Abstract
Purpose: :
Bevacizumab (BVZ), an anti-vascular endothelial growth factor (VEGF) agent has been used to treat several vasoproliferative disorders in adult eyes, and more recently, in developing eyes of neonates with Retinopathy of Prematurity. Extracellular matrix and cellular membrane proteoglycans (PG) are essentials for retinal cell development as part of control mechanisms of neuron differentiation, neurite and axon outgrowth, neural cell adhesion and apoptosis. Chondroitin (CSPG) and Heparan Sulfated Proteoglycans (HSPG) are the most abundant PG in the developing nervous central system as retina. Neurocan, a chondroitin sulfated PG, is an extracellular matrix molecule that can inhibit neurite and axon outgrowth. On the other hand, syndencan-3, a cellular membrane HSPG has the opposite function: stimulate axonal and neurite growth by binding with basis Fibroblast Growth Factor. The purpose of this research is to investigate the effects of BVZ on extracellular matrix and/or cellular membrane PGs during rat retina development.
Methods: :
Thirty 2-days-old Lister Hooded rats were euthanized and had their retinas dissected and cut in 1mm² explants. Retinal tissue was divided in two groups: control and treated (experimental group) with 0.5 mg/mL of BVZ for 48 hours. After incubation, retinas were analyzed with Real Time RT-PCR for mRNA evaluation for: neurocan and syndecan-3, using SYBR Green. The mRNA expression analysis was done with relative comparison method (ΔΔCt) and data were analyzed by Rotor Gene ScreenClust HRM (Qiagen) software. The amount of neurocan and syndecan-3 mRNA was calculated relative to the amount of ribosomal protein ARBP as reference gene. Comparisons between the groups were performed using the Student’s unpaired, considering a p value less than 0.05 as statistically significant.
Results: :
According to mRNA expression analysis, similar patterns were observed for the controls and experimental group for neurocan mRNA content. However, a significant decrease in syndecan-3 mRNA level, about 6.5 times lower, was observed in the BVZ-treated group compared to controls.
Conclusions: :
Our results demonstrated that BVZ could alter PG gene expression of syndecan-3, therefore could affect retinal cells differentiation during development, since syndecan-3 is important in neurite and axon outgrowth for adequate formation of neural networks in retina. However, more studies are necessary to investigate BVZ action, and suggest caution of its use, particularly on developing retina, as currently used in children with retinopathy of prematurity.
Keywords: retinal development • retinopathy of prematurity • vascular endothelial growth factor