Current Pharmaceutical Biotechnology, 2012, 13, 1257-1265 1257

The Role of “Eye Platelet Rich Plasma” (E-Prp) for Wound Healing in
Ophthalmology

Jorge L. Alió1,2, Francisco Arnalich-Montiel3,4,* and Alejandra E. Rodriguez4

1
Department of Cornea and Refractive Surgery, VISSUM Corporation, Alicante, Spain; 2School of Medicine, Miguel
Hernandez University, Alicante, Spain; 3Department of Cornea and Refractive Surgery, VISSUM Corporation, Madrid,
Spain; 4Research & Development Department, VISSUM Corporation, Alicante, Spain

Abstract: Blood derived products have demonstrated their capacity to enhance healing and stimulate the regeneration of
different tissues and this enhancing effect is attributed to the growth factors and bioactive proteins that are synthesized and
present in blood. Eye platelet rich plasma (E-PRP) provides higher concentration of essential growth factors and cell ad-
hesion molecules by concentrating platelets in a small volume of plasma as compared with autologous serum, the latter
being used widely in ophthalmology for epithelial wound healing of the cornea for the last two decades. These growth fac-
tors and cell adhesion molecules have a major role in wound healing and enhance the physiological process at the site of
the injury/surgery via eye drops or clot. E-PRP has been used more recently, and has achieved successful outcomes in
peer-review articles in the treatment of dormant ulcers (epithelial defects of the cornea that fail to heal), moderate to se-
vere dry eye syndrome, ocular surface syndrome post Laser In Situ Keratomileusis (LASIK), and for surface reconstruc-
tion after corneal perforation associated with amniotic membrane transplantation. Preparation of E-PRP in the two avail-
able formulations, eyedrops and clot, is inexpensive and easy although it requires following strict sterility conditions using
sterile and disposable materials and operating inside a laminar flow hood. No serious adverse effects have been described
with the use of these products, and it is generally well tolerated. In summary, Platelet enriched plasma in the form ob-
tained in ophthalmology, E-PRP, is a reliable and effective therapeutic tool to enhance epithelial wound healing in ocular
surface disease.
Keywords: Eye platelet rich plasma; corneal dormant ulcer; dry eye; corneal perforations; ocular surface syndrome post
LASIK.

INTRODUCTION decades, and it has been widely used in the treatment of ocu-
lar surface disease such as persistent epithelial defects or
The ocular surface comprises the conjunctival mucosa
neurotrophic keratopathy as well as following corneal graft-
that lines the globe and palpebral surfaces, the corneoscleral
ing [1-4]. Amniotic membrane or hemotherapy derivates can
limbus, the corneal epithelium and the tear film. This com-
actively stimulate the corneal epithelial turnover and migra-
plex structure needs a stable tear film, normal blink, func- tion. The release of growth factors and other cytokines that
tioning lacrimal system and healthy eyelids to maintain a
promote epithelialisation, suppress inflammation and have
balanced homeostasis. Several diseases can compromise the
microbicidal effects, has been the rationale for using amni-
stability of the ocular surface, ranging from chemical or
otic membrane [4, 5] or autologous serum in patients with
physical injuries such as Laser In Situ Keratomileusis (LA-
wound healing impairment [6].
SIK), to autoimmune diseases such as Sjögren disease or
ocular cicatrizial pemphigoid. If corneal wound healing does The goal of this article is to review the use of another
not occur promptly, it can lead to visual loss, severe scarring, blood derived product known as eye platelet-rich plasma (E-
infection and even corneal perforation. PRP) in ophthalmology, and outline its current clinical appli-
cations based on the evidence found in the literature.
There are several strategies to promote wound healing.
Firstly, lubrication can be enhanced with the frequent use of
BACKGROUND
artificial eye drops or ointment. By using bandage contact
lenses or patching the eye, closing the eyelids together with a Blood derived products have demonstrated their capacity
tarsorrhaphy procedure or inducing a ptosis with botulin-toxin to enhance healing and stimulate the regeneration of differ-
we can reduce the exposure of the ocular surface to the outside ent tissues and this enhancing effect is attributed to the
environment, facilitating cell proliferation and migration. growth factors and bioactive proteins that are synthesized
and present in blood [7].
Finally, the use of biologically active agents to promote
wound healing has been known in ophthalmology for several Different blood derived formulations, such as autologous
serum, plasma enriched with platelets and preparations rich
*Address correspondence to this author at the Department of Cornea and in growth factors have been used to promote wound healing
Refractive Surgery, VISSUM Corporation, Madrid, Spain; in multiple tissues.
E-mail: arnalich@hotmail.com

1873-4316/12 $58.00+.00 © 2012 Bentham Science Publishers

1258 Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 Alió et al.

The serum is the clear liquid part of full blood after cellu- ing a platelet concentration above baseline. They use a PRP
lar components and clotting proteins have been removed. device, concentrate platelets using a double centrifugation
Since Fox et al. [1], first used autologous serum eye drops in technique and activate PRP just when they are ready to use
the treatment of keratoconjunctivitis sicca, it has been the it. The final concentration is at least 1.000.000 platelets/ mi-
preferred blood derived topical preparation in the treatment crolitre. Therefore, it is an autologous concentration of plate-
of ocular surface diseases. Autologous serum is commonly lets and growth factors. After the complete release of growth
used and has been found effective for the treatment of persis- factors, platelets are able to synthesize and secrete additional
tent epithelial defects [8], neurotrophic ulcers [2], superior growth factors for the remaining several days of their
limbic keratoconjunctivitis [9], and other types of dry eye lifespan thus expanding the wound healing effect [23].
symptoms such as graft versus host disease [10] or after LA-
PRP and PRGF have been successfully used for over a
SIK [11]. Autologous serum has also been used as an adjunc- decade as a component for tissue regeneration procedures
tive treatment in ocular surface reconstruction with different
such as oral and maxillofacial surgery, reconstructive ortho-
results [3, 12]. Unlike artificial tears, serum eye drops have
paedics, cardiovascular surgery and plastic surgery [21, 24,
pH, osmolarity and biomechanical properties which resemble
25]. Dental implant surgery with guided bone regeneration is
natural tears, and they are non-preserved. When used topi-
one example in which an autologous platelet-rich clot has
cally they supply essential nutrients to the ocular surface
been shown to accelerate ossification after a tooth extraction
such as growth factors, vitamins and bacteriostatic products and/or around titanium implants, with marked reductions in
such as IgG, lysozyme and complement [13]. Therefore, not
the time required for implant stabilization and an improved
only do they provide lubrication, but they also have epithe-
success rate [26-28]. Articular surgery [29], tendon repair
liotrophic and antimicrobial properties which resemble natu-
[30], reversal of skin ulcers [31], macular hole repair in eye
ral tears, and cannot be found in commercialised artificial
surgery [32], and cosmetic surgery [33] are other situations
tears.
in which autologous platelets have shown to accelerate heal-
Plasma, unlike serum, does contain clotting proteins of ing. Concentrated platelet preparations are also used to in-
full blood such as fibrin. Although the acellular component duce bone regeneration when prosthetic devices are to be
of blood contains growth factors, it is well known that plate- implanted [34] in maxillofacial and implant surgery and also
lets are great reservoirs of growth factors that enhance pro- in traumatology, combined with hydroxyapatite, autologous
liferation and wound healing. The alpha-granules in platelets bone, and other biomaterials [35-37].
contain more than 30 bioactive proteins such as epidermal Other different blood derived preparations such as en-
growth factor (EGF), platelet derived growth factor AB
riched growth factors solution obtained from activated plate-
(PDGF-AB), vascular endothelial growth factor (VEGF),
lets with human thrombin previously washed with a buffer
insulin-like growth factor (IGF-1), transforming growth fac-
solution, has been used to study the epitheliotrophic capacity
tor beta (TGF-
), as well as cytokines including proteins
in cell culture models, including human and rabbit corneal
such as PF4 and CD40L, which promote tissue repair and
epithelial cells [13].
influence the reactivity of vascular and other blood cells in
angiogenesis and inflammation [7]. Growth factors released A meticulous review in the literature shows us that blood
from activated platelets initiate and modulate wound healing derived products like plasma enriched with platelets and
in both soft and hard tissues [14, 15]. The plasma also con- growth factors has not been as widely used in ophthalmology
tains concentrated quantities of some important cell-adhesion as it has been in other disciplines that deal with wound heal-
molecules which promote epithelial migration such as fibrin, ing.
fibronectin and vitronectin [16]. Laboratory research with
different cultured cellular lines such as tendon cells, synovial AUTOLOGOUS E-PRP PREPARATION IN OPH-
and skin fibroblasts and corneal epithelial cells, shows the THALMOLOGY: EYEDROP AND CLOT FORMULA-
multiple benefits and biological effects of growth factors [13, TIONS
17-19].
The E-PRP (eye platelet rich plasma) used in ophthal-
In the literature there are different autologous blood de- mology is an autologous preparation of plasma rich in plate-
rived products with variable amounts of platelets and growth lets, but different from the PRP described by Marx et al. [23]
factors which are used to promote wound healing and tissue and slightly different from PRGF [38]. The E-PRP uses so-
regeneration. dium citrate as an anticoagulant and when necessary calcium
chloride is used for the activation of the E-PRP clot. The
Autologous PRGF has been used for multiple purposes
main difference with PRGF is that the E-PRP preparation
by Anitua et al. [20] since 1999. At the beginning PRGF was
referred to as “Plasma Rich in Growth Factors” [21], and uses commercial tubes to obtain the blood and a laboratory
centrifuge is used for plasma separation. E-PRP does not
more recently has changed to “Preparation Rich in Growth
require specific devices.
Factors” [22] due to the possibility of obtaining different
formulations for regenerative purposes in multiple medical The E-PRP obtention is carried out using a one-step cen-
conditions. PRGF technology identifies 100% autologous trifugation process and the final rate of the platelet concen-
and biocompatible products elaborated using a one-step cen- tration depends on whether it will be used as eyedrops (with-
trifugation process, sodium citrate and calcium chloride as an out activate) or as a clot (activated).
anticoagulant and activator, respectively.
The autologous E-PRP in the form of topical eyedrops is
Platelet-rich plasma (PRP) is defined by Marx et al. [23] used for surface applications and as a clot for surgical proce-
as a portion of the plasma fraction of autologous blood hav- dures for ocular reconstruction.
The Role of E-PRP for Healing in Ophthalmology Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 1259

E-PRP EYEDROP PREPARATION ous adverse events have been described using PRP for ocular
surface diseases. Only one patient from our series developed
80-100 ml of blood is obtained from the patient by
tolerability problems and discomfort with the topical admini-
venipuncture and collected in 10 ml sterile tubes containing
stration of E-PRP [40]. There is not a single case reported of
1ml of 3,2% sodium citrate to avoid coagulation. E-PRP
infection secondary to microbial contamination of the eye
preparation is made following strict sterility conditions using drops, as it is prepared under strict sterile conditions.
sterile and disposable materials and operating inside a lami-
nar flow hood.
CLINICAL APLICATIONS
Citrated tubes containing blood are spun at 5ºC, 1400
Dormant Ulcers
rpm for 10 minutes and ninety percent of plasma obtained
after centrifugation is collected and used as the final product. Dormant ulcers are epithelial defects of the cornea that
Nevertheless, the speed and time vary depending on the fail to heal in spite of at least 2 weeks of conventional treat-
characteristics of each centrifuge and the size of tubes used. ment, and are most commonly caused by neurotrophic kera-
A haemocytometer is needed to quantify the number of topathy (including metaherpetic disease), dry eye, or immu-
platelets in whole blood after the centrifugation in order to nological disorders such as rheumatoid arthritis or ocular
obtain the maximum enrichment. We find that PRP made cicatricial pemphigoid.
with our conditions yields a 1,6-2,5 fold enrichment of plate-
lets when compared to full blood. In a prospective study Alió et al. [41] included 26 eyes
with dormant corneal ulcers, 12 had neurotrophic keratopa-
Three to four millilitres aliquots of E-PRP are placed into thy, 9 had herpetic keratopathy and 5 had ulcers of immu-
new, sterilized 10 ml amber glass bottles with eye drop ap- nological origin Fig. (1). They were treated with eye drops
plicators. Patients are instructed to wash their hands before made from autologous platelet rich plasma at a dose of 6
using the product, to keep the area of application of the eye times a day in addition to routine medication. Primary out-
drops clean and to avoid touching the eyedropper. The bottle come measures were the reduction in size or depth of the
in use can be kept at +4ºC for one week, after which, the corneal ulcer and improvement in best-corrected vision. Sec-
bottle in use needs to be discarded. All these preventions are ondary outcome measures were the reduction of pain or dis-
carried out in order to avoid contamination. Until used, the comfort, decrease in conjunctival or ciliary hyperemia, or
rest of the PRP tubes should be kept at -20ºC. conjunctival edema if present.
As eye drops need to be used in the liquid form, there is
no activation of the coagulation until the drops are instilled
and aggregation takes place. Endogenous release of activa-
tors of the coagulation in the site of application results in a
slower release of growth factors and chemical mediators,
providing a longer effect.

E-PRP CLOT PREPARATION

For the preparation of E-PRP clot, 40-60 ml of blood is
obtained from the patient just before the surgery and centri-
fuged by the one-step process as previously described for the
E-PRP eye drop formulation. However in this case, only the
plasma nearest to the red cells is harvested, avoiding the
white blood cell layer due to the unknown effects of leuko-
cytes [39]. 1 ml of E-PRP is placed into each 4 well tissue
culture plates (Nunc®) and 50 μl of 10% calcium chloride Fig. (1). Patient with neurotrophic keratopathy, showing intense
inflammation.
(Braun®) are added to each well for activation. After mixing
carefully with a sterile pipette, the plates are incubated at
37ºC for 30 minutes. After that time the E-PRP clot is Significant clinical improvement was found in 92 % of
formed and it is ready to be applied onto the ocular defect the eyes (24/26), with a complete resolution of the ulcer in
immediately afterwards. In the E-PRP clot the rate of the 50% of the cases (13/26) Fig. (2). Only two eyes did not
platelets enrichment is about 2-3 times over the full blood show significant changes after the treatment. Reduction in
values. The manipulation of the tubes to obtain the E-PRP inflammation and decrease of ocular pain were the other
must be done under strict sterile conditions using a laminar parameters that clearly improved in the majority of the cases.
flow hood. Two eyes with a relapsing epithelial defect, defined as ob-
servation of a new epithelial defect located at the level of the
SAFETY previous ulcer with positive fluorescein staining under slit-
lamp examination, occurred after 6 months to 1 year and
The concern about transmissible diseases such as HIV,
were treated successfully by keratoplasty.
hepatitis, or Creutzfeldt-Jakob disease, which arises when
using allogenic blood derivates do not apply to either serum Visual acuity also improved in more than half of the pa-
drops or E-PRP as they are entirely autologous. Bovine tients, with 31% of the eyes gaining 1-3 lines of visual acu-
thrombin is not used anymore as the clotting initiator so the ity, 15% with an improvement of 4-5 lines of visual acuity,
stimulation of antibody production is not an issue. No seri- and 12% with more than 6 lines of improvement. A reduc-
1260 Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 Alió et al.

tion in inflammation occurred in 1 to 2 weeks, with a de- also compromise visual acuity if it is severe enough. Artifi-
crease in ocular pain. cial tears are the main standard treatment for this disease, but
very often not sufficient enough to stop all the symptoms or
signs of the disease.

Fig. (2). Same patient as in fig. (1), two weeks after treatment with
E-PRP. Inflammation has been reduced and there is no epithelial
defect. Fig. (3). Patient with severe dry eye. Fluorescein stains in green and
shows punctate keratopathy with blue cobalt light.
The same study [41] included 14 eyes treated surgically
with clot of autologous E-PRP and amniotic membrane in
Some studies have shown that dry eye is often associated
perforated corneas or with impending perforation due to cor-
to a low-grade inflammatory reaction on the ocular surface
neal melting following chronic corneal ulcer. We will review with production of pro-inflammatory cytokines, and a status
this subgroup of patients in detail in the paragraph dedicated
of activation of T cells in the lacrimal glands and conjunctiva
to surgical use of E-PRP.
either with or without Sjögren’s syndrome [42, 43]. Dry eye
This report shows that E-PRP improved photophobia, symptoms usually correlate with the severity of the disease
pain and inflammation; facilitated reepithelialisation, pro- but, according to recent reports, when accompanied by in-
moting corneal wound healing and improving the clinical flammation, they may cause a down regulation of sensory
condition, which in the end resulted in improved vision in receptors leading to a decrease in symptoms while milder
the majority of the patients included in the study Fig. (1a and conditions may cause the opposite effect.
1b). Although based on a single study and therefore further
Alio et al. [44] conducted a prospective, nonrandomized,
studies are necessary, this article shows that autologous E-
observational consecutive pilot study that included 36 eyes
PRP is a reliable and efficient procedure to restore the cor-
of 18 patients with moderate to severe dry eye syndrome
neal epithelial surface in patients with dormant corneal ul- according to the triple classification of Madrid [45]. Twelve
cers. As it has been described for autologous serum, E-PRP
patients were suffering from moderate dry eye and 6 patients
facilitates reepithelialisation and promotes corneal wound
had severe dry eye symptoms related to Sjögren’s syndrome
healing in intractable corneal ulcers. The advantage of E-
or Stevens-Johnson disease. All the patients had punctate
PRP over autologous serum is that E-PRP has a large quan-
keratitis, with fluorescein staining on at least 50% of the
tity of growth factors that are released from the platelets
corneal surface, were highly symptomatic and showed severe
once these are activated in the ocular surface [41]. Whether ocular surface disturbances Fig. (4). Main outcome meas-
this new therapeutic approach is better than previous ap-
urements included the disappearance of subjective symptoms
proaches is a question that should be answered with a larger
after treatment; the increase in visual acuity, tear meniscus
series of studies with a contemporaneous comparison group.
height and tear breakup time (BUT); the decrease in inflam-
mation and fluorescein staining and improvement of impres-
Dry Eye Syndrome
sion cytology. E-PRP was given topically as eye drops (4-6
Dry eye is a disorder of the tear film caused by a distur- times a day per eye) to patients suffering from moderate to
bance in the composition and quantity of tears, and can be severe dry eye symptoms. Patients were monitored for im-
due to: a) deficiency of the aqueous phase of the tear, as in provement every week. Data was analyzed after 1 month of
Sjögren´s syndrome or lacrimal gland disease b) deficiency treatment.
of the mucin layer caused by a vitamin A deficiency, After 1 month of treatment with E-PRP eye drops Fig.
trachoma, diphtheric keratoconjunctivitis, mucocutaneous (5), 89% of the patients (16/18 patients) experienced relevant
disorders and certain topical medications or c) abnormalities improvement or full disappearance of subjective symptoms,
of the lipid tear layer caused by blepharitis and rosacea. and none of the patients’ symptoms deteriorated whilst re-
Dry eye is very common in ophthalmology and causes ceiving treatment. Twenty-eight percent of patients (5/18)
disturbing subjective symptoms such as dryness, burning, had at least 1 line or more of visual acuity, with no cases of
and a sandy-gritty eye irritation and is accompanied by su- visual loss documented. Improvement of the quality of the
perficial punctate keratopathy Fig. (3), tiny abrasions on the tear film was also reported in more than half of the patients
surface of the eyes and conjunctival hyperemia. Dry eye can with an increase of the tear meniscus height and tear break
The Role of E-PRP for Healing in Ophthalmology Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 1261

up time (BUT). As in the report on the use of E-PRP in dor- preservative component present in these steroid solutions can
mant ulcers, a significant reduction of the inflammation was also affect the bonds between epithelial cells in the cornea
found in 89% of those patients suffering from conjunctival inducing punctate keratopathy that may develop into a cor-
inflammation (6 out of 7 patients). Moreover, improvement neal ulcer [46].
of fluorescein staining of the cornea due to superficial punc- A comparative study with other therapeutic options such
tate keratitis was found in 72% of the cases (13 out of 18).
as autologous serum, which has also shown to be effective
Lastly, with the available data of impression cytology 12 out
[1], is needed. No formal pilot study has been designed yet
of the 18 patients presented an increase in the density of gob-
but we have observed from data obtained in our institute, that
let cells (mucin producing cells) that was statistically signifi-
E-PRP has a slight benefit compared to 100% autologous
cant on the superior bulbar conjunctiva, and that almost
serum, probably due to the fact that the presence of platelets
reached significance (p value 0.053) in the inferior bulbar produces a prolonged release of growth factors. E- PRP
conjunctiva.
should be considered the product of choice in cases of severe
dry eye where the presence of growth factors acting for
longer periods of time is more beneficial.

Ocular Surface Syndrome Post Laser In Situ Kera-

tomileusis (LASIK)
Ocular surface syndrome is a well-known side effect fol-
lowing refractive procedures using LASIK technology.
Clinically it is characterized by dry eye, micropunctate
keratitis, decreased and unstable tear film, and decreased
best spectacle-corrected visual acuity (BSCVA). Although
the exact mechanism for this disease is not completely un-
derstood, it is thought to be secondary to corneal nerve dam-
age which occurs following the cutting of the corneal stroma
in refractive procedures which involve creating a corneal
flap. One of the main functions of the ocular nerves is the
Fig. (4). Patient with severe dry eye prior to E-PRP treatment show-
regulation of secretion activities of the lacrimal and mei-
ing intense keratitis.
bomian glands. Their damage causes neurotrophic epithe-
liopathy and affects tear composition [47]. All lipidic, aque-
ous and mucus components of tears are involved in the qual-
ity of the tear film and any misbalance can compromise the
ocular surface. When a deficiency on the aqueous phase is
present in the tear film, the risk of infection by pathogens
increases. A decrease in the quantity or quality of the mucus
associated with goblet cells of the conjunctiva decreases tear
stability. The same is observed when the alteration of the
lipid composition of the tears affects the tears’ evaporation
control [48]. Treatment of ocular surface syndrome post LA-
SIK surgery with artificial tears is often disappointing [49].
Eye platelet-rich plasma has a lubricating effect and has
been effective in regenerating the ocular surface in cases of
micropunctate keratitis, decreasing inflammation in patients
suffering from dry eye [44] and stimulating wound-healing
Fig. (5). Same patient as in fig. (4), two weeks after treatment with processes in dormant corneal ulcers [41]. This motivated a
E-PRP. prospective study [40] by Alió et al. which included 26 eyes
of 13 patients who had had LASIK surgery to correct myo-
pia. All surgeries were performed using a mechanical mi-
In this clinical study, E-PRP eye drop application im- crokeratome (Moria M2, disposable head; Moria, Paris,
proves regeneration of the ocular surface and relieves symp- France). A 9.5-mm flap was created with a superior hinge of
toms in patients with symptomatic dry eye, with no adverse
3.5 to 4 mm. Flaps were tentatively programmed to be 130-
events with a follow-up of up to 6 months. Autologous E- μm thick based on an intended flap thickness of 160 μm.7
PRP is a preservative-free biological product obtained di- Laser in situ keratomileusis was performed using the Esiris
rectly from the patient’s own blood. Its main components are excimer laser (Schwind, Kleinostheim, Germany). They
platelets and growth factors known for their properties of were all affected by severe to moderate dry eye syndrome
increasing and accelerating the normal healing process by (according to the Triple Classification of Madrid REF) for at
stimulation of cell growth and function, and the symptoms
least 6 months after the surgery. Six of the 13 patients suf-
are relieved rapidly. Topical steroids have also been proved fered from moderate ocular surface dysfunction symptoms
to quickly relieve symptoms, but their long-term usage may prior to treatment Fig. (6) showed severe symptoms. Best
result in side effects such as an increased risk of infection, spectacle-corrected visual acuity decreased from 1 line to 4.
intraocular pressure elevation and cataract formation. The
1262 Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 Alió et al.

All patients were positive for fluorescein staining and factors, thus increasing the endurance of their effects with
showed a tear break-up time between 4 and 9 seconds. Main fewer applications if compared to autologous serum. In a
outcome measures included subjective symptom disappear- previous study [11], no differences were noted in the subjec-
ance, increased visual acuity, increased tear meniscus and tive scores for dryness between the autologous serum eye
tear break-up time, decreased inflammation and fluorescein drops and artificial tears groups. However, further studies
staining and improvement in impression cytology. comparing conventional treatment such as autologous serum
and E-PRP are necessary.

E-PRP as Adjuvant for Ocular Surface Reconstruction:

Role of E-PRP Clot as a Potential Surgical Tool
Ocular surface reconstruction includes limbal autograft or
allograft keratoplasty, amniotic membrane transplantation
(AMT), sectorial epitheliectomy, etc. Theoretically, the ad-
juvant use of PRP would enhance the regenerative effect of
these interventions, by release of growth factors that promote
wound healing and decrease inflammation. Other biologi-
cally active products such as autologous serum have been
used successfully for this purpose [3].
Alió et al. [41] presented a series of cases with perforated
eyes or high risk of perforation due to deep chronic corneal
ulcers treated with amniotic membrane transplantation com-
Fig. (6). Fluorescein staining of a patient with punctate keratitis
bined with a clot of autologous E-PRP Figs. (8 and 9). Sur-
suffering ocular surface syndrome post LASIK before treatment.

E-PRP eye drops applied 6 times daily improved subjec-

tive symptoms in the vast majority of patients and the im-
provement was considered good or excellent in 84% of the
enrolled patients (11/13 patients). Fluorescein staining analy-
sis showed complete resolution of punctate keratitis in 69%
of the cases (18/26 eyes) Fig. (7) and almost complete in
another 23% of the cases (6/26 eyes). E-PRP also improved
visual acuity, providing a gain in vision in 69% of the cases
(18/26 eyes) ranging from 1 to 4 lines of improvement in
visual acuity. Tear break-up time increased to 2 seconds in
46% (12/26 eyes). Fourteen (54%) eyes showed a 0 to 2 sec-
ond tear break-up time increase. One patient (8%) developed
intolerance after 4 weeks and has been the only case reported
in the literature.

Fig. (8). Perforated eye before the treatment with amniotic mem-
brane transplantation combined with a clot of autologous E-PRP.

Fig. (7). Same patient as in Fig. (6), one month after treatment with
E-PRP.

Autologous E-PRP adds a new tool for ocular surface

syndrome treatment after LASIK as it provides subjective
and objective improvement. E-PRP is rich in vitamins and
growth factors. It also contains platelets, which upon activa- Fig. (9). Amniotic membrane transplantation with a clot of E-PRP
tion are capable of providing a prolonged release of growth before placing it underneath.
The Role of E-PRP for Healing in Ophthalmology Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 1263

gery consisted of wound debridement, excision and removal bined with AMT should further promote corneal wound-
of devitalized tissue and an application of amniotic mem- healing processes and further decrease inflammation due to
brane to the wound site with the epithelial side up. A clot of the intrinsic characteristics of E-PRP mentioned throughout
autologous platelet-rich plasma was placed under the amni- the review.
otic membrane to seal the impending or actual corneal perfo- We also have experience (unpublished data) with the use
ration and to increase the therapeutic effect of the amniotic
of E-PRP clot in other surgical interventions to restore the
membrane Figs. (10 and 11). The membrane was sutured
ocular surface such as limbal keratoplasty Figs. (12 and 13)
into place with a continuous 10-0 nylon suture to the con-
with good results, as seen with autologous serum by other
junctiva and a running purse-string suture was applied so that
authors [3, 12]. We believe that the prolonged synthesis and
the membrane tightly adhered to the entire corneal surface.
release of growth factors by the E-PRP clot provides addi-
The eye was closed with a temporary tarsorrhaphy. Primary tional long acting that would increase the benefit of E-PRP
outcome measures were the reduction in size or depth of the
over autologous serum.
corneal ulcer and improvement in best-corrected vision.

Fig. (10). Introducing the clot of E-PRP under the amniotic mem- Fig. (12). 360º Limbal keratoplasty previous to autologous clot of
brane. E-PRP and Tutopatch membrane application, in a patient with lim-
bal deficiency.

Fig. (11). Final result after amniotic membrane transplantation with

a clot of E-PRP in a perforated eye.
Fig. (13). Same patient as in fig. (8) after limbal keratoplasty fol-
lowed by autologous clot of E-PRP and Tutopatch membrane (bo-
All patients showed an improvement in the size of the vine pericardium).
ulcer, and 71% (10/14 eyes) had a complete resolution of the
Other Uses in Ophthalmology
ulcer. Vision also improved in 57% of the cases (8/14 eyes)
along with a decrease in inflammation that occurred 1 to 2 The use of autologous platelets concentrates has also
weeks after the surgery. been described to enhance the healing of the idiopathic
It cannot be ascertained in this study whether the healing macular hole. A significant improvement in the anatomic
effect of the E-PRP in combination with AMT was higher success rate of surgery for idiopathic macular holes of less
than using AMT alone. However, theoretically, E-PRP com- than 3 years' duration was found, but postoperative visual
acuity of the group treated with autologous platelets concen-
1264 Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 Alió et al.

trates was not statistically different from the control group [11] Noda-Tsuruya, T.; Asano-Kato, N.; Toda, I; Tsubota, K. Autolo-
[50]. gous serum eye drops for dry eye after LASIK. J. Refract. Surg.,
2006, 22(1), 61-66.
[12] Tsubota, K.; Shimazaki, J. Surgical treatment of children blinded
CONCLUSIONS by Stevens-Johnson syndrome. Am. J. Ophthalmol., 1999, 128(5),
573-581.
Platelet enriched plasma in the form obtained in oph- [13] Liu, L.; Hartwig, D.; Harloff, S.; Herminghaus, P.; Wedel, T.;
thalmology, E-PRP, is a reliable and effective therapeutic Kasper, K.; Geerling, G. Corneal epitheliotrophic capacity of three
tool to enhance epithelial wound healing in ocular surface different blood-derived preparations. Invest. Ophthalmol. Vis. Sci.,
disease. PRP provides a high concentration of essential 2006, 47(6), 2438-2444.
growth factors and cell adhesion molecules by concentrating [14] Anitua, E.; Sánchez, M.; Nurden, A.T.; Nurden, P.; Orive, G.;
Andía, I. New insights into and novel applications for platelet-rich
platelets in a small volume of plasma. These growth factors fibrin therapies. Trends Biotechnol., 2006, 24(5), 227-234.
and cell adhesion molecules have a major role in wound [15] Nurden, A.T.; Nurden, P.; Sánchez, M.; Andía, I.; Anitua, E. Plate-
healing and enhance the physiological process at the site of lets and wound healing. Front. Biosci., 2008, 13, 3532-3548.
the injury/surgery via eye drops or clot. [16] Marx, R.E. Platelet-rich plasma: evidence to support its use. J. Oral
Maxillofac. Surg., 2004, 62(8), 489-496.
There are only a few controlled clinical studies that pro- [17] Anitua, E.; Andía, I.; Sánchez, M.; Azofra, J.; Zalduendo, M.;, De
vide evidence that the use of autologous E-PRP promotes la Fuente, M.; Nurden, P.; Nurden, A.T. Autologous preparations
wound healing. Larger series and comparative studies using Rich in growth factors promote proliferation and induce VEGF and
HGF production by human tendon cells in culture. J. Orthop. Res.,
other treatment options such as autologous serum should be
2005, 23(2), 281-286.
conducted to determine clinical differences between thera- [18] Anitua, E.; Sánchez, M.; Nurden, A.T.; Zalduendo, M.; de la
peutic options. In theory, E-PRP should provide a higher Fuente, M.; Azofra, J.; Andía, I. Platelet-released growth factors
amount of growth factors in an initial burst as well as late enhance the secretion of hyaluronic acid and induce hepatocyte
synthesis and secretion of growth factors for the remaining 7 growth factor production by synovial fibroblasts from arthritic pa-
tients. Rheumatology, 2007, 46(12), 1769-1772.
days of the life span of a platelet.
[19] Anitua, E.; Sánchez, M.; Zalduendo, M.M.; de la Fuente, M.;
Prado, R.; Orive, G.; Andía, I. Fibroblastic response to treatment
CONFLICT OF INTEREST with different preparations rich in growth factors. Cell Prolif.,
2009, 42(2), 162-170.
None declared. [20] Anitua, E. Plasma rich in growth factors: preliminary results of use
in the preparation of sites for implants. Int. J. Oral Maxillofac. Im-
ACKNOWLEDGEMENT plants, 1999, 14(4), 529-535.
[21] Anitua, E. (2001) The use of plasma-rich growth factors (PRGF) in
None declared. oral surgery. Pract Proced Aesthet Dent., 2001, 13(6), 487-493.
[22] Anitua, E.; Sánchez, M.; Orive, G.; Andía, I. The potential effects
REFERENCES of the preparation rich in growth factors (PRGF) in different medi-
cal fields. Biomaterials, 2007, 28(31), 4551-4560.
[1] Fox, R.I.; Chan, R.; Michelson, J.B.; Belmont J.B.; Michelson, P.E. [23] Marx, R.E. Platelet-rich plasma (PRP): what is PRP and what is not
Beneficial effect of artificial tears made with autologous serum in PRP? Implant Dent., 2001, 10(4), 225-228.
patients with keratoconjunctivitis sicca. Arthritis Rheum., 1984, [24] Foster, T.E.; Puskas, B.L.; Mandelbaum, B.R.; Gerhardt, M.B.;
27(4), 459-461. Rodeo, S.A. Platelet-rich plasma: from basic science to clinical ap-
[2] Matsumoto, Y.; Dogru, M.; Goto, E.; Ohashi, Y.; Kojima, T.; Is- plications. Am. J. Sports Med., 2009, 37(11), 2259-2272.
hida, R.; Tsubota, K. Autologous serum application in the treat- [25] Intini, G. The use of platelet-rich plasma in bone reconstruction
ment of neurotrophic keratopathy. Ophthalmology, 2004, 111(6), therapy. Biomaterials, 2009, 30(28), 4956-4966.
1115-1120. [26] Anitua, E.; Orive, G.; Pla, R.; Roman, P.; Serrano, V.; Andía, I.
[3] Tsubota, K.; Satake, Y.; Ohyama, M.; Toda, I.; Takano, Y.; Ono, The effects of PRGF on bone regeneration and on titanium implant
M.; Shinozaki, N.; Shimazaki, J. Surgical reconstruction of the ocu- osseointegration in goats: a histologic and histomorphometric
lar surface in advanced ocular cicatricial pemphigoid and Stevens- study. J. Biomed. Mater. Res. A., 2009, 91(1), 158-165.
Johnson syndrome. Am. J. Ophthalmol., 1996, 122(1), 38-52. [27] Roldan, J.C.; Jepsen, S.; Schmidt, C.; Knüppel, H.; Rueger, D.C.;
[4] Dua,, H. S.; Azuara-Blanco, A. Amniotic membrane transplanta- Açil, Y.; Terheyden, H. Sinus floor augmentation with simulta-
tion. Br. J. Ophthalmol., 1999, 83(6), 748-752. neous placement of dental implants in the presence of platelet-rich
[5] Azuara-Blanco, A.; Pillai, C.T.; Dua, H.S. Amniotic membrane plasma or recombinant human bone morphogenetic protein-7. Clin.
transplantation for ocular surface reconstruction. Br. J. Ophthal- Oral Implants Res., 2004, 15(6), 716-723.
mol., 1999, 83(4), 399-402. [28] Garg, A.K.; Gargenese, D.; Peace, I. Using platelet-rich plasma to
[6] Kojima, T.; Higuchi, A.; Goto, E.; Matsumoto, Y.; Dogru, M.; develop an autologous membrane for growth factor delivery in den-
Tsubota, K. Autologous serum eye drops for the treatment of dry tal implant therapy. Dent. Implantol. Update, 2000, 11(6), 41-44.
eye diseases. Cornea, 2008, 27(Suppl 1), S25-30. [29] Sánchez, M.; Anitua, E.; Azofra, J.; Aguirre, J.J.; Andia, I. Intra-
[7] Anitua, E.; Andia, I.; Ardanza, B.; Nurden, P.; Nurden, A.T. articular injection of an autologous preparation rich in growth fac-
Autologous platelets as a source of proteins for healing and tissue tors for the treatment of knee OA: a retrospective cohort study.
regeneration. Thromb. Haemost., 2004, 91(1), 4-15. Clin. Exp. Rheumatol., 2008, 26(5), 910-913.
[8] Tsubota, K.; Goto, E.; Shimmura, S.; Shimazaki J. Treatment of [30] Sánchez, M.; Anitua, E.; Azofra, J.; Andía, I.; Padilla, S.; Mujika,
persistent corneal epithelial defect by autologous serum applica- I. Comparison of surgically repaired Achilles tendon tears using
tion. Ophthalmology, 1999, 106(10), 1984-1989. PRGF. Am. J. Sports Med., 2007, 35(2), 245-251.
[9] Goto, E.; Shimmura, S.; Shimazaki, J.; Tsubota, K. Treatment of [31] Anitua, E.; Aguirre, J.J.; Algorta, J.; Ayerdi, E.; Cabezas, A.I.;
superior limbic keratoconjunctivitis by application of autologous Orive, G.; Andia, I. Effectiveness of autologous preparation-rich in
serum. Cornea, 2001, 20(8), 807-810. growth factors for the treatment of chronic cutaneous ulcers. J
[10] Ogawa, Y.; Okamoto, S.; Mori, T.; Yamada, M.; Mashima, Y.; Biomed. Mat. Res. B., 2008, 84(2), 415-421.
Watanabe, R.; Kuwana, M.; Tsubota, K.; Ikeda, Y.; Oguchi, Y. Au- [32] Blumenkranz, M.S.; Ohana, E.; Shaikh, S.; Chang, S.; Coll, G.;
tologous serum eye drops for the treatment of severe dry eye in pa- Morse, L.S.; De Bustros, S. Adjuvant methods in macular hole sur-
tients with chronic graft-versus-host disease. Bone Marrow Trans- gery: intraoperative plasma-thrombin mixture and postoperative
plant., 2003, 31(7), 579-583. fluid-gas exchange. Ophthalmic. Surg Lasers, 2001, 32(3), 198-
207.
The Role of E-PRP for Healing in Ophthalmology Current Pharmaceutical Biotechnology, 2012, Vol. 13, No. 7 1265

[33] Man, D.; Plosker, H.; Winland-Brown, J.E. The use of autologous [42] Pflugfelder, S. C.; Wilhelmus, K. R.; Osato, M. S.; Matoba, A. Y.;
platelet-rich plasma (platelet gel) and autologous platelet-poor Font, R. L. The autoimmune nature of aqueous tear deficiency.
plasma (fibrin glue) in cosmetic surgery. Plast. Reconstr. Surg., Ophthalmology, 1986, 93(12), 1513-1517.
2001, 107(1), 229-237. [43] Pepose, J.S.; Akata, R.F.; Pflugfelder, S.C; Voigt, W. Mononuclear
[34] Mazor, Z.; Peleg, M.; Garg, A.K.; Luboshitz, J. Platelet-rich plas- cell phenotypes and immunoglobulin gene rearrangements in la-
ma for bone graft enhancement in sinus floor augmentation with crimal gland biopsies from patients with Sjogren's syndrome. Oph-
simultaneous implant placement: patient series study. Implant thalmology, 1990, 97(12), 1599-1605.
Dent., 2004, 13(1), 65-72. [44] Alio, J.L.; Colecha, J.R.; Pastor, S.; Rodriguez, A.; Artola, A.
[35] Velich, N.; Nemeth, Z.; Hrabak, K.; Suba, Z.; Szabo, G. Repair of Symptomatic dry eye treatment with autologous platelet-rich plas-
bony defect with combination biomaterials. J. Craniofac. Surg., ma. Ophthalmic. Res., 2007, 39(3), 124-129.
2004, 15(1), 11-15. [45] Murube, J.; Benitez del Castillo, J.M.; Chenzhuo, L.; Berta, A.;
[36] Siebrecht, M.A.; De Rooij, P.P.; Arm, D.M.; Olsson, M.L.; Aspen- Rolando, M. The Madrid triple classification of dry eye. Arch. Soc.
berg, P. Platelet concentrate increases bone ingrowth into porous Esp. Oftalmol., 2003, 78(11), 595-601.
hydroxyapatite. Orthopedics, 2002, 25(2), 169-72. [46] Yang, C.Q.; Sun, W.; Gu, Y.S. A clinical study of the efficacy of
[37] Lieberman, J.R.; Daluiski, A.; Einhorn, T.A. The role of growth topical corticosteroids on dry eye. J. Zhejiang Univ. Sci. B., 2006,
factors in the repair of bone. Biology and clinical applications. J. 7(8), 675-678.
Bone Joint Surg. Am., 2002, 84-A(6), 1032-1044. [47] Wilson, S.E.; Ambrosio, R. Laser in situ keratomileusis-induced
[38] Anitua, E.; Sánchez, M.; Orive, G.; Andía, I. PRGF technology: a neurotrophic epitheliopathy. Am. J. Ophthalmol., 2001, 132(3),
biological delivery system of growth factors. Trends Pharmacol. 405-406.
Science, 2008, 29(2), 37-41. [48] Holly F.J. Physical chemistry of the normal and disordered tear
[39] Anitua, E.; Sánchez, M.; Orive, G.; Andía, I. Shedding light in the film. Trans. Ophthalmol. Soc. UK, 1985, 104(Pt 4), 374-380.
controversial terminology for platelet rich products. J. Biomed. Ma- [49] Ambrosio R.Jr.; Tervo, T.; Wilson, S.E. LASIK-associated dry eye
ter. Res. A., 2009, 90(4), 1262-1263. and neurotrophic epitheliopathy: pathophysiology and strategies for
[40] Alio, J.L.; Pastor, S.; Ruiz-Colecha, J.; Rodriguez, A.; Artola, A. prevention and treatment. J Refract Surg., 2008, 24(4), 396-407.
Treatment of ocular surface syndrome after LASIK with autolo- [50] Pâques, M.; Chastang, C.; Mathis, A.; Sahel, J.; Massin, P.; Dos-
gous platelet-rich plasma. J. Refract. Surg., 2007, 23(6), 617-619. quet, C.; Korobelnik, J.F.; Le Gargasson, J.F.; Gaudric, A. Effect of
[41] Alio J.L; Abad M.; Artola A.; Rodriguez-Prats J.L.; Pastor S.; autologous platelet concentrate in surgery for idiopathic macular
Ruiz-Colecha J. Use of autologous platelet-rich plasma in the hole: results of a multicenter, double-masked, randomized trial.
treatment of dormant corneal ulcers. Ophthalmology, 2007, 114(7), Platelets in Macular Hole Surgery Group. Ophthalmology, 1999,
1286-1293. 106(5), 932-938.

Received: April 21, 2010 Revised: October 15, 2010 Accepted: October 18, 2010