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ARCH SOC ESP OFTALMOL 2007; 82: 475-482
ORIGINAL ARTICLE
SUBCONJUNCTIVAL APPLICATION OF PLASMA
PLATELET CONCENTRATE IN THE TREATMENT
OF OCULAR BURNS. PRELIMINARY RESULTS
APLICACIÓN SUBCONJUNTIVAL DE CONCENTRADO
DE PLAQUETAS PLASMÁTICAS EN EL TRATAMIENTO
DE QUEMADURAS OCULARES. RESULTADOS
PRELIMINARES
MÁRQUEZ-DE-ARACENA R1, MONTERO-DE-ESPINOSA I1, MUÑOZ M2, PEREIRA G3
ABSTRACT
RESUMEN
Purpose: The efficiency of the subconjunctival
application of autologous platelet concentrate in
patients with ocular burns was assessed.
Methods: This was carried out by analysing the
effect of treatment in the eyes of 10 patients suffering from ocular burns as a result of work-related
accidents. Two types of treatment were evaluated:
the first group only received conventional topical
medical treatment; and the second group, in addition, had subconjunctival injection of plasma platelet concentrate. The clinical condition of the patient
and the period in which the disease prevented the
patient from working were studied; monitoring was
carried out until the burns had fully healed.
Results: Statistically significant differences were
found between the group treated with subconjunctival autologous platelet concentrate and the group
treated with conventional topical medications, with
Objetivo: Se valoró la eficacia de la aplicación subconjuntival de concentrado de plaquetas plasmáticas
autólogas en pacientes con quemaduras oculares.
Método: Para ello se analizaron 10 ojos de 10
pacientes que sufrieron accidentes laborales por
quemaduras oculares del mismo grado. Se establecieron dos tipos de tratamiento: al primer grupo,
control, se le aplicó tratamiento médico tópico convencional; al segundo, se le añadió además, inyección subconjuntival de concentrado de plaquetas
autólogas. Se estudió la evolución clínica, días que
le incapacitaron dicha patología para su actividad
laboral y seguimiento hasta su curación final.
Resultados: Se encontraron diferencias entre el
grupo de los tratados con concentrado de plaquetas
autólogas subconjuntivales respecto al grupo con
tratamiento médico tópico convencional, siendo
más corto el tiempo de cicatrización de córnea (sig-
Received: 17/7/06. Accepted: 12/6/07.
FREMAP Hospital, Sevilla. Centro Cid Clinic, Sevilla. Surgery Dept. (Ophthalmology Area). Medicine Faculty of Seville University.
1 Ph.D. in Medicine. Seville University. Spain.
2 Ph.D. in Medicine. Virgen del Rocío Hospital. Seville. Spain.
3 Graduate in Medicine. Virgen del Rocío Hospital. Seville. Spain.
Study financed by the Casa Real de los Godos Institution.
Correspondence:
Rafael Márquez de Aracena del Cid
Servicio de Oftalmología
Hospital FREMAP de Sevilla
Avda. de Jerez, s/n
Sevilla
Spain
E-mail: [email protected]
MÁRQUEZ-DE-ARACENA R, et al.
a shorter period of time in corneal and conjunctival
healing, time on sick leave and time needed for full
healing.
Conclusions: Subconjunctival infiltration of autologous platelet concentrate should be considered as
a straightforward, economical and possibly effective form of treatment for traumatic accidents (burns)
of the ocular surface (Arch Soc Esp Oftalmol 2007;
82: 475-482).
nificativamente estadístico), conjuntiva, baja laboral y tiempo hasta curación final.
Conclusiones: Podemos considerar la autoplaquetoterapia subconjuntival (infiltración subconjuntival
de concentrado de plaquetas autólogas) como un
tratamiento sencillo y económico en el tratamiento
de las alteraciones traumáticas (quemaduras) de la
superficie ocular. Cuya eficacia deberá ser confirmada por series mayores.
Key words: Ocular surface, ocular burns, platelet,
growth factors, cornea.
Palabras claves: Superficie ocular, quemaduras
oculares, plaquetas, factores de crecimiento, córnea.
INTRODUCTION
morphnuclear (PMN) to the cornea (9). These will
produce leucotrienes, chemotactics for new neutrophiles and collagenase which would cause the
enzymatic degradation of the collagen and therefore corneal perforation. The epithelial growth factor which stimulate keratocytes to form new anomalous collagen which will cause a whitish scar and
develop intra-cytoplasmatic contractile fibrils
which will cause scarring and possibly irregular
astigmatism (4).
The objective of the treatment is to achieve
corneal and conjunctival epithelialization as fast as
possible with the least possible permanent damage.
Prolonged episodes of corneal de-epithelialization
increase the production of collagenase in the
corneal stroma, which may reach corneal perforation. The basis of conventional treatment, comprises abundant ocular washing, cleaning product
remains from the ocular surface, debridling necrotic areas and applying topical treatment with corticoids, antibiotics and cyclopegics (3). Severe
lesions may require, in addition to the above, systemic medical treatment such as tetracyclines (10),
corticoids, antioxidants and collagen synthesis
enhancers such as ascorbic acid (11). In serious
burns, surgical treatment options may be necessary
such as amniotic membrane transplant (12), or self
grafting of sclerocorneal limbus (13) in order to
regenerate limbar stem cells to promote epithelial
formation and suppressing fibrosis. The classic
techniques include self-hemotherapy —subconjunctival injection of autologous blood— in patients
with important ocular burns, the inefficiency of
which has been proven (14-17) even when associated to other substances (18).
On the other hand, it is known that autologous
serum seems to facilitate epithelial regeneration and
Ocular burns account for 12-19% of eye traumatisms (1), generally involving young people in their
working or domestic activities (2). This pathology
requires quick and efficient action from which the
final prognosis will depend. Burns can be chemical
or physical. Most of chemical burns are produced
by acid or alkaline substances. The severity of a
chemical burden is related to the concentration of
the product, the pH of the solution and the exposure
time. Alkaline products cause more serious damages due to a denaturalization of proteins and
saponification of membrane lipids, which allows
the alkaline material to quickly penetrate the cornea
and enter the eye. In turn, acid precipitate corneal
proteins, which act as a buffer and barrier against
deeper penetration. In general, the health professional must consider the degree of involvement of
the cornea, of the conjunctiva and the sclerocorneal
limbus, with the letter had been increased relevance
because it hosts the stem cells in charge of epithelium regeneration (3).
After a corneal burn, there is a first attempt at
recovery, whereby the basal epithelial cells on the
edges of the ulceration lose their hemidemosomes
and migrate to cover the defect. The definitive
recovery will be at the expense of the transitory
cells and limbar stem cells (4). When these are
destroyed, due to the action of chemical mediators
found in the stroma and the absence of fibroblasts
—which produce keratocyte growth factors— (5),
the conjunctival epithelium undergoes a transformation and covers the cornea (6,7). The increased
vascularization will end up conjunctivalizing the
cornea (8). In the corneal stroma, the collagen is
degraded, which stimulate the migration of poly-
476
ARCH SOC ESP OFTALMOL 2007; 82: 475-482
Treatment of ocular burns with plasma platelet
healing of ocular surface pathologies (19-22)
because, among other things, it provides growth
factors.
Said growth factors are proteins which play an
essential function in complex tissue repair and
regeneration processes. They intervene in intercellular communication, transmitting their information
by interacting with receptors in the cellular membrane. In addition, they enhance cellular chemothaxis, proliferation and differentiation (23). Growth
factors can be found inside platelets and stored in
alpha granules, and several endogenous factors
have been found to be involved in the repair of tissues (24), including the eye (25).
The possible application of growth factors
derived from platelet concentrates on epithelium
corneal cells could enhance their trophism (26). In
cell cultures, it has been observed that the application of specific serum and plasma fractions
enhance the growth, migration and differentiation
of cells (27). In addition, a possibility has opened
for the utilization of asthmatic platelet concentrates in ocular surface pathologies such as transplants (28).
The efficiency of topical application of autologous serum and of autohemotherapy in specific
alterations of the ocular surface may lead to the
conclusion that the therapeutic potential of this
technique resides in the content of the platelet fraction containing the blood. To date, it has not been
proved that growth factors are the only blood derivatives which enhance corneal epithelialization
processes.
At this point, it is necessary to determine whether
the application of plasma platelet concentrates act
efficiently in the regeneration of ocular surface
alterations. For this, the regeneration of burned ocular surface tissues after the subconjunctival application of plasmatic platelet concentrates was clinically analyzed, assessing the utility and inefficiency of
the said technique. The preliminary results are presented below.
SUBJECTS, MATERIAL AND
METHOD
We analyzed ten eyes of ten patients who attended the urgency practice of our hospital (Labor
pathology monographic) due to Labor accidents:
grade 3 ocular burns as per Dua classification (29),
that is with corneal defects with three to six hours
of corneal involvement and with 30-50% of conjunctival lesion. After receiving prescribed treatment according to method and protocol, we made
clinical follow up until healing.
The patients were selected in sequential manner
among the first who attended the urgency service
with said pathologies and authorized the corresponding treatments. In those who had burns in
both eyes the one with the more severe injury was
chosen for the study.
Distribution in groups
The patients were divided in two groups depending on the treatment they received:
— Group A (5 patients) received conventional
topical treatment.
— Group B (5 patients) received topical treatment + subconjunctival platelet concentrate (SPC).
Conventional medical treatment was given to all
patients of the study, as follows:
After a profuse ocular wash with saline solution
and cleaning with swab, the following medical
treatment was established: cycloplentolate eye
drops, 1 drop every 12 hours; dexametasone eye
drops, 1 mg and tobramycine 3 mg (tobradex,
Alcon-Cusí Labs) 1 drop every 6 hours: retinol
palmitate cream, 10.000 U.I. + gentamicine sulphate 3 mg + DL-Methionine 5 mg (epithelializing
eye cream, Novartis) every 6 hours + occlusion of
the eye. When found necessary, antitetanus procedure was adopted.
For Group B the following protocol was used:
1. Detailed information was given to the patient
who then signed the informed consent.
2. Extraction of 20 cc of peripheral blood in
4.5 ml sterile tubes with 0.5 ml sodium citrate
base as anticoagulant. In addition, blood was
extracted for clinical analysis comprising complete hemogram and biochemistry, hepatitis and
HIV.
The tubes with the blood samples were centrifuged 7 minutes at 2,100 rpm, separating the
plasma in fractions. For our study we utilized the
plasma fraction with more platelets, i.e., the 200400 µl (0.2-0.4 cc.) immediately above the red
series. 1-2 ml were obtained after centrifugation
when hematocrite is within the usual clinical parameters.
ARCH SOC ESP OFTALMOL 2007; 82: 475-482
477
MÁRQUEZ-DE-ARACENA R, et al.
Subconjunctival treatment
RESULTS
In a sterile environment (operating theatre) and
after 3 applications in the eye to be treated of 1 drop
every 3 minutes of 1mg clorhydrate tetracaine +
4mg of clorhydrate oxybuprocaine (Colircusi double anesthetic®, Alcon-Cusí), the platelet concentrate was subconjunctivally injected in the bulbar
conjunctiva (fig. 1) 0.5-1 ml as 12 o’clock and
another one at 6 o’clock. Antibiotic cream + antiinflammatory eye drops were applied, with 24-hour
occlusion.
Clinical evaluation method
The patient’s subjective symptoms and the clinical evaluation of the lesions were assessed by slit
lamp biomicroscopy with/without topical fluorescein dyeing. To measure the injuries as well as the
evolution thereof, the eye was divided in 12 hourly
Due sections (29), analyzing the fluorescein-dyed
area and hyperemia.
Data were collected in a Type Excel base..
The statistical study was made as follows:
mean values with standard deviation; percentages by means of Tukey pivots and signification
by means of the Kruskal-Wallis test, as well as
Pearson correlations (parametric and non-parametric).
Ten male patients who had suffered Type III
caustic eye burns as per Dua classification (35) due
to labor accidents. The mean age of these patients
was of 32.25 years (SD=15.52). Four were right
eyes (40%), five were left eyes (50%) and one had
both eyes involved (10%). In what concerns the
labor field of the patients, 50% (five patients) were
from the construction industry, followed by the services sector (30%, three patients) and 20% from
manufacturing (two patients). The most frequent
etiological agents were six patients with alkali
burns (60%), three due to solvents (30%) frequently used in construction and services, and one
patient due to acids (10%).
As regards the cicatrization time of the cornea, it
was of 6 days for group A (conventional treatment)
and 3 days for group B (platelet self-therapy), with
statistically significant differences being found
(0,001) between both groups.
The conjunctival cicatrization time was longer in
group A (6 days) than in group B (4 days), although
the difference was not statistically significant
(0.017).
In what concerns keratitis, shortening was evident in group B patients (6 days) vis-à-vis group A
(9 days) but, as above, the difference was not statistically significant.
As for the time off work, no statistically significant differences were found between group A (7
days) and group B (6 days).
In contrast, the final healing time exhibited differences between group A (10 days) and B (6 days)
but, as with other parameters, these were not statistically significant (0.017).
The analytical study of patients treated with
autologous plasma concentrates did not reveal any
correlations with the parameters of the study (parametric and non-parametric Pearson correlation).
Side effects of the treatments
Fig. 1: Conjunctival chemosis after infiltration of subconjunctival plasmatic platelet concentrate in an eye
with alkali burns.
478
Group A did not evidence undesirable effects. In
Group B, 100% of the patients treated with subconjunctival platelet concentrate exhibited no chemosis
after 24 hours (fig. 2). Three patients (60%) referred
ocular irritation one minute after injection, which
disappeared within 3-5 minutes. Three patients
(60%) exhibited highly localized and slightly ele-
ARCH SOC ESP OFTALMOL 2007; 82: 475-482
Treatment of ocular burns with plasma platelet
(EGF) and neural growth factor (NGF) seem to
enhance the healing of corneal defects (36-38),
improving vision and corneal sensitivity (39).
Endothelial vascular factors could play an important role in the physiology of epithelial conjunctival
cells (25). It has also been proven that the exogenous application of factors (KGF-2) in experimental animals (rabbits) could stimulate the migration
of limbar stem cells to the center of the cornea, thus
enhancing the healing of a burnt cornea (40). Said
stimulation seems to be related to the concentration
of growth factors (41). The growth factors in the
amniotic membrane enhance the growth of cultivated limbar stem cells (42).
The topical application of mice growth factors in
three patients with persistent epithelial defects due
to neurotrophic keratitis improved the condition,
but application problems were noted due to costs as
well as technical difficulties (43).
In this study it can be seen that the subconjunctival injection of autologous platelet concentrates
produces a statistically significant reduction of the
cicatrization time in Type III corneal burns (0.001).
In what concerns the cicatrization of the conjunctiva, a certain degree of activity of endogenous
growth factors had been described in the regeneration thereof (25). Our study evidences a reduction
of the conjunctival cicatrization time in Type III
burns with platelets vis-à-vis the conventional treatment (group A) which is not statistically significant
(0.017). These results should be verified in larger
trials.
In relation to keratitis, a shorter healing period
was seen in group B patients (6 days) in comparison
to group A (9 days) which, however, was not statistically significant.
Fig. 2: Bulbar conjunctiva 24 hrs after subconjunctival
infiltration of plasmatic platelet concentrate.
vated small orange-colored subconjunctival
deposits in the infiltration points, which disappeared after 8 days (average). No permanent
sequels were found in any of the treated eyes.
DISCUSSION
The treatments for correcting corneal epithelializing defects include the application of autologous
serum, in neuroparalytic, neurotrophic keratitis
(30), chronic eye surface diseases (31), keratoconjunctivitis sicca (32,33) and persistent defects of the
corneal epithelium (34,35).
Autologous serum seems to facilitate the epithelializing healing and regeneration of eye surface
pathologies (19-22) because it provides growth factors and improves corneal sensitivity after application (30). In addition, the topical use of other
growth factors such as the epidermic growth factor
Table I. Summary of results
Days off
Type III
Healing time
Type III
Corneal healing
Type III
Conjunctival healing
Type III
Group A
Group B
M
5
5
P50
7
6
[P25-P75]
[6-9]
[4-6]
P
0.053
Group A
Group B
5
5
10
6
[8-15]
[6-7]
0.017
Group A
Group B
5
5
6
3
[4-7]
[2-3]
0.001
Group A
Group B
5
5
6
4
[6-7]
[4-6]
0.017
ARCH SOC ESP OFTALMOL 2007; 82: 475-482
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MÁRQUEZ-DE-ARACENA R, et al.
Even though it is not a strictly clinical factor, the
time off work of patients seemed to be a convenient
value to be included due to its social and economic
relevance. The results are very similar (7 days in
group A and 6 days in group B). As for the final
healing time of the burns, differences were found
between group A (10 days) and group B (6 days)
which were not statistically significant (0.017).
The shorter corneal cicatrization period (statistically significant), of the conjunctiva, the time off
work and the final healing time of the patients treated with plasmatic platelet concentration (group B)
as compared to the group treated with conventional
procedures (group A) could indicate that the subconjunctival application thereof plays an active and
efficient function in the external ocular regeneration
processes of traumatic etiology (burns). Therefore,
the topical utilization thereof could be considered.
The patients treated with autologous platelet concentrates (group B) did not evidence any correlation
between the different analytical results (hematic
and biochemical) vis-à-vis the parameters of the
study.
The efficacy of this treatment is evidenced in the
generally shorter healing and cicatrization time of
the conjunctiva and healing time of Type III ocular
burns (35), which becomes statistically significant
in corneal cicatrization.
The presence of small localized conjunctival elevations seems to indicate the location of platelet
concentrate deposits which could be released later
on, prolonging the effect of said treatment.
The subconjunctival application of plasma rich in
plasmatic platelets seems to accelerate the healing
of clinical signs existing in ocular pathologies such
as corneal and conjunctival ulcers. In processes
appearing several days after said application, such
as keratitis, it does not seem to be particularly efficient. Accordingly, the topical use thereof could be
considered.
In our experience, the clinical efficacy of subconjunctival application of platelet fraction allows
us to deduce that it is not necessary to activate the
platelet concentrate for it to be efficient and that the
possibility of a new application for treating ocular
burns could be of interest. In addition, the tissular
regeneration action of blood can be found mainly in
the platelet fraction.
The technique utilized in our study allows for the
exclusive obtainment of platelets, excluding inflammatory interleukines of leucocytic origin. Disease
480
transmission risks are avoided because the plasma
is autologous and in addition it is prepared immediately on an outpatient basis, as well as having a null
antigenic effect and a very low cost. The fact that
the platelet concentrate evidences regenerating
activity in the ocular surface opens a new range of
applications for treating ocular surface pathologies.
The subconjunctival application of autologous
plasmatic platelets (platelet self-therapy) in traumatic eye lesions – burns – is a simple and economic treatment for ocular surface burns, free of
undesirable side effects. However, the efficiency
thereof must be confirmed with larger series.
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