essential for platelet activation, attraction, and aggregation. In this context, re-
ceptors expressed on platelets’ surface support platelets’ secretion and their irreversible
aggregation: among them, the most significative are P2Y1 receptor, which regulates the
mobilization of Ca2+ and morphology changes, and the P2Y12, essential in platelet secre-
tion and irreversible aggregation [7]. The α-granules instead present large adhesive pro-
teins (e.g., vWF, TSPI, vitronectin, fibronectin), but also mitogenic factors (e.g., PDGF,
VEGF, TGFβ), coagulation factors (e.g., factors V, VII, XII, XIII), and protease inhibitors,
released after the activation stage [1,7].
Another significant event for platelet physiology is their aggregation: this stage is
characterized by the accumulation of platelets into a hemostatic clot and is completely
regulated by GPIIb/IIIa, the central platelet receptor. The platelet stimulation during the
aggregation induces an increase in GPIIb/IIIa, through α-granules exocytosis and activa-
tion of surface-exposed GPIIb/IIIa. The linkage between GPIIb/IIIa and its ligands pro-
motes considerable molecular and morphological changes, leading to phosphorylation of
tyrosines of the cytoplasmatic GPIIIa-chain [7]. Disulfide variations in the GPIIb/IIIa com-
plex, induced by the disulfide isomerase, enhance high-affinity binding sites for fibrino-
gen, which usually binds discoid platelets during platelet–platelet contact. In the end, low-
density lipoproteins (LDL) are also considered a ligand of GPIIb/IIIa, with potential reg-
ulative effects on platelet function.
During secondary hemostasis, the generation of a platelet clot is determined by a
thrombin-mediated conversion of fibrinogen to fibrin. At this stage, the platelet plasma
membrane is essentially formed by phospholipids, cholesterol, and glycolipids. Among
phospholipids, phosphatidylcholine (PC), sphingomyelin (SphM), phosphatidylethanola-
mine (PE), phosphatidylserine (PS), and phosphatidylinositol (PI) are located in the
plasma membrane: specifically, SphM and PC are diffused in the outer left, while PE in
the inner left [7]. The activation process of platelets is linked to a flip-flop move of PS,
whose exposure on aggregated platelets supplies a catalytic surface for pro coagulation,
preventing the synthesis of thrombin in the site of injury. The shedding of microvesicles
from the platelet surface is due to the combination of collagen and thrombin: this process
is determined by activation of calpain, induced by Ca 2+, which promotes the formation of
procoagulant vesicles.
Biomedicines 2022, 10, 218 5 of 14

At the same time, a stimulatory platelet signaling enhances the production of several
intracellular messenger molecules: among them, Ca2+, products of the phospholipase C
(PLC) phosphoinositol hydrolysis, diacylglycerol, inositol-1, 4, 5-triphosphate (IP3), and
thromboxane (TxA2). ADP and TxA2 interconnect with seven transmembrane receptors
associated with GTP-binding heterotrimeric G proteins to activate different signaling
pathways. PLC activation is due to receptors coupled to the Gq family of G proteins, and
this activation induces PIP2 hydrolysis, with mobilization of Ca 2+ from the dense tubular
system [1,7]. The increase in intracellular Ca2+ enhances the phosphorylation of the myosin
light chain as an essential event for morphology change. Granule secretion is another sig-
nificant response to Ca2+ mobilization, which determines the release of ADP from dense
bodies: after this release, ADP links to P2Y12, supporting platelet activation. Regarding
the TxA2 activation pathway, it is confirmed that TxA2 binds to Gq-coupled TP receptors
to enhance the stimulatory process [1,7].
In the absence of a wound, prostaglandin I2 and nitric oxide (NO), released by the
endothelial cells, represent platelet inhibitors. They promote an increase in cAMP through
adenylate cyclase and guanylate cyclase, respectively, and promote a decrease in IP3 and
Ca2+ levels, with a consequential decrease in platelet activation [7].

2. Platelets’ Pathologies Relevant for Oral Surgery

Platelets, therefore, play a crucial role in the coagulation cascade as constituents of
factor X and prothrombin conversion complexes through the release of platelet factor 3
(PF3) [10]. In some cases, platelets may not complete the release of this factor due to de-
fective thromboxane production because of the administration of anti-inflammatory
drugs such as aspirin, NSAIDs, tricyclic antidepressants, lactam antibiotics, and calcium
channel blocker drugs, but also due to a deficiency in the production of dense granule
ADP [11]. While in healthy subjects, drug-induced impairment of platelet function has no
clinical significance, in patients with coagulation disorders, uremic or thrombocytopenic
patients, and those taking heparin or coumarin anticoagulants, drug-induced platelet dys-
function can cause severe bleeding [4]. Screening tests should therefore be carried out on
these patients before any surgical procedure [4].
Structural or enzymatic alterations of platelet cause functional deficits and platelet
disorders, which can be congenital or acquired [12].
Depending on the function that is compromised, we can distinguish [13]:
− Platelet diseases with an adhesion defect;
− Thrombocytopathies with an aggregation defect;
− Thrombocytopathies with a defect in the release reaction.
One of the causes of hemorrhagic diathesis, therefore, is platelet deficiency or dys-
function. The severity of hemorrhagic diathesis is very variable, concerning the varying
degrees of platelopenia and the possible coexistence of a defect in platelet function.
A reduction in the number of platelets, or thrombocytopenia, causes hemorrhages if
the platelet count falls below 100,000 per mm3; however, spontaneous bleeding does not
occur if the count does not reach a value of fewer than 20,000 platelets. Individuals with
low platelet counts and no alterations in function do not bleed spontaneously but may
experience prolonged bleeding during trauma, surgery, dental extractions, or biopsies [4].
Thrombocytopenias can be caused by [14]:
− Reduced platelet production: aplastic anemia, leukemia, vitamin B12, or folic acid
deficiency cause a reduced development and faster destruction of bone marrow cells,
including the megakaryocytes. In cases of aplastic anemia, which can be inherited or
acquired, the bone marrow stem cells are incapacitated to proliferate and differenti-
ate.
Dental management of patients affected by aplastic anemia includes an accurate
medical history and an interdisciplinary approach with the cooperation of the hematolo-
gist. In cases of surgical intervention, the treatment should be performed on the
Biomedicines 2022, 10, 218 6 of 14

transfusion day, and antifibrinolytics should be prescribed to control the bleeding. In

cases of some forms of acute leukemia, dental treatment is recommended before starting
the oncologic therapy.
− Destruction of platelets is due to [14]:
• Immunological mechanisms: circulating antibodies or immune complexes can
cause immune-mediated destruction of platelets following autoimmune, drug-
induced, or HIV-associated diseases.
• Mechanical damage: caused by platelets colliding with prosthetic heart valves,
thrombi, or narrowed vessel walls.
• Hypersplenism: due to increased platelet sequestration by the splenic phagocyte
system.

2.1. Immunological Mechanisms: IMMUNE Thrombocytopenic Purpura (ITP)

The ITP is caused by the formation of anti-platelet autoantibodies (IgG) directed
against the glycoproteins of the platelet membrane, which cause their destruction [15].
The diagnosis depends on excluding other causes of platelopenia, so it is a disease of ex-
clusion.
ITP can be chronic and may occur more frequently in adult women under 40 years of
age, in whom immune-mediated platelet destruction may be due to systemic lupus ery-
thematosus, AIDS, complications of drug therapies, or it may be idiopathic [15]. Bleeding
from the skin may occur as spot hemorrhages (petechiae), which may coalesce into ecchy-
moses acute: this is a disease of childhood, with equal frequency in both sexes. Its onset is
sudden and is preceded by a viral illness, with an interval of 2 weeks, but it usually re-
solves spontaneously.
Recently, some cases of ITP have been described after administration of mRNA
SARS-CoV-2 vaccines and after adenovirus-based SARS-CoV-2 vaccine [16]. Most pa-
tients responded to treatment with corticosteroids and intravenous immunoglobulins
(IVIG) but showed little benefit following platelet transfusion, as is the case with ITP. Fur-
thermore, it is unclear whether these cases are self-limiting or will persist and lead to
chronic ITP.

2.2. Immunological Mechanisms: Heparin-Induced Thrombocytopenia

This type of thrombocytopenia is caused by the immune-mediated destruction of
platelets by heparin, which binds to platelet factor IV and changes its conformation [17].
At this point, the immune system no longer recognizes platelet factor IV, which, bound to
heparin, is attacked by antibodies, resulting in the formation of immune complexes. These
immune complexes, even under conditions of thrombocytopenia, can paradoxically cause
thrombosis by activating the platelets [17].

2.3. Immunological Mechanisms: HIV-Associated Thrombocytopenia

Thrombocytopenia is the most common hematological manifestation in HIV infec-
tion due to reduced production and increased destruction of platelets [18]. Indeed, the
CD4 molecule, the central HIV receptor, in megakaryocytes undergo apoptosis. In addi-
tion, HIV infection also induces hyperplasia and dysregulation of B cells, predisposing to
the development of immune-mediated thrombocytopenia, characterized by the formation
of anti-platelet antibodies [18].

2.4. Mechanical Damage: Thrombotic Microangiopathies

Thrombotic microangiopathies comprise a variety of clinical syndromes, including
thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS) [19].
Both are characterized by fever, thrombocytopenia, microangiopathic hemolytic anemia,
and renal failure, more severe in HUS [14]. Transient neurological deficits are also present
in TTP, absent in HUS, which has a higher incidence in children. A common feature of
Biomedicines 2022, 10, 218 7 of 14

both conditions is the widespread formation of hyaline thrombi within the microcircula-
tion, which causes platelet consumption (thrombocytopenia) and microangiopathic he-
molytic anemia due to the clash of red blood cells with these thrombi [19]. However, the
pathogenetic mechanism responsible for thrombus formation is different. In TTP, there is
a deficiency of an enzyme known as ADAMTS 13 or “von Willebrand factor metallopro-
teinase”, which degrades von Willebrand factor multimers, representing the form in
which it circulates in the blood. In the absence of this enzyme, von Willebrand factor mul-
timers accumulate in the plasma and promote the formation of platelet microaggregates
in the microcirculation, causing TTP. In HUS, the cause is gastroenteritis caused by E. coli,
which produces a toxin that binds to endothelial cells in the renal glomerulus and other
organs, damaging them and initiating platelets activation and aggregation [19]. HUS can
also affect adults, following pathologies that cause endothelial damage (drugs, radiother-
apy). TTP and HUS are differentially diagnosed with disseminated intravascular coagu-
lation (DIC) since they are all characterized by microvascular occlusion and microangio-
pathic hemolytic anemia [19], but in the former, activation of the coagulation cascade does
not play a primary role; therefore, coagulation tests such as PT and PTT usually show
typical values.

3. Hemorrhagic Diseases of Impaired Platelet Function

Qualitative defects in platelet function may be congenital or acquired. Acquired dis-
orders mainly include ingestion of aspirin and other non-steroidal anti-inflammatory
drugs [20] due to irreversible inhibition of cyclooxygenase, an enzyme required for throm-
boxane A2 synthesis, and uremia, during which numerous alterations in platelet function
have been observed. However, the pathogenesis of bleeding during uremia is complex
and not well understood.
Congenital disorders of platelet function, on the other hand, can be due to:
− Defects in platelet adhesion to the subendothelial matrix [21], as in the case of von
Willebrand disease or Bernard-Soulier syndrome [22], an autosomal recessive disor-
der caused by the hereditary deficiency of the membrane glycoprotein complex Ib-
IX, which acts as a receptor for von Willebrand factor, essential for the physiological
adhesion of platelets to the subendothelial matrix.
Von Willebrand disease is classified as the most common inherited bleeding disorder
due to an inherited defect that causes both deficits in platelet function and the coagulation
cascade [23]. Indeed, the von Willebrand factor is crucial in platelet adhesion to the ex-
posed extracellular matrix following vascular injury and in transporting coagulation fac-
tor VIII, a component of the intrinsic pathway required for factor X activation and ensur-
ing its stability [24]. The half-life of circulating factor VIII is approximately 12 h if von
Willebrand factor levels are standard but only 2–4 h if von Willebrand factor is defective
or deficient, with alteration of the coagulation cascade [24]. Unbound factor VIII is there-
fore destroyed in the circulation.
Depending on the severity of the genetic expression, the disease has several variants,
most of which are transmitted as autosomal dominant traits (types 1 and 2) [25]. The dif-
ferent variants of von Willebrand disease tend to cause mild to moderate clinical bleeding
problems: notably, in mild cases, bleeding occurs only after surgery or trauma; in severe
cases, spontaneous nosebleeds or bleeding of the oral mucosa may be noted [25].
Among the variants of von Willebrand disease [26], we can distinguish:
• Diseases associated with a reduction in circulating von Willebrand factor:
− Type 1 (70–80% of cases) is the most common form of von Willebrand disease,
due to non-sense mutations, with mild but variable clinical manifestations: the
more significant the vWF deficiency in type 1 disease, the more likely we are to
find signs and symptoms of hemophilia A [26].
− Type 3 is a rare autosomal recessive form characterized by severe clinical mani-
festations caused by frameshift mutations or deletions [26].
Biomedicines 2022, 10, 218 8 of 14

• Diseases associated with qualitative deficiencies of von Willebrand factor:

− Type 2, responsible for 15–20% of cases, is an autosomal dominant form due to
non-sense mutations causing defective assembly of the multimers, with mild to
moderate symptoms. Treatment involves desmopressin, which stimulates vWF,
or factor VIII and vWF concentrate [26].
− Defects in platelet aggregation, such as Glanzmann’s thrombasthenia, an auto-
somal recessive disorder characterized by a deficiency or malfunction of glyco-
protein IIB-IIIa, a protein complex that contributes to the formation of “bridges”
between platelets [26].
Platelet secretion disorders are characterized by a normal initial response to the ag-
gregating stimulus of collagen but by a defect in the secretion reactions of thromboxane
and the release of ADP in the granules [14].
Hereditary thrombocytopathies, therefore, derive from genetic alterations that lead
to a deficiency in the number of platelets: there are diseases transmitted by a “dominant”
and “recessive” mechanism [27].
In the first case, it is enough for only one of the two genes inherited from the parents
to be defective for the development of thrombocytopenia. The recognition of the genetic
nature of the defect is generally easy thanks to the medical history, which reveals the fa-
ther or mother is also ill and, often, that other blood relatives have the same defect [27].
In the second case, the mutation is less serious and induces thrombocytopenia only
when both genes are defective: the parents are “healthy carriers” of the disease, and only
the son shows thrombocytopenia [27].
Finally, “sex-linked recessive” transmission is rare and occurs when a mother is a
carrier, and only the male child/children develop the disease [27].
The clinical conditions derived from platelets’ defects are represented by various
bleeding manifestations, from mild to moderate to very severe, affecting the skin, oral and
nasal mucosa, intestine, and uterine mucosa [14]. In case of mucosal bleeding manifesta-
tions, with average platelet count, it is advisable to refer to the specialist center to perform
bleeding time and in vitro platelet function tests (Born platelet aggregation or PfA-100
test) and perform the differential diagnosis with von Willebrand disease [14].
In order to reduce both the risk of gum disease and the occurrence of dental diseases
requiring gum surgery, it is necessary to pay great attention to oral hygiene from an early
age and prevent, in general, traumas and injuries [4].

4. Role of Platelets Derivates in Regenerative Dental Procedures

In dentistry, platelets find various applications in oral and maxillofacial surgery due
to the possible use as a grafting material for bone rehabilitation to improve bone regener-
ation, especially in bone volume augmentation [28]. Regenerative dentistry represents a
discipline continuously in evolution, where not only platelets and dentinal derivates are
used as autologous biomaterial, but also the experimental use of stem cells in vitro studies
gave promising results [29,30].
As known, the repair or regeneration of damaged tissue is a biological process in
which various factors are involved, such as immune cells, growth factors, proteins, and in
which we can distinguish four main phases [31]: hemostasis, inflammation, proliferation,
and maturation.
The whole process is very complex, as it requires the combination of many factors
and the perfect timing of their activation [31].
The technology advancement and the available centrifuges allowed the possibility to
obtain platelet concentrates on an outpatient basis: the centrifugation of the blood taken
from the patient separates the plasma part, rich in platelets, for autologous therapeutic
use [32]. The fractioned blood is used to obtain both platelet gel (with a higher concentra-
tion of platelets than physiological) and fibrin glue with a high adhesive and hemostatic
property [32].
Biomedicines 2022, 10, 218 9 of 14

The two most common platelet concentrates are platelet-rich plasma (PRP) and plate-
let-rich fibrin (PRF), which represent the first and second generation (Table 1), respec-
tively [33].

Table 1. Resuming table of the properties of PRP and L-PRF.

Blood Product L-PRF PRP

Protocol 2700 rpm for 12′ 2400 rpm 10′
Flow One step continuous Two steps cloth activation
PDGF levels (ng/mL) [32] High Low
VEGF levels(ng/mL) [34] Highest Low
TGF-b 1 (ng/mL) [34] High Low
Reproducibility No bias Possible bias
Use of anticoagulants No Yes
Fibrin density High Low
Speed of fibrin formation High Low
Fibrin morphology Tetramolecular Tetramolecular
Handling Easy Complex
PDGF—platelet-derived growth factor, VEGF—vascular endothelial growth factor, TGF-β 1—
transforming growth factor β 1.

The PRP is obtained from the heparinization, which makes it incoagulable of the
blood taken from the patient and can be stored for a few days at −19 °C, or it can be pre-
pared immediately after the blood collection [35]. In any case, before centrifuging the
blood (the protocol includes two centrifugations) to remove the red blood cells, anticoag-
ulant factors such as bovine thrombin or CaCl2 must be added to trigger the coagulation
and fibrin formation cascade [35]. This process leads to a tumultuous fibrin network for-
mation process, resulting in somewhat irregular fibrin [35]. PRP should also be used
within 4 h of isolation, as growth factors are secreted within 10 min of preparation and
reach 95% after 1 h [35].
This type of platelet concentrate presents disadvantages [36]:
• The presence of anticoagulants such as bovine thrombin could cause allergic reac-
tions and coagulopathies due to the action of antibodies against factors V, XI, and
consequent thrombus formation [36];
• Final preparation without rigidity requires the further addition of bone grafts to
maintain a stable volume [36].
The PRF belongs to a new generation of hemo-concentrates obtained by the sole cen-
trifugation, which do not require additives such as heparin or thrombin, allowing a slow
release of growth factors [37]. Therefore, the blood is not manipulated in any way, and the
obtained PRF must be used at the same time as its production since it is not storable [37].
The fibrin formation process is much more regular, and the cytokines and the growth fac-
tors remain trapped in its structure, making it stronger and more elastic and considerably
increasing regenerative capacity [37,38].
PRF has the following bioactive properties:
• Stimulation, through the growth factors contained within it, of the proliferation,
differentiation, chemotaxis, and adhesion of stem cells, promoting angiogenesis and
immune processes [34];
• Increased expression of alkaline phosphatase (ALP) in stem cells, leading to faster
mineralization of the newly formed tissue [34];
• Induction of mineralization of the defect, thanks to the growth factors it contains
(TGF-β1 and PDGF) [39];
• The creation of an epithelial barrier by the PRF membrane [40].
Biomedicines 2022, 10, 218 10 of 14

The capacity for clot formation and organization is increased, thanks to the formation
of a regular fibrin network, which provides an osteoconductive scaffold suitable for neo-
angiogenesis and cell migration, which is essential for the bone apposition process
[34,35,38–40].
This increases the speed of healing and the quality of the newly formed tissues.
The different types of PRF preparation found in the literature are based on different
relative centrifugation forces (RCF).
Choukroun’s L-PRF or leukocyte-PRF (solid PRF) protocol includes [37]:
• A blood sample (10 to 100 cc of blood) taken and placed in a 10 mL glass tube (not
plastic as the latter activates fewer coagulation factors);
• Centrifugation at 3000 rpm for 10 min.
This centrifugation produces a total concentration with a high percentage of growth
factors, immune cells, and platelets that release important cytokines.
At the end of centrifugation, the patient’s blood is separated into three phases:
• Superficial layer: cellular plasma;
• Middle layer: fibrin + platelets + leucocytes (PRF);
• Deep layer: blood cells.
The intermediate layer takes the form of a gelatinous substance, which can be used
either as a membrane or as an adjunctive biomaterial [37]. The L-PRF does not have rigid-
ity and always requires the support of a filler (even crushed PRF). In addition, it does not
need to be cut or sutured: its consistency allows it to adapt perfectly to the bone. In clinical
practices, the L-PRF is placed between two surfaces (even between two glass plates used
to mix the cement, wrapped in sterile gauze) and exerts pressure with their weight for a
certain period, transforming it into a resorbable membrane. Due to its rapid resorption
time, the L-PRF represents a “reservoir” of growth factors [39].
The Ghaanatis PRF (also known as advanced PRF or A-PRF) is a solid PRF more mal-
leable than the L-PRF. It is obtained using a reduced centrifugal force, 208 G on plastic
tubes (whereas in the previous PRF, it is 708 G on glass tubes) [41].
The main advantages include a high concentration of leukocytes (due to slow centrif-
ugation), a high concentration of platelets and neutrophil granulocytes, and a good release
of different growth factors [41]. The matrix is more porous, allowing a better angiogenesis
and blood vessel penetration; the soft consistency allows different use of the material such
as:
• Pellets for bone grafting;
• Membrane.
The I-PRF is a platelet concentrate in the liquid form obtained using centrifugal force
60 G (at higher speed for less time), so no coagulation takes place [42].
Due to the lower centrifugation speed and time, it contains a high concentration of
leukocytes and blood plasma proteins, and it can be used in different ways, such as injec-
tion into the deep tissue space or mixed with other graft materials [42].
The main advantages represented by the platelet concentrations are the contained
growth factors [43]. The growth factors are released by platelets, macrophages, and leu-
kocytes, which play a role in inflammation and wound healing by stimulating or inhibit-
ing the mechanisms of migration, adhesion, proliferation, and differentiation [43]. They
can exist in two forms: active and inactive and have short biological half-lives.
Growth factors are linked to the different types of cells that secrete or stimulate the
release of these factors [44]. Depending on the centrifugation, therefore, a different per-
centage of growth factors can be obtained in different types of platelet concentrate [44].
Among the various growth factors, mention must be made of the following [43]:
• Endothelial growth factor (EGF): it stimulates endothelial cell chemotaxis, mesen-
chymal cell mitosis, epithelialization and increases tissue tensile strength [43].
Biomedicines 2022, 10, 218 11 of 14

• Transforming growth factor β1 (TGF-β1): it regulates angiogenesis, upregulates

vascular endothelium growth factor (VEGF), regulates the tissue repair process,
immune modulation, synthesis of extracellular matrix, and plays a key role during
bone regeneration, stimulating the chemotaxis and mitogenesis in osteoblast pre-
cursors and the deposition of mineralized tissue on the bone collagen matrix [43].
• Platelet-derived growth factor (PDGF): this factor is mainly produced by platelets,
and it can be in the fibrin matrix in high quantities. PDGF regulates the migration,
the proliferation of mesenchymal cells, stimulates collagen production, the mito-
genesis of osteoblasts, fibroblasts, smooth muscle cells, and glial cells [43].
• Insulin-like growth factors (IGF): the IGF is secreted by platelets and over-regulates
the proliferation and differentiation of various cells involved in tissue repair mecha-
nisms, the differentiation and proliferation of mesenchymal cells, and at the same
time, it is a regulator of programmed apoptosis [43].
• VEGF: macrophages and thrombocytes secrete VEGF, the main factor in stimulating
angiogenesis, modulating tissue remodeling, and when added to bone materials,
stimulates the formation of new bone tissue [43].

Clinical Use of Platelets Derivates

Several studies and literature reviews, due to the regenerative properties mentioned
above, have highlighted the effectiveness of the regenerative potential of these prepara-
tions [45].
The clinical applications of these preparations are in various areas such as guided
periodontal and bone regeneration, sinus floor elevation, and support for the regeneration
of the pulp-dentine complex [46–48]. However, there is no shortage of studies highlight-
ing the limited effects of, for example, the efficacy of PRP in bone formation during sinus
floor elevation procedures, as noted in the systematic review by Pocaterra et al. [49].
PDGF has also been used, in combination with advanced surgical techniques, to treat
infrabony defects, as shown in the study by Del Fabbro M. et al. [50]. The study evaluated
the effects of autologous platelet concentrates together to open periodontal surgery, with
or without bone grafting, guided tissue regeneration (GTR), or in addition or without
enamel matrix derivatives. However, the authors did not find that the addition of autolo-
gous platelet concentrates (APCs) to specific surgical techniques such as open flap with
or without bone graft, or GTR or interventions with the presence of enamel matrix deriv-
atives, would improve pocket depth, clinical attachment level, or radiographic results of
filling the infraosseous defect, due to the high risk of bias in performing their revision [50].
APCs have also been used in the treatment of furcation defects, as shown in the re-
view by Panda et al. [51], achieving an improvement in horizontal and vertical clinical
attachment loss and furcation depth, in addition to open surgery and bone grafting, but
not with GTR and enamel matrix derivatives. Furthermore, again regarding the treatment
of furcation defects, the study by Tarallo et al. [46] demonstrated the effectiveness of the
additional use of PRF to open periodontal surgery to improve periodontal regeneration,
particularly in the presence of grade II furcation.
In addition to regenerative properties, it has been shown in other reviews by Del
Fabbro et al. [52,53] that platelet concentrates provide additional benefits during post-
surgical healing, such as reduced pain and improved quality of life for patients.
Recently, the clinical efficacy of PRGF in addition to non-surgical periodontal ther-
apy, such as scaling and root planing (SRP) in patients with periodontitis was evaluated
by Panda et al. [51]; the authors concluded that the use of PRGF proved effective in both
reducing pocket depth and bleeding index and gaining clinical attachment level.
L-PRF has also been used in the treatment of single and multiple gingival recessions
in conjunction with the coronal repositioning flap (CAF) technique; the efficacy of this,
compared with CAF alone and with the addition of connective graft, was the subject of
the study by Mancini et al. [47], who concluded their review by reporting that there were
statistically significant results in mean root coverage, reduction in recession, gain in
Biomedicines 2022, 10, 218 12 of 14

keratinized tissue width, gain in gingival thickness, patient perception of pain and dis-
comfort with the use of L-PRF compared to CAF alone, but not in relation to the addition
of connective graft in single and multiple CAF. The latter, however, showed statistically
significant results only in relation to the patient’s perception of pain and discomfort. In
recent years, PRP has also gained interest in regenerative endodontics, particularly in
pulpotomy and apical surgery, being considered an ideal scaffold in this field. Indeed,
autologous PRP acts as an ideal natural three-dimensional scaffold, supporting cell
growth and differentiation and releasing various growth factors that influence tissue re-
generation. The study by Sequeira et al. [48] evaluated the potential of using human apical
papilla stem cells (SCAP) embedded in a PRP scaffold for endodontic regenerative proce-
dures, in combination with ProRoot MTA or Biodentine. The results of this study pro-
vided evidence of de novo pulp and dentin-like tissue formation when SCAP and PRP
were applied in the empty root canal space of root portions filled with cultured cells after
they were isolated from apical papillae and implanted in the subcutaneous space of im-
munodeficient rats [48]. The results derived from the study are probably due to the bio-
physical properties of the existing dentin interacting with SCAP-derived odontoblasts or
the increased concentration of bioactive molecules on the root dentin wall [48]. In addition
to the presence of pulp tissue, blood vessels, and nerve fibers, a typical layer of odonto-
blast-like cells could also be observed in this study’s pulpal part of the new dentin tissue
[48].
Finally, although no specific clinical studies have been conducted to date, there is
substantial preclinical evidence of the antimicrobial effect of autologous platelet concen-
trates against certain species commonly found in the oral cavity, suggesting that they may
be a valuable tool for controlling the onset of post-surgical infection [54,55].

5. Conclusions
From the bone marrow to the wound through the vascular supply, platelets have a
crucial role in hemostasis, immune modulation, and repair mechanisms.
The described diseases affecting platelets production and quality, with also severe
consequences on the survival and patients’ quality of life, prove the unicity and the indis-
pensability of these small elements and the necessity of an accurate medical history also
in cases of dental surgery procedures.
In addition, the secretive property of growth factors and of fibrin network, whose
physical characteristics can vary and be artificially controlled, made them an important
source in regenerative medicine.
The fibrin network and the growth factors give to the platelets concentrates inductive
and conductive properties and, therefore, a potential aid in regenerative bone procedures.

Author Contributions: Conceptualization, S.B. (Sara Bernardi); validation, S.B. (Serena Bianchi),
G.V., and M.P.; investigation, F.R. and D.T.; writing—original draft preparation, S.B. (Sara Ber-
nardi); writing—review and editing, G.V., supervision, S.B. (Serena Bianchi), G.V., and M.P. All
authors have read and agreed to the published version of the manuscript.
Funding: This research received no external funding.
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: Not applicable.
Conflicts of Interest: The authors declare no conflict of interest.

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