Trial Information

Vascular Endothelial Growth Factor Levels in Aqueous, Vitreous and Subretinal Fluid in Patients With Retinopathy of Prematurity Stage V.

Vascular Endothelial Growth Factor (VEGF) is a dimeric glycoprotein, naturally expressed in
epithelial and tumor cells (1). In conditions of hypoxia it is secreted by the pericytes,
the Retina Pigmented epithelium (RPE)and glial cells, including a the Müller cells (1,2,4,5)
favoring the formation of new abnormal vessels (1). Its activity is mediated by two high
affinity receptors: the receptor VEGF 1 (FLT-1) and VEGF 2 (FLT-2) (1,9). The FLT-1
promotes cellular proliferation whereas FLT-2 the migration and cell- extracellular matrix
interaction (9). The VEGF is essential as much for the normal retinal vascularization as for
pathological (5,14).

Normal Retinal vascularization has two phases: In the first fase, cells of mesenquimatous
origin form the first superficial plexus (14-21 weeks of gestation). In the second phase,
denominated "angiogenesis phase", the superficial and deep capillary plexus are formed
(15,20). The second phase is highly dependent of VEGF induced by hypoxia (15,20). In the
normal retina the growth occurs from the optical nerve to the periphery (17).

The scientific progress in the neonatal intensive care units in the last years has allowed
the survival of a larger number of premature newborns(15).

The Retinopathy of Prematurity (ROP)(ROP) was described for the first time in 1942 (4), at
the present moment it is a public health problem in the developing countries. Of each
100.000 blind child in Latin America, 24.000 are due to ROP (240/1000 blind child) (17). It
is the main cause of blindness in premature new born (1,4,5). It is of multifactorial
etiology (17) but it has been related to a short gestational age (smaller to 32 weeks of
gestation) (4), to low birth weight (less of 1500gr) or very low birth weight (less
1250gr)(1,4,5,17,18,20), to the administration of high oxygen concentration immediately
after birth (1,3,18), low levels of the Insulin-like growth factor(IGF-I) and excessive
production of VEGF (3,17).

All this, combined with large periods of hyperoxia-hypoxia, Produces that the normal
vascularization of the retina stops. This also produces vascular-occlusive changes,
arteriovenous anastomoses in the edge of the vascularized with the none-vascularized retina
(18), begins the formation of abnormal vessels towards the vitreous, finally lead to retina
detachment in the most severe cases (2,3,4,5).

The International Classification of ROP classifies it in 5 stages, dividing it in 3
anatomical zones. A threshold stage is considered when we have the presence of stage 3, in
zone 1 or 2 in 5 continuous hour uses or 8 discontinuous with plus disease (15,16,17). The
pre-threshold stage is divided in two types: type 1 is any stage of the disease in zone 1
with plus disease, stage 3, without plus disease in zone 1 or stage 2 or 3 with plus in zone
2. Type 2 is stage 1 or 2 without plus disease in zone 1 or stage 3 without plus disease in
zone 2 (17,19). The threshold stage happens approximately in 5% of the patients with very
low weight to the birth, of which the 10-15% develop blindness (6,8)

It is a public health problem that continuous without having an effective prophylaxis (5),
nevertheless the use of constant oxygen concentrations, avoiding the periods of
hyperoxia-hypoxia, has demonstrated to produce a decrease of the presentation of advanced
stages of the disease and the necessity of ophthalmological surgery (5). The early diagnosis
and treatment in thresholds stages have changed the prognosis of this disease (11,12). The
use of the laser as treatment in the threshold and prethreshold stages is successful in the
91-95% of the cases(10), the ablative effect on the neurons, probably diminishes the VEGF
production (15). The use of the vitrectomy in the early cases of retinal detachment also has
good results (13).

For the determination and the quantification of VEGF several laboratory techniques have been
used, as they are the Westernblot (1), the flow cytometry (7), CBA (7). Nevertheless, the
Enzyme-linked immunoabsorbent assay (ELISA) remains the gold standard for the measurement of
interleukins, growth factors and other cytokines in corporal fluids, including the vitreous,
aqueous and subretinal fluid (1,5,7).

It has been demonstrated in humans that there are high vitreous concentrations of VEGF in
eyes with active neovascularization (6,14). In the same way, the animal models of ROP report
high levels of VEGF (2,21). The levels of this growth factor are altered in both stages,
with a more marked increase during the first stage (2). Some clinical studies in patients
with ROP in stage 4 and 5 have reported an increase of up to 90 times greater of the
concentration of VEGF in the subretinal liquid in comparison with the found in patients with
retinal detachment of any other cause (1). Finally, when it has been possible to examine
retinas immediately after the treatment (due to death of the patient) have been found a low
expression of mRNA of VEGF in the treated cells (6).