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Nicola Maffulli (Eds.) - Platelet Rich Plasma in Musculoskeletal Practice-Springer-Verlag London (2016) PDF
Nicola Maffulli (Eds.) - Platelet Rich Plasma in Musculoskeletal Practice-Springer-Verlag London (2016) PDF
Editor
123
Platelet Rich Plasma
in Musculoskeletal Practice
Nicola Maffulli
Editor
vii
Contents
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 241
Chapter 1
Contents and Formulations
of Platelet Rich Plasma
Amy S. Wasterlain, Hillary J. Braun, and Jason L. Dragoo
Platelets
Leukocytes
a b
c d
Fibrin
Platelet-Rich Formulations
Two main issues muddy the current PRP literature. First,
there is little consensus regarding the exact constituents of
PRP; the only definition consistently upheld in the literature
10 A.S. Wasterlain et al.
PRP Gel
GenesisCS EmCyte 60a 4–5 [73] 12–16a [73] + 10a 68–96 %a 5a P4-Aα 1A
[74]
GPS III Biomet 60a 6a 27 [73] + + 2.1–8.1 [7, 23–90 %a 4.3–5 [7, P2-Aα 1A/B,
A.S. Wasterlain et al.
Magellan Arteriocyte/ 60 [72] 1–10 [73] 17–22 [72, + 2.8–7.1a 66 % [45] 2–2.9 [45, P2-Aα 1A/B
Medtronic 73] [45, 72] 72]
SmartPReP 2 Harvest 60 [72] 10 [73] 15–17a [72] + 6–8.2a [72] 62–72 % 3.5–4a [72] P2-Aα 1A
[65, 67,
69]
Leukocyte-Poor PRP
Cascade MTF/ 18 [45] 7.5 [45] 21 [73] + 1.6 [45] 68 % [45] 0.2 [45] P2-Bβ 3B or
CONMED 4B
PRGF- BTI 9a 2–4 [75] 28a + 2–3a P2-Bβ 3B or
Endoret 4B
PRP gel
AutoloGel Cytomedix + 78 %a
platelet-poor plasma, PRGF plasma rich in growth factors, PRP platelet-rich plasma, LR-PRP leukocyte-rich platelet-
rich plasma
15
16 A.S. Wasterlain et al.
Activation Procedures
The term “activation” refers to two key processes within PRP
preparations that may be initiated: (1) degranulation of plate-
lets to release α-granules containing growth factors and (2)
fibrinogen cleavage to initiate matrix formation. The bulk of
current literature simply indicates the means by which gen-
eral PRP activation occurs but rarely specifies which cellular
components are targeted by these techniques. The addition of
thrombin and calcium chloride, or collagen are both effective
in activating both platelets and fibrinogen, while activation
via freeze/thaw cycles initiates degranulation only.
Accordingly, rapid platelet activation can be achieved by the
following three mechanisms: (1) addition of calcium chloride
and thrombin, (2) freeze/thaw cycles, and (3) direct exposure
to collagen in vivo. Once activated, the PRP composition is
often referred to as the PRP releasate.
Chapter 1. Contents and Formulations 17
Fibroblasts
Collagen
Tenocytes
Systemic Effects
Conclusion
References
1. Rozman P, Bolta Z. Use of platelet growth factors in treating
wounds and soft-tissue injuries. Acta Dermatoven APA.
2007;16(4):156–65.
2. Rehfeldt F, Engler AJ, Eckhardt A, Ahmed F, Discher DE. Cell
responses to the mechanochemical microenvironment – implica-
tions for regenerative medicine and drug delivery. Adv Drug
Deliv Rev. 2007;59(13):1329–39.
3. Foster TE, Puskas BL, Mandelbaum BR, Gerhardt MB, Rodeo
SA. Platelet-rich plasma: from basic science to clinical applica-
tions. Am J Sports Med. 2009;37(11):2259–72.
4. Marx RE. Platelet-rich plasma (PRP): what is PRP and what is
not PRP? Implant Dent. 2001;10(4):225–8.
5. Creaney L, Hamilton B. Growth factor delivery methods in the
management of sports injuries: the state of play. Br J Sports Med.
2008;42(5):314–20.
6. Anitua E, Sanchez M, Zalduendo MM, de la Fuente M, Prado R,
Orive G, et al. Fibroblastic response to treatment with different
preparations rich in growth factors. Cell Prolif. 2009;42(2):162–70.
7. Sundman EA, Cole BJ, Fortier LA. Growth Factor and Catabolic
Cytokine Concentrations Are Influenced by the Cellular
Composition of Platelet-Rich Plasma. Am J Sports Med.
2011;39(10):2135–40.
Chapter 1. Contents and Formulations 23
39. de Mos M, van der Windt AE, Jahr H, van Schie HT, Weinans H,
Verhaar JA, et al. Can platelet-rich plasma enhance tendon
repair? A cell culture study. Am J Sports Med.
2008;36(6):1171–8.
40. Anitua E, Andia I, Ardanza B, Nurden P, Nurden AT. Autologous
platelets as a source of proteins for healing and tissue regenera-
tion. Thromb Haemost. 2004;91(1):4–15.
41. Min JK, Lee YM, Kim JH, Kim YM, Kim SW, Lee SY, et al.
Hepatocyte growth factor suppresses vascular endothelial
growth factor-induced expression of endothelial ICAM-1 and
VCAM-1 by inhibiting the nuclear factor-kappaB pathway. Circ
Res. 2005;96(3):300–7.
42. He L, Lin Y, Hu X, Zhang Y, Wu H. A comparative study of
platelet-rich fibrin (PRF) and platelet-rich plasma (PRP) on the
effect of proliferation and differentiation of rat osteoblasts
in vitro. Oral Surg Oral Med Oral Pathol Oral Radiol Endod.
2009;108(5):707–13.
43. Marx RE, Carlson ER, Eichstaedt RM, Schimmele SR, Strauss
JE, Georgeff KR. Platelet-rich plasma: growth factor enhance-
ment for bone grafts. Oral Surg Oral Med Oral Pathol Oral
Radiol Endod. 1998;85(6):638–46.
44. Weibrich G, Kleis WK, Hafner G, Hitzler WE. Growth factor
levels in platelet-rich plasma and correlations with donor age,
sex, and platelet count. J Craniomaxillofac Surg.
2002;30(2):97–102.
45. Castillo TN, Pouliot MA, Kim HJ, Dragoo JL. Comparison of
growth factor and platelet concentration from commercial
platelet-rich plasma separation systems. Am J Sports Med.
2011;39(2):266–71.
46. Dragoo JL, Braun HJ, Durham JL, Ridley BA, Odegaard JI,
Luong R, Arnoczky SP. Comparison of the acute inflammatory
response of two commercial platelet-rich plasma systems in
healthy rabbit tendons. Am J Sports Med. 2012 Jun;40(6):1274–
81. doi: 10.1177/0363546512442334. Epub 2012 Apr 10. PMID:
22495144.
47. Felisaz N, Boumediene K, Ghayor C, Herrouin JF, Bogdanowicz
P, Galerra P, et al. Stimulating effect of diacerein on TGF-beta1
and beta2 expression in articular chondrocytes cultured with and
without interleukin-1. Osteoarthritis Cartilage. 1999;7(3):255–64.
48. Roman-Blas JA, Stokes DG, Jimenez SA. Modulation of TGF-
beta signaling by proinflammatory cytokines in articular chon-
drocytes. Osteoarthritis Cartilage. 2007;15(12):1367–77.
Chapter 1. Contents and Formulations 27
Summary
For the past 20 years, autologous Platelet-Rich Plasma (PRP)
has been safely employed and its use has been documented in
many areas, including orthopedics, sports medicine, dentistry,
Introduction
Platelet-rich plasma (PRP) is an autologous biomaterial
obtained by centrifuging whole blood. PRP may be defined
as a fraction of autologous plasma with platelet concentration
above baseline level. Studies have shown that ideal concen-
tration is at least a fourfold increase in initial concentration,
or around 1,000,000 mm3 [1].
Platelets are responsible for hemostasis promotion, new
connective tissue formation and revascularization. A sample
of whole blood usually contains 93 % red blood cells, 6 %
platelets and 1 % white cells (leukocytes).
Justification for the benefits of PRP rests on inverting the
proportion of these cells in the blood, reducing the red layer
to 5 %, since red blood cells are less useful in the healing
process, and increasing platelets and leukocytes to 94 % to
stimulate tissue regeneration [1].
2 Platelet-Rich Plasma in Pain Medicine 33
Other benefits that justify PRP use are the diverse growth
factors contained in the concentrate, such as transforming
growth factor-beta (TGF-β), platelet-derived growth factor
(PDGF), basic fibroblast growth factor (bFGF), insulin-like
growth factor-l (IGF-l), vascular endothelial growth factor
(VEGF), and epidermal growth factor (EGF) [2–4]. There
have been in vitro and in vivo reports of the effects of PRP
and growth factors on the stimulation of fibroblast prolifera-
tion by IGF-l, bFGF, and PDGF and of collagen synthesis and
extracellular matrix synthesis by TGF-β in tendons and liga-
ments [5–8], resulting in enhanced regeneration and increased
tissue strength and consistency.
PRP Processing
PRP processing involves the separation and concentration of
platelets, leukocytes, and growth factors, considered initiators
in any repair process. However, there is no standard tech-
nique for obtaining PRP, and different preparations are
described in the literature. There is wide variability in the
ability to concentrate these cells and, for the most part, this
variability relies on equipment, manufacturer and materials
utilized.
Several authors have presented their proposals for classi-
fying and suggestions for standardizing the infinite number of
terms observed in current studies. These classifications are
directly related to the cell composition of each concentrate.
Mishra [9] classifies PRP on the basis of the presence or
absence of leukocytes, on the utilization of activating agents
and on final platelet concentration, enabling a framework
wherein products are divided into four types: type 1, high
leukocyte concentration, non-activated; type 2, high leuko-
cyte concentration, activated; type 3, non-activated, no leuko-
cytes or low leukocyte concentration; and type 4, activated, no
leukocytes or low leukocyte concentration. All these product
types may be further classified as A, corresponding to platelet
concentration equal to or five times higher than the baseline
34 J.F.S.D. Lana et al.
Injection Technique
Muscle
Cartilage
The social impact of degenerative diseases such as joint carti-
lage conditions is increasing, and it is a consequence of the rise
in the average age of the active population [65, 66]. Joint car-
tilage injuries have limited potential for repair, they are diffi-
cult to treat and remain a problem for doctors and orthopedic
surgeons. Capacity for cartilage regeneration is limited
because of its isolation from systemic regulation and because
of vascular deficiency [67–69]. Unlike the majority of tissues,
in an inflammatory process, chondrocytes do not migrate to
the joint injury from a healthy, intact site [41, 43]. Biomechanical,
metabolic, and biological changes, as well as trauma and iso-
lated chondral injuries, may lead to loss of tissue homeostasis,
resulting in faster degeneration of joint surfaces, leading to
cartilage injury or osteoarthritis (OA). These pathologies are
generally known to be crippling diseases, for they are con-
stantly associated to severe pain and mobility difficulties.
2 Platelet-Rich Plasma in Pain Medicine 47
Spine
Spinal pain includes all painful conditions originating
from the spine, whether from intervertebral disks, mus-
cles, ligaments, joints or bones. The structures responsible
for spinal pain are the vertebrae themselves, interverte-
bral discs, spinal cord, nerve roots, facet joints, muscles
and ligaments 1-10. It is a public health problem in
Western industrialized societies. Prevalence rates range
from 12 to 35 % and around 10 % of these patients
become chronically disabled. In the U.S.A., 1.5–4 million
adults suffer from lumbar pain secondary to spinal degen-
eration for whom conservative treatment was ineffective,
and many of these patients eventually undergo surgical
procedures [77].
Back pain is closely related to intervertebral disk degen-
eration (IDD). This condition, though asymptomatic in
many cases, is also associated with sciatica and interverte-
bral disk herniations or prolapses. Studies have shown that
discogenic pain is the most common cause of chronic back
pain, with rates ranging from 30 to 60 % in all cases. IDD
alters disk height and the resting mechanisms of the spinal
column, possibly adversely affecting the behavior of other
spinal structures, such as muscles and ligaments. In the long
run, it may lead to spinal stenosis, an important cause of
pain and disability in the elderly; its incidence is increasing
exponentially with current demographic changes and
increase in life expectancy.
Disks degenerate earlier than any other musculoskeletal
tissue structures. There is a significant reduction in end-plate
50 J.F.S.D. Lana et al.
Cell Therapies
Conclusion
Platelet-rich Plasma emerges as an autologous, non-
immunogenic, therapeutic option capable of stimulating the
supraphysiological release of cells responsible for wound
healing, with the aim of augmenting healing potential, medi-
ating inflammatory processes and relieving pain. The fre-
quently reported reduction in pain and in healing time
renders the PRP technique an important therapeutic tool in
interventional pain treatment. Correct association with
adjunctive therapies is also related to a good pain prognosis.
It is important to remember that being an autologous bioma-
terial, the success of PRP therapy depends on the patient’s
general clinical conditions. The present literature reports
2 Platelet-Rich Plasma in Pain Medicine 53
References
1. Marx R, Garg A. Dental and craniofacial applications of
platelet-rich plasma. Chicago: Quintessence Publishing Co, Inc;
2005.
2. Landesberg R, Roy M, Glickman RS. Quantification of growth
factor levels using a simplified method of platelet-rich plasma
gel preparation. J Oral Maxillofac Surg. 2000;58(3):297–300; dis-
cussion 00-1.
3. Weibrich G, Kleis WK, Hafner G, Hitzler WE. Growth factor
levels in platelet-rich plasma and correlations with donor age,
sex, and platelet count. J Craniomaxillofac Surg. 2002;30(2):
97–102.
4. Eppley BL, Woodell JE, Higgins J. Platelet quantification and
growth factor analysis from platelet-rich plasma: implications for
wound healing. Plast Reconstr Surg. 2004;114(6):1502–8.
5. Molloy T, Wang Y, Murrell G. The roles of growth factors in ten-
don and ligament healing. Sports Med. 2003;33(5):381–94.
6. Abrahamsson SO, Lundborg G, Lohmander LS. Long-term
explant culture of rabbit flexor tendon: effects of recombinant
human insulin-like growth factor-I and serum on matrix metabo-
lism. J Orthop Res. 1991;9(4):503–15.
7. Kurtz CA, Loebig TG, Anderson DD, DeMeo PJ, Campbell
PG. Insulin-like growth factor I accelerates functional recovery
from Achilles tendon injury in a rat model. Am J Sports Med.
1999;27(3):363–9.
8. Chan BP, Fu S, Qin L, Lee K, Rolf CG, Chan K. Effects of basic
fibroblast growth factor (bFGF) on early stages of tendon heal-
ing: a rat patellar tendon model. Acta Orthop Scand.
2000;71(5):513–8.
9. Mishra A. Platelet-rich plasma. Orthopedics. 2010;33(7):486–7.
54 J.F.S.D. Lana et al.
10. Everts PA, van Zundert A, Schonberger JP, Devilee RJ, Knape
JT. What do we use: platelet-rich plasma or platelet-leukocyte
gel? J Biomed Mater Res A. 2008;85(4):1135–6.
11. Anitua E, Sanchez M, Orive G, Andia I. Shedding light in the
controversial terminology for platelet rich products. J Biomed
Mater Res A. 2009;90(4):1262–3.
12. Dohan Ehrenfest DM, Bielecki T, Del Corso M, Inchingolo F,
Sammartino G. Shedding light in the controversial terminology
for platelet-rich products: platelet-rich plasma (PRP), platelet-
rich fibrin (PRF), platelet-leukocyte gel (PLG), preparation rich
in growth factors (PRGF), classification and commercialism.
J Biomed Mater Res A. 2010;95(4):1280–2.
13. Anitua E. Plasma rich in growth factors: preliminary results of
use in the preparation of future sites for implants. Int J Oral
Maxillofac Implants. 1999;14(4):529–35.
14. Everts PA, Overdevest EP, Jakimowicz JJ, Oosterbos CJ,
Schonberger JP, Knape JT, et al. The use of autologous platelet-
leukocyte gels to enhance the healing process in surgery, a
review. Surg Endosc. 2007;21(11):2063–8.
15. Moojen DJ, Everts PA, Schure RM, Overdevest EP, van Zundert
A, Knape JT, et al. Antimicrobial activity of platelet-leukocyte
gel against Staphylococcus aureus. J Orthop Res.
2008;26(3):404–10.
16. Scott A, Khan KM, Roberts CR, Cook JL, Duronio V. What do
we mean by the term “inflammation”? A contemporary basic
science update for sports medicine. Br J Sports Med.
2004;38(3):372–80.
17. Toumi H, Best TM. The inflammatory response: friend or enemy
for muscle injury? Br J Sports Med. 2003;37(4):284–6.
18. Sundman EA, Cole BJ, Fortier LA. Growth factor and catabolic
cytokine concentrations are influenced by the cellular composition
of platelet-rich plasma. Am J Sports Med. 2011;39(10):2135–40.
19. Dohan Ehrenfest DM, Bielecki T, Jimbo R, Barbe G, Del Corso
M, Inchingolo F, et al. Do the fibrin architecture and leukocyte
content influence the growth factor release of platelet concen-
trates? An evidence-based answer comparing a pure platelet-
rich plasma (P-PRP) gel and a leukocyte- and platelet-rich fibrin
(L-PRF). Curr Pharm Biotechnol. 2012;13(7):1145–52.
20. Smith C, Kruger MJ, Smith RM, Myburgh KH. The inflamma-
tory response to skeletal muscle injury: illuminating complexi-
ties. Sports Med. 2008;38(11):947–69.
2 Platelet-Rich Plasma in Pain Medicine 55
Introduction
Common chronic musculoskeletal injuries, including tendi-
nopathies, cartilage disorders, and spine disease are difficult
to manage. Their treatments are often only palliative, aiming
to reduce pain. Traditional interventions include non-steroi-
dal anti-inflammatory medications, corticosteroid injections,
S. Sampson, DO ()
The Orthohealing Center and The Orthobiologic Institute (TOBI),
David Geffen School of Medicine at UCLA, Los Angeles, CA, USA
e-mail: drsampson@orthohealing.com
K. Mautner, MD
Rehabilitation Medicine, Orthopaedics and Sports Medicine,
Emory University, Atlanta, GA, USA
A.G. Via, MD
Department of Orthopaedic and Traumatology, University of
Rome “Tor Vergata”, School of Medicine, Viale Oxford 81,
Rome 00133, Italy
A.B.-v. Bemden, PhD, ATC, CSCS
Musculoskeletal Research International (MRI), Clinical Research
Experts (CRE), Florida International University, Miami, FL, USA
PRP Preparation
Many PRP formulations are available for clinical uses, yet the
products may vary greatly, and this may influence the efficacy
of treatment and the results of clinical trials. The presence of
WBCs is largely dependent on the centrifugation and tech-
nique used to process the PRP. The role of leukocytes in PRP
(L-PRP) is controversial [5]. Those in favor of excluding WBCs
argue that neutrophils may have a detrimental effect on muscle
and bone [6, 7]. Furthermore, neutrophils can release degrada-
tive matrix metalloproteinase (MMP)-3,8,9 and 13, and free
radicals [6]. This could lead to a delayed healing response in
muscle [7]. Regarding bone healing, mice with temporary neu-
tropenia and femur fractures healed the fracture with a higher
bending moment of the bone callus compared to the non-neu-
tropenic mice [8].
66 S. Sampson et al.
Current Applications
Tendinopathy
Ligament Pathology
Muscle Pathology
Cartilage
Proliferative phase
Inflammatory phase Maturation phase
• Collagen
accumulation
• Remodeling
Granulation Wound
Inflammation
tissue contraction
[64]. Phase III begins around the sixth week. During this
phase, new extracellular matrix is laid down primarily through
accumulation of Type I collagen, the foundation of healthy
tendons [64]. This phase lasts for several months, even up to
a year. Thus, measuring outcomes from PRP injections should
require adequate time for healing to occur.
Based on the above framework, rehabilitation following
PRP injections can be divided into three phases as well
(Fig. 3.7). The first is the acute phase immediately after a PRP
injection is performed. The principal treatments of this phase
are pain control and tissue protection. It is recommended to
minimize excessive motion of the involved area and allowing
local platelet activation while avoiding disruption of the
fibrin plug during this period. This can range from limiting
weight bearing or resistance to frank immobilization. There
are no studies to demonstrate that immobilization enhances
80 S. Sampson et al.
Expert Opinion
Among the earliest users of PRP in musculoskeletal disease,
the authors have seen good outcomes in challenging condi-
tions. However, PRP is not a first line option for most
patients, and should generally be considered when traditional
measures fail. However, every day our clinics are filled with
patients who are told they have no conservative options
available and must “live with the pain.” These patients, which
84 S. Sampson et al.
stem cell based therapies offer great promise, its research and
practical applications are in their infancy.
Conclusions
Because of its autologous status, ease of use and preparation,
and given its safety profile, PRP is gaining popularity and is
introducing physicians and the community to the realm of
biological therapies. Most likely, PRP use will persist for years
to come. However, it may be recommended for particular
diagnoses or may serve as an adjunct for other autologous cell
based therapies such as Bone Marrow Concentrate (BMC),
Fat Grafts or Adipose Derived Stem Cells. Much research is
still needed to elucidate the optimal concentration, need for
activation, leukocyte and RBC presence, timing and fre-
quency of injections, specific clinical indications, post injec-
tion rehabilitation, treatment adjuncts, and the role of
nutrition, hormone optimization, medical history and psycho-
logical factors. The authors have had good outcome using
PRP in thousands of patients while continually exploring new
ways to improve outcomes. Growing worldwide collaboration
with controlled trials is necessary to advance our understand-
ing of musculoskeletal disease, and to institute novel biologi-
cal therapies to maximize healing. Although challenging to
conduct robust trials in a clinical setting, practitioners should
collaborate with colleagues, analyze and share data and pub-
lish work to further establish and advance biologic medicine.
References
1. Troiano M, Schoenhaus H. A closer look at orthobiologics for
tendon repair. Podiatry Today. 2009;22(10).
2. Roback J, Combs M, Grossman B, Hillyer C. Technical Manual
of the American Association of Blood Banks. 16th ed. Bethesda:
American Association of Blood Banks (AABB); 2008.
3. Alsousou J, Thompson M, Hulley P, Noble A, Willet K. The biol-
ogy of platelet-rich plasma and its application in trauma and
86 S. Sampson et al.
Introduction
Though highly debated, platelet rich plasma (PRP) has been
increasingly used in all the fields of orthopaedic and trauma
surgery. Its mechanism of action is still unclear, and no clear
evidences of its effectiveness are available. PRP is supposed
to act as a biological healing enhancer, and is considered to
contain platelets and, in some formulations, white blood cells.
Platelets do not have nucleus but they contain small granules
rich in growth factors (GF) responsible for hemostasis and
Tendons
The incidence of tendinopathies is rising up since the increas-
ing popularity of playing sports among the general popula-
tion and also among sedentary individuals. Chronic
tendinopathy is a major problem, since the healing process
often produces a painful tissue with abnormal histological
features. Conservative measures are the main treatment
option for tendinopathies. Eccentric exercises, Extracorporeal
Shock Wave Therapy, peritendinous injections produce vari-
able outcomes. Surgery is considered the last available option
when nonoperative management has failed, though even the
outcomes of surgery are unpredictable. Tendinopathies are
supposed to result from an unbalance between external
stimuli and adaptive changes. Traditional treatments are
often unpredictable; thus, great efforts have been made to
enhance the healing process. Studies on animal models
showed that PRP is able to lower the COX-1 and COX-2
expression, as well as PGE2, which is involved in pain produc-
tion, decreased cell proliferation, and collagen production
[40]. PRP is able to increase cell number, pro- mote progeni-
tor cells differentiation and increase collagen fiber density,
thus restoring the normal tendon tissue architecture [3, 10, 33,
35, 38]. Clinical use of PRP is widespread, though its effec-
tiveness is still controversial. A recent meta-analysis on con-
trolled studies pointed out that there are no evidences
supporting or hindering the clinical use of PRP. Pooling data
by sites of tendinopathy showed mid-term (36 months) ben-
eficial effects on pain, comparing L-PRP (leukocite rich
PRP) to other existing methods [1].
Chapter 4. PRP in Tendons and Other Non-bone Tissues 95
Achilles Tendinopathy
Patellar Tendinopathy
Lateral Epicondylitis
Rotator Cuff
Plantar Fasciopathy
Knee Osteoarthritis
Knee osteoarthritis is disabling. Current available conserva-
tive treatments include physical therapy, oral drugs and intra-
articular injections, with variable effectiveness. Recently,
intra-articular PRP injections have been widely used, though
good quality level I evidence is lacking.
Four level I RCTs are currently available.
Cerza et al. [9] reported on 120 patients randomized into
two groups: 60 patients received 4 intra-articular injections of
PRP, and 60 patients received 4 intra-articular injections of
Hyaluronic Acid (HA) (20 mg/2 mL). Treatment with PRP
showed a significantly better clinical outcome than did treat-
ment with HA, with lower WOMAC scores up to 24 week
followup.
Patel et al. [28] in a double-blind RCT on a total of 78
patients (156 knees) with bilateral knee osteoarthritis were
randomized into 3 groups. Group A (52 knees) received a
single injection of PRP, group B (50 knees) received 2 injec-
tions of PRP 3 weeks apart, and group C (46 knees) received
a single injection of normal saline. At 6 month, the two
groups treated with PRP injections showed significantly bet-
ter outcomes compared with the saline group. There were no
differences between the single versus multiple PRP injection
groups.
100 S. Vasta et al.
Conclusion
PRP preparations contain high concentrations of platelets
that, once activated, undergo degranulation to release growth
factors with healing properties. PRP also contains plasma and
other growth factors implicated in wound healing, in addition
to thrombin, which has inherent biological and adhesive
properties [2]. PRP has an anti-inflammatory properties. In
addition, the GF contained in PRP can stimulate the healing
processes by acting as a scaffold, and thus attracting sur-
rounding cells [38].
Though it has been receiving increasing emphasis and its
use has been increasingly expanding in clinical practice,strong
evidences supporting the use of PRP have not been produced
yet. Many variables play a primary role in PRP effectiveness.
Some are related to PRP itself, and others to the various clini-
cal settings. Among the first, there are different preparation
methods, thus influencing the amount of platelets, of white
blood cells, and the presence of molecules that have a role in
activating platelets (e.g. thrombin). Moreover, PRP can be
administered as an injectable fluid or as a implantable fibrin-
containing patch or gel.
With regard to clinical setting, there are a number of fac-
tors that vary from one investigation to another. Features of
the patients (age, level of activity), previous treatments, num-
ber of PRP injection, post-treatment rehabilitation are some
of the possible variables.
Chapter 4. PRP in Tendons and Other Non-bone Tissues 101
References
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THRO04010004.
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Arthroscopy Assoc North Am Int Arthroscopy Assoc.
2011;27(8):1029–35. doi:10.1016/j.arthro.2011.06.010.
6. Bergeson AG, Tashjian RZ, Greis PE, Crim J, Stoddard GJ,
Burks RT. Effects of platelet-rich fibrin matrix on repair integ-
rity of at-risk rotator cuff tears. Am J Sports Med. 2012;40(2):286–
93. doi:10.1177/0363546511424402.
7. Boileau P, Brassart N, Watkinson DJ, Carles M, Hatzidakis AM,
Krishnan SG. Arthroscopic repair of full-thickness tears of the
supraspinatus: does the tendon really heal? J Bone Joint Surg
Am. 2005;87(6):1229–40. doi:10.2106/JBJS.D.02035.
8. Castricini R, Longo UG, De Benedetto M, Panfoli N, Pirani P,
Zini R, Maffulli N, Denaro V. Platelet-rich plasma aug-
102 S. Vasta et al.
Biological Rationale
The biological rationale behind PRP treatment is the topical
administration of platelet-derived GFs, a group of polypep-
tides normally involved in joint homeostasis, healing and
regenerating of the damaged tissue, including cartilage [16].
Platelets are traditionally known for their hemostatic and
coagulation functions, but they also play a key role in the heal-
ing mechanism of damaged tissues, primarily thanks to their
GFs. Platelets contain three types of granules: lisosomal gran-
ules, dense granules, and alpha granules. Alpha granules are
the source of GFs, including platelet-derived growth factor
(PDGF), transforming growth factor (TGF-β), platelet-derived
epidermal growth factor (PDEGF), vascular endothelial
growth factor (VEGF), insulin-like growth factor 1 (IGF-1),
fibroblastic growth factor (FGF), and epidermal growth factor
(EGF); they also contain cytokines and chemokines, which are
involved in stimulating chemotaxis, cell proliferation and mat-
uration, modulating inflammatory molecules and attracting
leukocytes [10]. Dense granules store ADP, ATP, calcium ions,
histamine, serotonin and dopamine, which also play a complex
role in tissue modulation and regeneration [10, 17]. Finally,
lisosomial granules contain acid hydrolases, cathepsin D and E,
elastases and lisozyme [18], and many other proteins whose
physiological role is not well characterized. Furthermore, the
plasma itself contains important molecules involved in the
healing mechanism of connective tissues, also contributing to
the platelet stimulus in tissue regeneration.
5 Platelet Rich Plasma in Articular Cartilage Lesions 111
Knee Application
(ACP) – with HA; 120 patients were divided into two groups,
and underwent 4 weekly injections. After 6 months’ follow-
up, the ACP group showed a significantly better performance
than that of the HA group, with clinical superiority even in
patients with Ahlback grade 3 knee OA. A double-blinded
randomized controlled trial comparing leukocyte-rich PRP
and HA has recently been published by Filardo et al. [35],
who reported the preliminary results of the first 109 patients
treated (55 PRP and 54 HA) and followed up for 12 months.
The evaluation was performed by IKDC objective and sub-
jective, Tegner score, KOOS score and VAS for general
health status. A significant increase in clinical scores was
observed in both treatment groups, with no statistical inter-
group difference reported. A tendency towards better results
was found only in the subgroup of patients with low grade
cartilage degeneration (Kellgren Lawrence up to 2).
Therefore, the authors concluded that PRP should be limited
to early OA, but it should not be considered as a first line
treatment in moderate/severe OA.
Patel et al. [36] studied 78 patients with bilateral OA
(Ahlback grade up to 2). Both knees of each patient were
included in the study and three treatment groups were identi-
fied: the first one received a single injection of leukocyte-free
PRP, the second group 2 PRP injections, whereas the last
group received one injection of normosaline. Follow-up
evaluation was performed at 6 months: a superior clinical
outcome was evident for PRP-treated knees, with no signifi-
cant difference between one or two PRP injections. Moreover,
even in this study, low-grade degenerated knees (Ahlback
grade 1) presented far better results than those of more
severely degenerated knees.
Hip Applications
Discussion
The clinical application of PRP is supported by increasing
interest and evidence, but debate is still inconclusive because
of the lack of robust clinical trials [42]. A critical review of the
available literature allows to gain a better understanding of
the potential and feasibility of applying PRP in the manage-
ment of cartilage pathology. Concerning surgical application,
it is not possible to draw definite conclusions about the effi-
cacy of this treatment. In many surgical trials, PRP has been
administrated as an augmentation, together with mesenchy-
mal stem cells [22, 39, 40] or scaffold [20–22]. Therefore, it is
not possible to understand the role of PRP in determining the
clinical outcome. Moreover, the studies available are often
just case series treating disparate conditions in biomechani-
cally very different joints. Further studies are needed to define
guidelines for using PRP in such procedures and to determine
the persistence of the effect of PRP, since at present the lon-
gest reported follow-up is only 24 months. Moreover, PRP
augmentation of hydoxyapatite-collagen scaffold has a nega-
tive effect on its ability to repair cartilage [43].
With regards to the conservative use of PRP for hip and
talar osteochodral lesions, few studies have been published.
Two case series [37, 38] deal with management of hip OA, and
confirmed the safety of PRP and its usefulness in providing
temporary symptomatic relief. However, given the lack of
randomized trials, there is no evidence to support PRP as a
first line treatment in degenerative pathology of the hip joint.
Concerning talar osteochondral lesions, the only comparative
quasi-randomized trial published [38] showed statistically
5 Platelet Rich Plasma in Articular Cartilage Lesions 117
superior results for the PRP group. However, the low number
of patients treated and the short follow-up evaluation do not
allow, even in this case, the use of PRP to be fully endorsed.
Conversely, with regards to PRP application as conserva-
tive management of OA of the knee, several trials are now
available. Musch still needs to be clarified, but the consider-
able amount of studies published allows us to draw some
conclusions [44, 45]. Firstly, PRP is safe, as confirmed by all
trials, which reported only minor adverse events, such as post-
injective swelling and pain. Looking at the clinical outcome,
trials confirmed the benefit of treatment, despite a gradual
worsening occurring over time. Filardo et al. [27] were the first
to calculate the average duration of the PRP effect, which is
estimated at about 9 months. Four randomized clinical trials
were recently published, three of them comparing PRP versus
HA and the last one against saline solution: although better
results have been found in the PRP group at 6 months’ follow-
up [33, 34], Filardo et al. [35] reported no overall difference
between PRP and HA in terms of clinical outcome at 12
months’ follow-up. Nevertheless, a tendency towards better
results was found in young patients with a lower degree of
cartilage degeneration, whereas moderate or severe OA show
less favorable outcomes without difference compared to vis-
cosupplementation: the response to PRP may therefore be
linked to the stage of the condition. No definitive conclusions
can be drawn about the possibility of applying this approach
to a specific phase of cartilage degenerative pathology, espe-
cially if we consider that, until now, patients treated in the
aforementioned randomized trials were not homogeneous in
terms of disease stages, showing from chondropathy to severe
OA. What emerges can be considered as just a “suggestion”,
i.e. to avoid the indiscriminate use of PRP, the application of
which should be reserved to patients who can obtain the best
results from PRP (young patients with less articular degenera-
tion), and only as second line treatment in case of no response
to other traditional conservative managements for the other
patients, taking care to inform them not to expect miracles
from this biological treatment.
118 E. Kon et al.
Conclusions
Many questions are still open concerning PRP treatment of
cartilage lesions. At present, no conclusive indications are
available about this topic because only a few and heteroge-
neous high quality trials have been published. The only aspect
to emerge with enough scientific evidence is the safety of the
procedure, but clear superiority compared to other traditional
treatments has not been fully proven [44]. Further studies
should allow to dispel these doubts, but the indiscriminate use
of PRP should be avoided. PRP should not be a first line treat-
ment for cartilage pathology, and should be used in patients
who, according to the current scientific evidence, can obtain
the best results from this approach. The best PRP formulation,
the best protocols of administration, and the profile of the
patient and lesion that can benefit more from this biological
therapy are aspects which remain to be clarified.
References
1. Brittberg M, Imhoff A, Madry H, Mandelbaum B. Cartilage
repair current concepts, ESSKA, 1st ed. DJO publication,
Guildford, UK. 2010. p. 3–10.
2. Falah M, Nierenberg G, Soudry M, Hayden M, Volpin
G. Treatment of articular cartilage lesions of the knee. Int
Orthop. 2010;34(5):621–30.
3. Gomoll AH, Filardo G, de Girolamo L, Espregueira-Mendes J,
Marcacci M, Rodkey WG, Steadman JR, Zaffagnini S, Kon
E. Surgical treatment for early osteoarthritis. Part I: cartilage
repair procedures. Knee Surg Sports Traumatol Arthrosc.
2012;20(3):450–66.
4. Gomoll AH, Filardo G, Almqvist FK, Bugbee WD, Jelic M,
Monllau JC, Puddu G, Rodkey WG, Verdonk P, Verdonk R,
Zaffagnini S, Marcacci M. Surgical treatment for early osteoar-
thritis. Part II: allografts and concurrent procedures. Knee Surg
Sports Traumatol Arthrosc. 2012;20(3):468–86.
5. Kon E, Filardo G, Drobnic M, Madry H, Jelic M, van Dijk N,
Della VS. Non-surgical management of early knee osteoarthritis.
Knee Surg Sports Traumatol Arthrosc. 2012;20(3):436–49.
5 Platelet Rich Plasma in Articular Cartilage Lesions 119
Introduction
Osteoarthritis (OA) is a degenerative disease of synovial
joints resulting from the combination of mechanical stress
and biochemical cellular changes which ultimately cause pain
and impair joint function. Osteoarthritis is the eighth leading
non-fatal burden of disease worldwide and a major cause of
disability [1]. The prevalence of symptomatic OA increases
with age: it is estimated that 18 % of women and 9.6 % of
men over age 60 have knee OA [1]. With increased longevity
in the population the burden and prevalence of OA is
expected to grow [2]. Furthermore, the epidemic of obesity
and resultant motivation to exercise, often through sports,
places more middle aged and older adults at risk of develop-
ing degenerative joint disease [3]. For the aging and former
athlete, OA may be a result of active adolescence and result
from prior ligamentous injury, meniscal trauma, or joint dis-
location. In a recent study, aging male athletes were 1.6 times
more likely to develop knee OA than controls [4].
Nociception in OA
Cartilage is aneural. However, the remainder of the knee
joint is highly innervated. The joint capsule, tendons, retinac-
ula, fat pads, synovium, subchondral bone, and ligaments
6 Platelet-Rich Plasma in Knee Osteoarthritis 127
contain type III and type IVa fibers that contribute to pain
generation in OA. Through the release of neuropeptides sub-
stance P and calcitonin gene-related peptide (CGRP), these
nerve endings respond to noxious stimuli in and around the
joint. Cadaveric immunohistochemical studies show increased
presence of substance P and CGRP in degenerative com-
pared to healthy knees [27]. Substance P has also been identi-
fied in the lateral retinaculum, a common site of anterior
knee pain in young adults [28]. Mechanoreceptors may also
provide nociceptive feedback. In a study of arthroscopy in
two unanesthetized patients, injecting fluid into the free
patellar space to increase pressure was associated with sig-
nificant pain [29]. Similarly, the presence of effusion and
increased effusion size are associated with worsening knee
pain [30]. The periosteum is also innervated by sensory and
sympathetic fibers that express CGRP and tropomyosin
receptor kinase A, respectively. These fibers become progres-
sively less dense in deeper structures such as mineralized
bone and bone marrow [31]. However, an MRI comparison
study in painful versus non-painful osteoarthritic knee joints
found pain to be associated with larger, more edematous, and
poorly delineated bone marrow lesions suggesting bone mar-
row as another pain generator [32].
Patient Selection
ITB
b
PT ST
SM
PP SA
BF
SA
MM
LCL LM GR
MCL
LCL **
* MCL
ITB
GR
LR
MR
ST PT
Methods
Methods for administration of PRP are variable and, to date,
there is no consensus on the frequency of injection, method of
PRP preparation, and injection technique. Thus, the following
methodology is based on clinical studies of PRP application.
6 Platelet-Rich Plasma in Knee Osteoarthritis 135
References
1. Murray CJL, Lopez A. The global burden of disease. A compre-
hensive assessment of mortality and disability from disease,
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2. Woolf AD, Pfleger B. Burden of major musculoskeletal condi-
tions. Bull World Health Organ. 2003;81(9):646–56.
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4. Tveit M, Rosengren BE, Nilsson JA, Karlsson MK. Former male
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5. McGarry JG, Daruwalla ZJ. The efficacy, accuracy and complica-
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6. Nichols AW. Complications associated with the use of corticoste-
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6 Platelet-Rich Plasma in Knee Osteoarthritis 141
Introduction
Basic Science
Several in vitro studies have investigated the effect of PRP on
bone healing (Table 7.2). Oprea et al. sought to determine the
effect of PRP on recruitment and migration of rat bone mar-
row derived cells within a fibrin matrix [36]. A fibrin gel over-
lay was added to cultured cells, and PRP applied to its surface.
PRP application increased the leading front of migration by
25 %, the number of migrating cells threefold, and cell prolif-
eration twofold. Additionally, these cells exerted a significant
contractile force on the fibrin matrix which was directly
related to the migratory activity of the cells. Kanno et al. stud-
ied the effect of PRP on osteoblast differentiation in human
osteosarcoma lines: it had both a dose-dependent and time-
dependent positive effects on the expression of alkaline phos-
phatase, procollagen type I, and osteopontin genes [37].
Kruger et al. studied the effect of PRP on human subcorti-
cal progenitor cells [38]. These cells are involved in cartilage
healing using the microfracture technique. They found
increased cell migration and cartilage matrix formation at the
site of microfracture. In addition, gene expression analysis of
typical chondrocyte, osteogenic and adipogenic markers
showed that PRP induced chondrogenic differentiation with-
out osteogenic or adipogenic differentiation, suggesting a
possible role in cartilaginous healing.
154 C. Cunningham et al.
Clinical Applications
One of the first reports of PRP use in the foot and ankle is by
Gandhi et al. [44], who studied 9 patients diagnosed with foot
or ankle fracture non-unions. Initial surgery occurred within
20 days of injury, and the diagnosis of non-union was formu-
lated 4–10 months later. Revision surgery consisted of PRP
and autologous bone graft applied locally to the site of non-
union. Ultimately, union was achieved in all patients within
60 days. Additionally, although plasma levels of PDGF and
TGF-b were consistent between groups, the levels in fracture
hematoma were significantly reduced between patients with
non-union compared to those with an acute fracture. The
authors concluded that the addition of PRP provided critical
early growth factors locally to the site of nonunion to aid the
healing process.
Chapter 7. Platelet Rich Plasma 157
after a year [50]. However, Schepull et al. did not use a fibrin
scaffold with PRP injections, raising the question of whether
its use significantly augmented the performance of PRP
therapy in the previous study.
The results of PRP use in chronic Achilles tendinopathy
have also been mixed. Gaweda et al. followed 14 patients
who received PRP injections following failure of other treat-
ment modalities [51]. They found a statistically significant
increase in functional outcome at 18 months follow-up as
well as improved tendon characteristics by ultrasound exam-
ination. At final evaluation, 2 patients reported persistent
functional limitations; their pain, however, was reduced from
initial presentation. Delos et al. and Monto et al. performed
similar studies that followed patients with chronic Achilles
tendinopathy that failed 6 months of conservative treatment
[52, 53]. Patients were treated with a single ultrasound
guided injection of PRP, and both studies reported signifi-
cant clinical improvement in most patients, and concluded
that PRP could be used safely and effectively to treat chronic
Achilles tendinosis. Sanchez et al. also showed that PRP was
able to improve patient symptoms and functionality when
used to treat post-surgical Achilles tendon complications
[54]. Conversely, in a level I study de Vos et al. found no sig-
nificant improvement in pain, activity, tendinous structural
organization, or degree of neovascularization compared to
placebo after treating 54 patients with a single ultrasound-
guided PRP injection [55, 56]. Similarly, Owens et al. reviewed
all patients treated with PRP injection for chronic Achilles
tendinopathy and found only modest improvement in func-
tional outcome measures with no changes in MRI appear-
ance of the diseased tendon [57].
Plantar fasciitis is also common in athletes, and several
recent studies have attempted to elucidate the role of PRP in
its treatment. Martinelli et al. treated 14 consecutive patients
with chronic plantar fasciitis with PRP injections at three dif-
ferent puncture sites [58]. Excellent results were reported in
9 of 12 patients, and pain scores were significantly reduced.
They concluded that PRP was safe and effective to use for the
Table 7.4 Summary of studies investigating the effect of PRP on soft tissue healing of the foot and ankle
160
Clinical applications
Sanchez et al. 2007 Open repair with PRP Acute Achilles Fewer complications, quicker return to
[49] and fibrin scaffold rupture normal
activities
Sanchez et al. 2009 Debridement followed Post-surgical Improved symptoms and functional
[54] with PRP complications outcome of both patients
Schepull et al. 2011 Open repair with PRP Acute Achilles No difference in mechanical variables
[50] injection rupture or
functional outcome
Gaweda et al. 2010 PRP injection Chronic Achilles Improved pain and tendon
[51] tendinopathy characteristics on
ultrasound
Delos et al. 2011 PRP injection Chronic Achilles Improved pain and tendon
[52] tendinopathy characteristics on
ultrasound
Montos et al. 2012 PRP injection Chronic Achilles Improved pain and tendon
[53] tendinopathy characteristics on
ultrasound
de Vos et al. 2011 PRP injection Chronic Achilles No change in tendon structure or
[55] tendinopathy vascularity on
ultrasound
de Vos et al. 2010 PRP injection Chronic Achilles No change in pain or functional level
[56] tendinopathy
Chapter 7.
Owens et al. 2011 PRP injection Chronic Achilles Modest improvement in function, no
[57] tendinopathy MRI changes
Martinelli et al. 2012 PRP injection Plantar fasciitis Decreased pain level
[58]
Ragab et al. 2012 PRP injection Plantar fasciitis Improved pain, decreased fascia
[59] thickness and
Platelet Rich Plasma
signal on ultrasound
Aksahin et al. 2012 PRP or corticosteroid Plantar fasciitis Improved pain, superior to
[60] injection corticosteroids
161
162 C. Cunningham et al.
Conclusion
Despite the encouraging results published by some foot and
ankle investigators, the authors of a recent meta-analysis of
studies comparing PRP with control therapy in orthopedic
Chapter 7. Platelet Rich Plasma 163
bone and soft tissue injuries concluded that, given the lack of
standardization of study protocols, platelet separation tech-
niques, and outcome measures, there is uncertainty about the
evidence to support the increasing clinical use of platelet-rich
plasma [62]. Four studies relevant to foot and ankle applica-
tions were included, and all four were found to have a “very
low” quality of evidence. Inconsistent results are also seen
when using PRP as an adjunct in the treatment of conditions
in other areas of the body. For example, the effectiveness of
PRP in foot and ankle applications differ from those reported
for the spine or maxillofacial conditions. This may be attrib-
uted to the different biological, physiological, and mechanical
factors which influence the local milieu for healing in differ-
ent anatomic areas. Better clinical study design in the form of
well-designed, prospective, randomized, clinical studies are
needed to better evaluate the efficacy of PRP therapy.
To date, no standardized method of preparation has been
adopted. In general, a two-step centrifugation technique is
used to separate various components of anticoagulated
blood. The initial “soft-spin” separates plasma and platelets
from red and white blood cells, while the second “hard spin”
separates the platelet-poor plasma. In some preparation tech-
niques, thrombin is added for exogenous platelet activation
facilitating immediate high-dose delivery of critical growth
factors [63, 64]. Calcium chloride may also be used to precipi-
tate fibrin, which acts not only as a scaffold but also as a res-
ervoir for sustained growth factor release [65]. Variations in
preparation techniques including the initial volume of blood
and centrifugation times yield differences in final PRP vol-
umes and platelet concentrations, and therefore the level of
growth factors delivered to the injury site are inconsistent.
Even when specific protocols are used, the potential for
inconsistency exists because of the variability in an individu-
al’s own platelet count and growth factor levels. Such varia-
tions may influence the efficacy of PRP treatment.
Several investigators have examined the difference in con-
tent and concentration of platelets and growth factors among
commercially available preparation devices. Differences in
164 C. Cunningham et al.
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Chapter 7. Platelet Rich Plasma 169
Introduction
Platelets are not only the primary effectors of hemostasis, but
also carriers of a variety of biologically active factors that can
turn useful for therapeutic purposes, especially, although not
exclusively, tissue healing. Recently characterized platelet-
derived growth factors are able to favor tissue regeneration,
wound healing and angiogenesis. The current availability of
commercial methods to prepare Platelet Rich Plasma (PRP)
has made its use quite popular, although there are still several
areas of uncertainty on the biological efficacy of the prepara-
tions and even on their exact composition, making it essential
to expand our knowledge on the events that characterize
G. Gobbi, PhD
Department of Biomedical, Biotechnological & Translational
Sciences (S.Bi.Bi.T.), University of Parma, Parma, Italy
M. Vitale, MD ()
Anatomy & Histology Unit, Department of Biomedical,
Biotechnological & Translational Sciences (S.Bi.Bi.T.),
University of Parma, Parma, Italy
e-mail: marco.vitale@unipr.it
Molecular Biology
PRP, defined by at least 1,000,000 platelets/µl in 5 ml of plasma
[1] and by a full array of clotting and growth factors [2], acts as
a biologically active factors reservoir, that induce mitogenesis,
chemotaxis and angiogenesis at the site of application [1, 3, 4].
However, several platelet-enriched products are collectively
named PRP, even though many differences in composition and
preparation techniques have been described, confounding the
evidence-based positive effects of “PRP” therapy.
PRP Preparation
Thrombin or
Calcium Chloride
A1 A2 B1 B2
C2
Application at
site of wound
C1
Injection at site
of wound
PB PRP NOTHING
AGONISTS
FIBRIN
COLLAGEN
PRP
LY
ON
HYDROGEL
P
BIOLOGICAL GRAFT
LE
UK RP & (i.e. DBM)
OC
YT
ES
SYNTHETIC GRAFT
(i.e. HA)
CELLS
(i.e. MSC; BMSC;
BM L-PRP CHONDROCYTES)
a b c
Fig. 8.3 Platelet precursor. Platelets origin from long extended pre-
cursors, named proplatelet, similar to strings of discoid structures
containing granules. Granules are distributed in platelet cytoplasm
(a). Platelet surface expresses several glycoproteins (GP), mediators
of platelet adhesion to the extracellular component. GPIIbIIIa (b)
represents the fibrinogen receptor, one of the most expressed pro-
teins on platelet surface. The merged image (c) shows the distribu-
tion of granules and surface receptor
180 G. Gobbi and M. Vitale
PRO-ANGIOGENIC ANTI-ANGIOGENIC
ACTIVITY ACTIVITY
PDGF-AA, -BB, -AB PF4
FGF Angiostatin
HGF Endostatin
VEGF TIMP-1, -4
EGF TSP-1
IGF-1 TGF-beta
PLATELET
BIOACTIVE FACTORS ARE
STORED IN PLATELET AND
RELEASED UPON
ACTIVATION
CHEMOTAXIS ACTIVITY
TGF-beta; PDGF-AA, -BB, -AB; FGF; HGF; VEGF; EGF;
IGF-1; BMP-2, -4, 6; CCL5; CCL3; CXCL-4, -7, -12
Clinical Applications
The rationale for the therapeutic use of PRP is to make
platelet-derived factors locally available for tissue to be
healed. Platelets present in PRP function as a tissue sealant
initiating wound repair [52], while fibrin matrix acts as drug
delivery system releasing platelet-derived bioactive factors
[3]. Local milieu conditioning modifies precursor cell differ-
entiation and angiogenesis, ultimately leading to tissue regen-
eration [59–61]. Indeed, PRP has gained success particularly
in the context of traumatic injuries of poorly vascularized or
ischemic tissues, where the local availability of autologous
pro-differentiative/pro-angiogenic factors more easily boosts
a positive clinical outcome. Moreover, although not yet
standardized, autologous PRP has less safety concerns than
8 Platelet Rich Plasma for Biological Therapy 185
as gelatin hydrogel [25, 96], that do not interfere with the rate
of bone regeneration. On the contrary, the beneficial effects
of PRP associated to grafts are still unclear. Both biological
and synthetic bone grafts support bone healing [97–99] and
the addition of PRP was beneficial in some cases [100–102],
irrelevant in others [103, 104], or even detrimental [105].
Limits
Despite the diffusion of PRP preparations in the treatment of
several injuries, there is a lack of solid knowledge on its real
benefits. This gap is due to several criticisms on the clinical
and experimental approaches to this topic. In a very sche-
matic way, we could summarize PRP flaws as follows.
1. Nomenclature. Indeed, the term PRP embraces a number
of different biological preparations not even well
characterized.
2. Processing. Frequently, details about the techniques used
to obtain PRP and to apply it are not well defined. Indeed,
platelet count is often not reported, as well as the centrifu-
gation step used for platelet enrichment. The amount of
PRP injected or applied is often left to the opinion of the
clinician, as well as the frequency of PRP applications. For
these reasons it is difficult to compare methods and out-
comes, hampering the necessary standardization.
3. Formulation. There is a lack of published information on
the formulation of different PRP-derived products, in
terms of cells and bioactive factors content. The content of
growth factors in platelets is highly variable between indi-
viduals, independently from platelet count. Moreover,
platelets are extremely sensitive to any kind of stress, from
blood drawing to PRP gel production. Thus the amount of
platelet-derived factors available at the end of the manipu-
lation process is likely to depend on the cumulative effects
over platelets, along the entire preparation process.
4. Exploitation. To date, only few randomized controlled tri-
als have been performed in humans to provide the level I
188 G. Gobbi and M. Vitale
Economical Issues
Both the cellular and molecular components of PRP are
autologous, as opposed to the use of recombinant human
growth factors, such as PDGF-BB, which have been recently
associated to malignant transformation in human cells [106].
PRP is autologous and, for this reason, safer than allogenic
preparations. It does not induce immunogenic reactions [2],
and disease transmission is not an issue [2, 10, 107].
Given the availability of simple techniques for PRP pro-
duction, PRP is considered an inexpensive way of enhancing
tissue healing, and its use may be a more cost-effective and
economical approach for the required treatments. Indeed,
PRP represents an easy, cost-effective way to obtain high
concentrations of growth factors at the site of application.
However, several devices have been developed and commer-
cialized for clinical preparations that are promoted in the
absence of solid clinical data on the outcome or effectiveness
in tissue repair [108].
Dougherty [109] compared the cost-effectiveness of PRP
in the treating non-healing diabetic foot ulcers to several
other therapies over a 5-year period in an hypothetical group
8 Platelet Rich Plasma for Biological Therapy 189
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Chapter 9
The Systemic Effects
of Platelet-Rich Plasma
Amy S. Wasterlain, Hillary J. Braun, and Jason L. Dragoo
Growth Factors
The use of other growth factors in athletes is prohibited
under section S2 of the 2013 WADA prohibited list [97], par-
ticularly because of concerns regarding insulin-like growth
Chapter 9. The Systemic Effects of Platelet-Rich Plasma 201
exerted its effects in tissues other than at the site of injury; its
ability to enhance oxygen conductance and exercise endur-
ance implicates VEGF as a candidate for doping regulations.
PRP Treatment
Multiple groups have reported that PRP contains elevated
concentrations of growth factors such as VEGF, PDGF, and
FGF [10, 23, 25, 90]. Even though PRP is not itself banned, its
systemic effects could potentially influence results of anti-
doping tests [86, 90]. For example, because PRP causes some,
but not all, of the changes expected after hGH doping, PRP-
treated athletes could mistakenly test positive for
hGH. Another concern about PRP is that athletes could use
PRP as a vehicle to inject exogenous substances, including
hGH. Testing for PRP could deter the intentional use of PRP
212 A.S. Wasterlain et al.
Conclusion
Sports physicians are in a unique position to uphold the prin-
ciples of clean competition, as well as to investigate new
technologies, such as PRP, with the potential to facilitate
athletes’ recovery from injury in a manner consistent with
anti-doping regulations. Creaney et al. and the broader sports
medicine community have recognized an ironic dichotomy in
which the general public may benefit from PRP, but recovery
from orthopaedic injuries is delayed in elite athletes because
of doping restrictions [17]. As recognized by the International
Olympic Committee (IOC) in a recent Medical Commission
Consensus Statement on the use of growth factor technolo-
gies in therapy, “Knowledge of the genomic and proteomic
impacts of growth factor-based therapies on the target cells,
and of the biomarkers reflecting stem cell differentiation
status, will underpin the development of tests capable of
monitoring therapeutic efficacy and minimizing adverse
events” [55].
Additional research into the potential systemic and ergo-
genic effects of PRP is warranted to ensure athlete safety, and
so that clear, evidence-based guidelines can be implemented
regarding the use of PRP. Future studies should directly com-
pare circulating growth factors in PRP-treated individuals to
those in untreated control participants. Other potential areas
for research include comparing intratendinous versus intra-
muscular injections; evaluating the effects of dry-needling
214 A.S. Wasterlain et al.
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Chapter 9. The Systemic Effects of Platelet-Rich Plasma 219
Introduction
Given the scarce cell number and reduced motility and pro-
liferation rates of tenocytes, and complicated by the absence
of blood vessels, tendons have poor ability to self-repair.
Healing may be also hindered by repetitive mechanical stress
without sufficient time to heal.
Unresolved Inflammation
Abnormal Angiogenesis
Inflammation
Angiogenesis
positively, not only with the number of platelets but also with
leukocyte concentration. However, the angiogenic effect of
PRP is not only attributed to pro-angiogenic factors such as
VEGF, and PRP contains molecular stimuli that up-regulate
the production of further angiogenic molecules [9].
Alpha-granules, the main molecular reservoir in platelets,
also contain established inhibitors of angiogenesis, such as
TSP-1, an adhesive protein modulating vascular cell behavior
by altering endothelial and vascular smooth muscle cell adhe-
sion, proliferation, motility, and survival. TSP-1 concentration
is proportional to the number of platelets [48]. TSP-1 inter-
feres with VEGF and bFGF binding, in doing so suppressing
their mitogenic effects. Other antiangiogenic proteins in
PRPs are angiostatin, endostatin, and fibronectin and the tis-
sue inhibitors of metalloproteinases (TIMPs-1 to -4) [46].
Pro- and anti-angiogenic proteins may well be stored sepa-
rately and differentially released because the secretion of
pro- versus anti-angiogenic stores may be agonist-specific.
For example, PAR1-activating peptide, ADP, and the glyco-
protein VI-targeting collagen-related peptide induced mas-
sive release of VEGF but a modest release of PF-4 or
endostatin. In contrast, PAR4-AP triggered marked PF-4 and
endostatin release. This suggests that depending on the
molecular milieu distinct secretion patterns of pro- and anti-
angiogenic factors will be achieved [17]. On this basis, PRP
therapies have the potential to produce selective pro- or
antiangiogenic environment depending on the host tissue
conditions.
Also, often neglected is the fact that a pool of small mol-
ecules, such as histamine, noradrenaline, dopamine, and sero-
tonin (stored in dense granules) can increase vascular
permeability, thereby permitting the extravasation of plasma
proteins into the injury.
Perspectives
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